Sarah Russell

A Scribble-E-cadherin complex controls daughter cell patterning by multiple mechanisms

Publisher: Cold Spring Harbor Laboratory

Date: 16-04-2021

DOI: 10.1101/2021.04.15.440081

Abstract: The fate of the two daughter cells is intimately connected to their positioning, which is in turn regulated by cell junction remodelling and orientation of the mitotic spindle. How multiple cues are integrated to dictate the ultimate patterning of daughters is not clear. Here, we identify novel mechanisms of regulation of daughter positioning in single MCF10A cells. The polarity protein, Scribble, links E-cadherin to NuMA and Arp2/3 signalling for sequential roles in daughter positioning. First Scribble transmits cues from E-cadherin localised in retraction fibres to control orientation of the mitotic spindle. Second, Scribble re-locates to the junction between the two daughters to allow a new E-cadherin-based-interface to form between them, influencing the width of the nascent daughter-daughter junction, generation of filopodia and subsequent cell patterning. Thus, E-cadherin and Scribble dynamically relocate to different intracellular sites during cell ision to orient the mitotic spindle and control placement of the daughter cells after cell ision.

A method for prolonged imaging of motile lymphocytes

Publisher: Wiley

Date: 04-11-2008

DOI: 10.1038/ICB.2008.79

Abstract: With new imaging technologies and fluorescent probes, live imaging of cells in vitro has revolutionized many aspects of cell biology. A key goal now is to develop systems to optimize in vitro imaging, which do not compromise the physiological relevance of the study. We have developed a methodology that contains non-adherent cells within the field of view. 'Cell paddocks' are created by generating an array of microgrids using polydimethylsiloxane. Each microgrid is up to 250 x 250 microm(2) with a height of 60 microm. Overlayed cells settle into the grids and the walls restrict their lateral movement, but a contiguous supply of medium between neighboring microgrids facilitates the exchange of cytokines and growth factors. This allows culture over at least 6 days with no impact upon viability and proliferation. Adaptations of the microgrids have enabled imaging and tracking of lymphocyte ision through multiple generations of long-term interactions between T lymphocytes and dendritic cells, and of thymocyte-stromal cell interactions.

Interplay of Polarity Proteins and GTPases in T-Lymphocyte Function

Publisher: Hindawi Limited

Date: 2012

DOI: 10.1155/2012/417485

Abstract: Polarity refers to the asymmetric distribution of different cellular components within a cell and is central to many cell functions. In T-cells, polarity regulates the activation, migration, and effector function of cytotoxic T-cells (CTLs) during an immune response. The regulation of asymmetric cell ision by polarity proteins may also dictate CTL effector and memory differentiation following antigen presentation. Small GTPases, along with their associated polarity and adaptor proteins, are critical for mediating the polarity changes necessary for T-cell activation and function, and in turn, are regulated by guanine exchange factors (GEFS) and GTPase activating proteins (GAPS). For ex le, a novel GEF, dedicator of cytokinesis 8 (DOCK8) was recently identified as a regulator of immune cell function and mutations in DOCK8 have been detected in patients with severe combined immunodeficiency. Both B and T-cells from DOCK8 mutant mice form defective immunological synapses and have abnormal functions, in addition to impaired immune memory development. This paper will discuss the interplay between polarity proteins and GTPases, and their role in T-cell function.

Delineation of the Regions of Interleukin-2 (IL-2) Receptor β Chain Important for Association of Jak1 and Jak3

Publisher: Elsevier BV

Date: 04-1998

DOI: 10.1074/JBC.273.17.10719

Maps of variability in cell lineage trees

Publisher: Cold Spring Harbor Laboratory

Date: 18-02-2018

DOI: 10.1101/267450

Abstract: New approaches to lineage tracking allow the study of cell differentiation over many generations of cells during development in multicellular organisms. Understanding the variability observed in these lineage trees requires new statistical methods. Whereas invariant cell lineages, such as that for the nematode Caenorhabditis elegans , can be described using a lineage map, defined as the fixed pattern of phenotypes overlaid onto the binary tree structure, the variability of cell lineages from higher organisms makes it impossible to draw a single lineage map. Here, we introduce lineage variability maps which describe the pattern of second-order variation throughout the lineage tree. These maps can be undirected graphs of the partial correlations between every lineal position or directed graphs showing the dynamics of bifurcated patterns in each subtree. By using the symmetry invariance of a binary tree to develop a generalized spectral analysis for cell lineages, we show how to infer these graphical models for lineages of any depth from s le sizes of only a few pedigrees. When tested on pedigrees from C. elegans expressing a marker for pharyngeal differentiation potential, the maps recover essential features of the known lineage map. When applied to highly-variable pedigrees monitoring cell size in T lymphocytes, the maps show how most of the phenotype is set by the founder naive T cell. Lineage variability maps thus elevate the concept of the lineage map to the population level, addressing questions about the potency and dynamics of cell lineages and providing a way to quantify the progressive restriction of cell fate with increasing depth in the tree. Multicellular organisms develop from a single fertilized egg by sequential cell isions. The progeny from these isions adopt different traits that are transmitted and modified through many generations. By tracking how cell traits change with each successive cell ision throughout the family, or lineage, tree, it has been possible to understand where and how these modifications are controlled at the single-cell level, thereby addressing questions about, for ex le, the developmental origin of tissues, the sources of differentiation in immune cells, or the relationship between primary tumors and metastases. Such lineages often show large variability, with apparently identical founder cells giving rise to different patterns of descendants. Fundamental scientific questions, such as about the range of possible cell types a cell can give rise to, are often about this variability. To characterize this variation, and thus understand the lineage at the population level, we introduce lineage variability maps. Using data from worm and mammalian cell lineages we show how these maps provide quantifiable answers to questions about any developing lineage, such as the potency of founder cells and the progressive restriction of cell fate at each stage in the tree.

T cell protein tyrosine phosphatase attenuates T cell signaling to maintain tolerance in mice

Publisher: American Society for Clinical Investigation

Date: 12-2011

DOI: 10.1172/JCI59492

E-cadherin in developing murine T cells controls spindle alignment and progression through β-selection

Publisher: American Association for the Advancement of Science (AAAS)

Date: 20-01-2023

DOI: 10.1126/SCIADV.ADE5348

Abstract: A critical stage of T cell development is β-selection at this stage, the T cell receptor β (TCRβ) chain is generated, and the developing T cell starts to acquire antigenic specificity. Progression through β-selection is assisted by low-affinity interactions between the nascent TCRβ chain and peptide presented on stromal major histocompatibility complex and cues provided by the niche. In this study, we identify a cue within the developing T cell niche that is critical for T cell development. E-cadherin mediates cell-cell interactions and influences cell fate in many developmental systems. In developing T cells, E-cadherin contributed to the formation of an immunological synapse and the alignment of the mitotic spindle with the polarity axis during ision, which facilitated subsequent T cell development. Collectively, these data suggest that E-cadherin facilitates interactions with the thymic niche to coordinate the β-selection stage of T cell development.

Polarization of excitation light influences molecule counting in single-molecule localization microscopy

Publisher: Springer Science and Business Media LLC

Date: 03-09-2014

DOI: 10.1007/S00418-014-1267-1

Abstract: Single-molecule localization microscopy has been widely applied to count the number of biological molecules within a certain structure. The percentage of molecules that are detected significantly affects the interpretation of data. Among many factors that affect this percentage, the polarization state of the excitation light is often neglected or at least unstated in publications. We demonstrate by simulation and experiment that the number of molecules detected can be different from -40 up to 100% when using circularly or linearly polarized excitation light. This is determined mainly by the number of photons emitted by single fluorescent molecule, namely the choice of fluorescence proteins, and the background noise in the system, namely the illumination scheme. This difference can be further exaggerated or mitigated by various fixation methods, magnification, and camera settings We conclude that the final choice between circularly or linearly polarized excitation light should be made experimentally, based on the signal to noise ratio of the system.

Cutting Edge: DNAX Accessory Molecule 1–Deficient CD8+ T Cells Display Immunological Synapse Defects That Impair Antitumor Immunity

Publisher: The American Association of Immunologists

Date: 15-01-2014

DOI: 10.4049/JIMMUNOL.1302197

Abstract: DNAX accessory molecule 1 (DNAM-1) is expressed on all CD8+ T cells and promotes their activation and effector function. DNAM-1 interacts with LFA-1, a critical molecule for immunological synapse formation between T cells and APCs, and for cytotoxic killing of target cells. Mice that lack DNAM-1 display abnormal T cell responses and antitumor activity however, the mechanism involved is unclear. In this article, we show that DNAM-1 deficiency results in reduced proliferation of CD8+ T cells after Ag presentation and impaired cytotoxic activity. We also demonstrate that DNAM-1–deficient T cells show reduced conjugations with tumor cells and decreased recruitment of both LFA-1 and lipid rafts to the immunological synapse, which correlates with reduced tumor cell killing in vitro. This synapse defect may explain why DNAM-1–deficient mice cannot clear tumors in vivo, and highlights the importance of DNAM-1 and the immunological synapse in T cell–mediated antitumor immunity.

Developing T cells form an immunological synapse for passage through the β-selection checkpoint

Publisher: Rockefeller University Press

Date: 19-01-2021

DOI: 10.1083/JCB.201908108

Abstract: The β-selection checkpoint of T cell development tests whether the cell has recombined its genomic DNA to produce a functional T cell receptor β (TCRβ). Passage through the β-selection checkpoint requires the nascent TCRβ protein to mediate signaling through a pre-TCR complex. In this study, we show that developing T cells at the β-selection checkpoint establish an immunological synapse in in vitro and in situ, resembling that of the mature T cell. The immunological synapse is dependent on two key signaling pathways known to be critical for the transition beyond the β-selection checkpoint, Notch and CXCR4 signaling. In vitro and in situ analyses indicate that the immunological synapse promotes passage through the β-selection checkpoint. Collectively, these data indicate that developing T cells regulate pre-TCR signaling through the formation of an immunological synapse. This signaling platform integrates cues from Notch, CXCR4, and MHC on the thymic stromal cell to allow transition beyond the β-selection checkpoint.

Dense small molecule labeling enables activator-dependent STORM by proximity mapping

Publisher: Springer Science and Business Media LLC

Date: 31-05-2016

DOI: 10.1007/S00418-016-1451-6

Abstract: Stochastic optical reconstruction microscopy (STORM) enables high-resolution imaging, but multi-channel 3D imaging is problematic because of chromatic aberrations and alignment errors. The use of activator-dependent STORM in which spectrally distinct activators can be coupled with a single reporter can circumvent such issues. However, the standard approach of linking activators and reporters to a single antibody molecule is h ered by low labeling density and the large size of the antibody. We proposed that small molecule labels might enable activator-dependent STORM if the reporter or activator were linked to separate small molecules that bound within 3.5 nm of each other. This would greatly increase the labeling density and therefore improve resolution. We tested various mixtures of phalloidin- or mCling-conjugated fluorophore to demonstrate this feasibility. The specific activation was dependent on the choice of activator, its density, a matching activating laser and its power. In addition to providing an effective means of multi-channel 3D STORM imaging, this method also provides information about the local proximity between labels, potentially enabling super-resolved mapping of the conformation of the labeled structures.

Different interleukin 2 receptor beta-chain tyrosines couple to at least two signaling pathways and synergistically mediate interleukin 2-induced proliferation.

Publisher: Proceedings of the National Academy of Sciences

Date: 05-03-1996

DOI: 10.1073/PNAS.93.5.2077

Abstract: One of the earliest events induced by interleukin 2 (IL-2) is tyrosine phosphorylation of cellular proteins, including the IL-2 receptor beta chain (IL-2Rbeta). Simultaneous mutation of three tyrosines (Y338, Y392, and Y510) in the IL-2Rbeta cytoplasmic domain abrogated IL-2-induced proliferation, whereas mutation of only Y338 or of Y392 and Y510 inhibited proliferation only partially. While Y392 and Y510 were critical for IL-2-induced activation of signal transducers and activators of transcription (STAT proteins), Y338 was required for Shc-IL-2Rbeta association and for IL-2-induced tyrosine phosphorylation of Shc. Thus, activation of both Jak-STAT and Shc-coupled signaling pathways requires specific IL-2Rbeta tyrosines that together act in concert to mediate maximal proliferation. In COS-7 cells, overexpression of Jak1 augmented phosphorylation of Y338 as well as Y392 and Y510, suggesting that the role for this Jak kinase may extend beyond the Jak-STAT pathway.

Structural basis of the zinc-induced cytoplasmic aggregation of the RNA-binding protein SFPQ

Publisher: Oxford University Press (OUP)

Date: 08-02-2020

DOI: 10.1093/NAR/GKAA076

Abstract: SFPQ is a ubiquitous nuclear RNA-binding protein implicated in many aspects of RNA biogenesis. Importantly, nuclear depletion and cytoplasmic accumulation of SFPQ has been linked to neuropathological conditions such as Alzheimer's disease (AD) and amyotrophic lateral sclerosis (ALS). Here, we describe a molecular mechanism by which SFPQ is mislocalized to the cytoplasm. We report an unexpected discovery of the infinite polymerization of SFPQ that is induced by zinc binding to the protein. The crystal structure of human SFPQ in complex with zinc at 1.94 Å resolution reveals intermolecular interactions between SFPQ molecules that are mediated by zinc. As anticipated from the crystal structure, the application of zinc to primary cortical neurons induced the cytoplasmic accumulation and aggregation of SFPQ. Mutagenesis of the three zinc-coordinating histidine residues resulted in a significant reduction in the zinc-binding affinity of SFPQ in solution and the zinc-induced cytoplasmic aggregation of SFPQ in cultured neurons. Taken together, we propose that dysregulation of zinc availability and/or localization in neuronal cells may represent a mechanism for the imbalance in the nucleocytoplasmic distribution of SFPQ, which is an emerging hallmark of neurodegenerative diseases including AD and ALS.

Defective lymphoid development in mice lacking expression of the common cytokine receptor γ chain

Publisher: Elsevier BV

Date: 03-1995

DOI: 10.1016/1074-7613(95)90047-0

Abstract: The common gamma chain (gamma c) of the IL-2, IL-4, IL-7, IL-9, and IL-15 receptors is defective in humans with XSCID. Mice lacking gamma c expression had hypoplastic thymuses the thymocytes responded to gamma c-independent mitogens, but not gamma c-dependent stimuli. Splenic T cells were diminished at 3 weeks of age, but CD4+ T cells markedly increased by 4 weeks. B cells were greatly diminished in contrast with the situation in XSCID. NK cells, gamma delta intestinal intraepithelial lymphocytes, dendritic epidermal T cells, peripheral lymph nodes, and gut-associated lymphoid tissue were absent. These findings underscore the importance of gamma c in lymphoid development. Moreover, differences in humans and mice lacking gamma c expression indicate species-specific differences in the roles of gamma c-dependent cytokines or in the existence of redundant pathways. These mice provide an important model for studying the pathophysiology provide an important model for studying the pathophysiology of and gene therapy for human XSCID.

An integrated transcriptional switch at the β-selection checkpoint determines T cell survival, development and leukaemogenesis

Publisher: Portland Press Ltd.

Date: 27-06-2019

DOI: 10.1042/BST20180414

Abstract: In T cell development, a pivotal decision-making stage, termed β-selection, integrates a TCRβ checkpoint to coordinate survival, proliferation and differentiation to an αβ T cell. Here, we review how transcriptional regulation coordinates fate determination in early T cell development to enable β-selection. Errors in this transcription control can trigger T cell acute lymphoblastic leukaemia. We describe how the β-selection checkpoint goes awry in leukaemic transformation.

The Scribble–Dlg–Lgl Module in Cell Polarity Regulation

Publisher: Springer International Publishing

Date: 2015

DOI: 10.1007/978-3-319-14463-4_4

Lethal giant larvae‐1 deficiency enhances the CD8⁺ effector T‐cell response to antigen challenge in vivo

Publisher: Wiley

Date: 22-09-2015

DOI: 10.1038/ICB.2015.82

Abstract: Lethal giant larvae-1 (Lgl-1) is an evolutionary conserved protein that regulates cell polarity in erse lineages however, the role of Lgl-1 in the polarity and function of immune cells remains to be elucidated. To assess the role of Lgl-1 in T cells, we generated chimeric mice with a hematopoietic system deficient for Lgl-1. Lgl-1 deficiency did not impair the activation or function of peripheral CD8(+) T cells in response to antigen presentation in vitro, but did skew effector and memory T-cell differentiation. When challenged with antigen-expressing virus or tumor, Lgl-1-deficient mice displayed altered T-cell responses. This manifested in a stronger antiviral and antitumor effector CD8(+) T-cell response, the latter resulting in enhanced control of MC38-OVA tumors. These results reveal a novel role for Lgl-1 in the regulation of virus-specific T-cell responses and antitumor immunity.

Different membrane cofactor protein (CD46) isoforms protect transfected cells against antibody and complement mediated lysis

Publisher: Elsevier BV

Date: 06-1993

DOI: 10.1016/0966-3274(93)90002-P

Abstract: The need for organ transplantation, especially of kidneys, exceeds the availability of human donors and the possibility of xenotransplantation from suitable animals is now being addressed. The immediate barrier to success is hyperacute graft rejection, resulting from naturally occurring xenoreactive antibodies and the activation of complement. It is proposed that the intensity of the hyperacute response can be reduced by providing additional regulatory molecules to limit activation of the complement cascade, initially as transfected gene products in cultured cells as an in vitro model and eventually as a transgene in potential donor animals, such as pigs. Limiting the activity of C3b reduces the production of the C3a, C4a and C5a anaphylotoxins, thus curtailing not only the immediate C3b-mediated lytic pathway but also the later effects of a cellular inflammatory response including endothelial and platelet cell activation. To develop and assess the first part of this strategy, we have transfected several cDNA's encoding isoforms of CD46 (membrane cofactor protein). At least four different CD46 isoforms are commonly expressed in almost all human cells, and we have compared two of these and a third form to determine if they mediate different functions. After transfection, CD46-expressing CHO-K1 cells were selected with methionine sulphoximine and identified using monoclonal antibodies. Transfectants with suitable CD46 expression were assayed for primary CD46 function using a lysis assay dependent on the reaction of antibody and complement. In this in vitro model of hyperacute rejection, normal human sera containing natural xenoreactive antibodies were shown to lyse CHO cells, but only in the presence of complement.(ABSTRACT TRUNCATED AT 250 WORDS)

Lethal Giant Larvae 1 Tumour Suppressor Activity Is Not Conserved in Models of Mammalian T and B Cell Leukaemia

Publisher: Public Library of Science (PLoS)

Date: 27-01-2014

DOI: 10.1371/JOURNAL.PONE.0087376

The Scribble and Par complexes in polarity and migration: friends or foes?

Publisher: Elsevier BV

Date: 12-2006

DOI: 10.1016/J.TCB.2006.10.005

Abstract: The Par complex [consisting of Bazooka (also called Par3), Par6 and aPKC] is a well-described regulator of cell polarity whose role in many aspects of cell morphogenesis is under intense investigation. Recently, another set of proteins known as the Scribble complex (consisting of Scribble, Discs large and Lethal giant larvae) has also been shown to be important in polarity regulation in several settings. Here, we describe the current status of Scribble in polarity and review evidence from various model systems that indicates an essential but context-dependent role for the Scribble and Par complexes in directed cell migration. Based on the known interactions of Scribble and Par complexes with each other and with other signalling pathways, we propose models by which Par and Scribble might interact to regulate cell migration.

Human astrocytes express membrane cofactor protein (CD46), a regulator of complement activation

Publisher: Elsevier BV

Date: 02-1992

DOI: 10.1016/0165-5728(92)90051-L

Abstract: Expression of membrane cofactor protein (CD46) on cultured human astrocytes was demonstrated by indirect immunofluorescence microscopy and flow cytometry following staining with a monoclonal antibody specific for CD46. Western transfer and immunoblotting detected a doublet of Mr 66,000 and 56,000. Analysis of astrocyte mRNA revealed the presence of multiple alternatively spliced transcripts encoding different extracellular regions or cytoplasmic tails of CD46. Astrocytes were also shown to express decay accelerating factor, but not the type 1 complement receptor. Upregulation of astrocyte CD46 occurred following cytomegalovirus infection. These results indicate that astrocytes express proteins involved in regulation of complement activation and protection against autologous complement.

Determination of T‐cell fate by dendritic cells: a new role for asymmetric cell division?

Publisher: Wiley

Date: 08-04-2008

DOI: 10.1038/ICB.2008.24

Abstract: The production, from a single naive T cell, of the many different activated T cell types required for an effective immune response has fascinated immunologists for decades. This process underpins the development of vaccines, immunosuppressive regimes in transplant patients, and immunotherapy in cancer among other things. Despite the enormous advances in detailing the mechanisms and influencing factors in the differentiation of each T-cell subtype, it is still not clear how the different T-cell progeny are produced in proportions that are appropriate for each situation. This review discusses the notion that asymmetric cell ision might allow for the regulated generation of different cell populations.

Divergent lymphocyte signalling revealed by a powerful new tool for analysis of time-lapse microscopy

Publisher: Wiley

Date: 02-10-2013

DOI: 10.1038/ICB.2012.49

Abstract: We describe a new approach for interactive analysis of time-lapse microscopy, and apply this approach to elucidating whether polarity regulation is conserved between epithelial cells and lymphocytes. A key advantage of our analysis platform, 'TACTICS', is the capacity to visualize in idual data points in the context of large data sets, similar to standard approaches in flow cytometry. Scatter plots representing microscopic parameters or their derivations such as polarity ratios are linked to the original data such that clicking on each dot enables a link to images and movies of the corresponding cell. Similar to flow cytometric analysis, subsets of the data can be gated and reanalyzed to explore the relationships between different parameters. TACTICS was used to dissect the regulation of polarization of the cell fate determinant, Numb, in migrating lymphocytes. We show here that residues of Numb that are phosphorylated by atypical protein kinase C (aPKC) to mediate apicobasal polarity in epithelial cells are not required for polarization of Numb in T cells, indicating that the role of aPKC is not conserved between lymphocytes and epithelia.

Alternatively spliced RNAs encode several isoforms of CD46 (MCP), a regulator of complement activation

Publisher: Springer Science and Business Media LLC

Date: 05-1991

DOI: 10.1007/BF00216692

Asymmetric Cell Division of T Cells upon Antigen Presentation Uses Multiple Conserved Mechanisms

Publisher: The American Association of Immunologists

Date: 07-2010

DOI: 10.4049/JIMMUNOL.0903627

Abstract: Asymmetric cell ision is a potential means by which cell fate choices during an immune response are orchestrated. Defining the molecular mechanisms that underlie asymmetric ision of T cells is paramount for determining the role of this process in the generation of effector and memory T cell subsets. In other cell types, asymmetric cell ision is regulated by conserved polarity protein complexes that control the localization of cell fate determinants and spindle orientation during ision. We have developed a tractable, in vitro model of naive CD8+ T cells undergoing initial ision while attached to dendritic cells during Ag presentation to investigate whether similar mechanisms might regulate asymmetric ision of T cells. Using this system, we show that direct interactions with APCs provide the cue for polarization of T cells. Interestingly, the immunological synapse disseminates before ision even though the T cells retain contact with the APC. The cue from the APC is translated into polarization of cell fate determinants via the polarity network of the Par3 and Scribble complexes, and orientation of the mitotic spindle during ision is orchestrated by the partner of inscuteable/G protein complex. These findings suggest that T cells have selectively adapted a number of evolutionarily conserved mechanisms to generate ersity through asymmetric cell ision.

A review of glass corrosion: the unique contribution of studying ancient glass to validate glass alteration models

Publisher: Springer Science and Business Media LLC

Date: 20-05-2023

DOI: 10.1038/S41529-023-00355-4

Abstract: Glass has been used in widespread applications within several sectors since ancient times and it has been systematically studied under different perspectives. However, its thermodynamic properties and the variety of its compositions, several aspects related to its durability and its alteration mechanisms remain still open to debate. This literature review presents an overview of the most relevant studies on glass corrosion and the interaction between glass and the environment. The review aims to achieve two objectives. On one hand, it aims to highlight how far research on glass corrosion has come by studying model systems created in the laboratory to simulate different alteration conditions and glass compositions. On the other, it seeks to point out what are the critical aspects that still need to be investigated and how the study of ancient, altered glass can add to the results obtained in laboratory models. The review intends also to demonstrate how advanced analytical techniques commonly used to study modern and technical glass can be applied to investigate corrosion marks on ancient s les.

High-speed and high-resolution 2D and 3D elemental imaging of corroded ancient glass by Laser Ablation-ICP-MS

Publisher: Royal Society of Chemistry (RSC)

Date: 2023

DOI: 10.1039/D2JA00337F

Abstract: Optimizing the LA-ICP-MS procedure to obtain 2D and 3D high-resolution multi-elemental imaging of heavily degraded Roman glass for studying glass weathering mechanisms by monitoring the lateral and in-depth distribution of elements.

Upsides and downsides to polarity and asymmetric cell division in leukemia

Publisher: Springer Science and Business Media LLC

Date: 24-11-2008

DOI: 10.1038/ONC.2008.350

Abstract: The notion that polarity regulators can act as tumor suppressors in epithelial cells is now well accepted. The function of these proteins in lymphocytes is less well explored, and their possible function as suppressors of leukemia has had little attention so far. We review the literature on lymphocyte polarity and the growing recognition that polarity proteins have an important function in lymphocyte function. We then describe molecular relationships between the polarity network and signaling pathways that have been implicated in leukemogenesis, which suggest mechanisms by which the polarity network might impact on leukemogenesis. We particularly focus on the possibility that disruption of polarity might alter asymmetric cell ision (ACD), and that this might be a leukemia-initiating event. We also explore the converse possibility that leukemic stem cells might be produced or maintained by ACD, and therefore that Dlg, Scribble and Lgl might be important regulators of this process.

Normalized Polarization Ratios for the Analysis of Cell Polarity

Publisher: Public Library of Science (PLoS)

Date: 25-06-2014

DOI: 10.1371/JOURNAL.PONE.0099885

Imaging of goblet cells as a marker for intestinal metaplasia of the stomach by one-photon and two-photon fluorescence endomicroscopy

Publisher: SPIE-Intl Soc Optical Eng

Date: 2009

DOI: 10.1117/1.3269681

Abstract: Goblet cells are a requirement for the diagnosis of intestinal metaplasia of the stomach. The gastric mucosa is lined by a monolayer of columnar epithelium with some specialization at the crypts, but there are no goblet cells in normal gastric epithelium. The appearance of goblet cells in gastric epithelium is an indicator of potential malignant progression toward adenocarcinoma. Therefore, in vivo three-dimensional imaging of goblet cells is essential for diagnoses of a premalignant stage of gastric cancers called intestinal metaplasia. We used mouse intestine, which has goblet cells, as a model of intestinal metaplasia. One-photon confocal fluorescence endomicroscopy and two-photon fluorescence endomicroscopy are employed for 3-D imaging of goblet cells. The penetration depth, the sectioning ability, and the photobleaching information of imaging are demonstrated. Both endomicroscopy techniques can three-dimensionally observe goblet cells in mouse large intestine and achieve an imaging depth of 176 microm. The two-photon fluorescence endomicroscopy shows higher resolution and contrast of the imaging sections at each depth. In addition, two-photon fluorescence endomicroscopy also has advantages of sectioning ability and less photobleaching. These results prove that two-photon fluorescence endomicroscopy is advantageous in diagnoses of a premalignant stage of gastric cancers.

Dlg, Scribble and Lgl in cell polarity, cell proliferation and cancer

Publisher: Wiley

Date: 19-05-2003

DOI: 10.1002/BIES.10286

Abstract: Dlg (Discs large), Scrib (Scribble) and Lgl (Lethal giant larvae) are evolutionarily conserved components of a common genetic pathway that link the seemingly disparate functions of cell polarity and cell proliferation in epithelial cells. dlg, scrib and lgl have been identified as tumour suppressor genes in Drosophila, mutations of which cause similar phenotypes, involving disruption of cell polarity and neoplastic overgrowth of tissues. The molecular mechanisms by which Dlg, Scrib and Lgl proteins regulate cell proliferation are not clear, but there is some evidence that epithelial polarisation is required for this regulation. Dlg, Scrib and Lgl are highly conserved between human and Drosophila, and we discuss evidence that these proteins also play a role in cancer progression in humans.

Calcium Signaling Is Required for Erythroid Enucleation

Publisher: Public Library of Science (PLoS)

Date: 05-01-2016

DOI: 10.1371/JOURNAL.PONE.0146201

Author Reply to Peer Reviews of Stepwise progression of β-selection during T cell development as revealed by histone deacetylation inhibition

Publisher: EMBO

Date: 16-07-2022

DOI: 10.15252/RC.2022541874

Stepwise progression of β-selection during T cell development as revealed by histone deacetylation inhibition

Publisher: Cold Spring Harbor Laboratory

Date: 04-10-2021

DOI: 10.1101/2021.10.03.462949

Abstract: During T cell development, the first step in creating a unique T Cell Receptor (TCR) is the genetic recombination of the TCRβ chain. The quality of the newly recombined TCRβ is assessed at the β-selection checkpoint. Most cells fail this checkpoint and die, but the coordination of the complex events that control fate at the β-selection checkpoint is not yet understood. We shed new light on fate determination during β-selection using a selective inhibitor of histone deacetylase 6, ACY1215. ACY1215 disrupted the β-selection checkpoint. Characterising the basis for this disruption revealed a new, pivotal stage in β-selection, bookended by upregulation of the TCR co-receptors, CD28 and CD2 respectively. Within this ‘DN3b Pre’ stage, CD5 and Lef1 are upregulated to reflect pre-TCR signalling and their expression correlates with proliferation. During this phase, ACY1215-mediated disruption of the organisation of the β-selection immunological synapse is associated with a breakdown in connectivity of expression of pre-TCR, CD5 and Lef1. Subsequent deregulation of pre-TCR-induced proliferation leads to bypass of the β-selection checkpoint and subsequent failure to progress. We propose that the progressive expression of CD28, CD5 and Lef1, then CD2 reports and modulates the pre-TCR signal to orchestrate passage through the β-selection checkpoint. These findings suggest a refined model of β-selection in which a coordinated increase in expression of pre-TCR, CD28, CD5 and Lef1 allows for modulating TCR signalling strength, and culminates in the expression of CD2 to enable exit from the β-selection checkpoint.

hScrib is a functional homologue of the Drosophila tumour suppressor Scribble

Publisher: Springer Science and Business Media LLC

Date: 18-12-2003

DOI: 10.1038/SJ.ONC.1207154

Quantifying subcellular distribution of fluorescent fusion proteins in cells migrating within tissues

Publisher: Wiley

Date: 19-10-2010

DOI: 10.1038/ICB.2010.122

Abstract: The movement of proteins within cells can provide dynamic indications of cell signaling and cell polarity, but methods are needed to track and quantify subcellular protein movement within tissue environments. Here we present a semiautomated approach to quantify subcellular protein location for hundreds of migrating cells within intact living tissue using retrovirally expressed fluorescent fusion proteins and time-lapse two-photon microscopy of intact thymic lobes. We have validated the method using GFP-PKCζ, a marker for cell polarity, and LAT-GFP, a marker for T-cell receptor signaling, and have related the asymmetric distribution of these proteins to the direction and speed of cell migration. These approaches could be readily adapted to other fluorescent fusion proteins, tissues and biological questions.

Asymmetric Proteasome Segregation as a Mechanism for Unequal Partitioning of the Transcription Factor T-bet during T Lymphocyte Division

Publisher: Elsevier BV

Date: 04-2011

DOI: 10.1016/J.IMMUNI.2011.03.017

DOCK8 deficiency impairs CD8 T cell survival and function in humans and mice

Publisher: Rockefeller University Press

Date: 17-10-2011

DOI: 10.1084/JEM.20110345

Abstract: In humans, DOCK8 immunodeficiency syndrome is characterized by severe cutaneous viral infections. Thus, CD8 T cell function may be compromised in the absence of DOCK8. In this study, by analyzing mutant mice and humans, we demonstrate a critical, intrinsic role for DOCK8 in peripheral CD8 T cell survival and function. DOCK8 mutation selectively diminished the abundance of circulating naive CD8 T cells in both species, and in DOCK8-deficient humans, most CD8 T cells displayed an exhausted CD45RA+CCR7− phenotype. Analyses in mice revealed the CD8 T cell abnormalities to be cell autonomous and primarily postthymic. DOCK8 mutant naive CD8 T cells had a shorter lifespan and, upon encounter with antigen on dendritic cells, exhibited poor LFA-1 synaptic polarization and a delay in the first cell ision. Although DOCK8 mutant T cells underwent near-normal primary clonal expansion after primary infection with recombinant influenza virus in vivo, they showed greatly reduced memory cell persistence and recall. These findings highlight a key role for DOCK8 in the survival and function of human and mouse CD8 T cells.

Tissue‐specific and allelic expression of the complement regulator CD46 is controlled by alternative splicing

Publisher: Wiley

Date: 06-1992

DOI: 10.1002/EJI.1830220625

Abstract: CD46 (membrane cofactor protein) is a human cell surface glycoprotein with cofactor activity for factor I-mediated cleavage of complement components C3b and C4b. The CD46 protein from normal lymphocytes resolves on sodium dodecyl sulfate-polyacrylamide gel electrophoresis as two major bands of 66 and 56 kDa. CD46 cDNA encodes four extracellular short consensus repeat domains, a Ser/Thr/Pro (STP)-rich region, a transmembrane region and a cytoplasmic tail. We now show that exquisite control of mRNA splicing is responsible for the heterogeneous expression of CD46 isoforms. Differential splicing of 5 exons generates at least 14 CD46 mRNA variants whose expression is stringently regulated by allelic, tissue-specific and malignancy-related factors, as: (a) leukemic cells and Epstein-Barr virus-transformed B cells preferentially incorporate the first of three STP exons (exon 7) into mRNA, and produce a larger CD46 isoform of 74 kDa, (b) an allelic difference in the proportion of 66- and 56-kDa CD46 isoforms on lymphocytes corresponds to the preferential inclusion or exclusion of the second STP exon (exon 8), (c) the third STP exon (exon 9) is specifically deleted in some placentae, (d) spermatozoa delete both exons 12 and 13, encoding a shorter transmembrane region and a unique cytoplasmic tail and (e) all tissues tested differentially splice exon 13, resulting in two alternative cytoplasmic tails. The distribution of the 14 alternatively spliced RNA transcripts correlated with the presence of protein isoforms of the predicted size, indicating that alternative splicing leads to heterogeneity of CD46 glycoproteins.

Compartmentalization in T‐cell signalling: Membrane microdomains and polarity orchestrate signalling and morphology

Publisher: Wiley

Date: 02-2006

DOI: 10.1111/J.1440-1711.2005.01415.X

Abstract: Lymphocyte function is regulated by complex signalling responses to erse extracellular inputs, and a cell will often receive multiple, conflicting signals at one time. The mechanisms by which a lymphocyte integrates these signals into a single cellular response are not well understood. An important factor in the integration of signals likely involves the regulation of access of signalling molecules to cell surface receptors and of receptor signals to morphological determinants within the cell. Recent studies have led to important advances in our understanding of both the mechanisms by which signals are compartmentalized in T cells and the physiological role played by such compartmentalization. We review progress in the field, with a particular focus on membrane microdomains or lipid rafts and on cell polarity.

Polymorphic expression of CD46 protein isoforms due to tissue-specific RNA splicing

Publisher: Elsevier BV

Date: 10-1993

DOI: 10.1016/0161-5890(93)90038-D

Abstract: CD46 is a member of the regulators of complement activation (RCA) family and serves to protect autologous cells from complement mediated lysis. The CD46 gene consists of 14 exons and extensive RNA splicing produces protein isoforms of different molecular weight. Predominant protein isoforms of 66 and 56 kDa arise from splicing in or out of exon 8 which encodes a region rich in serine, threonine and proline residues known to be heavily O-glycosylated. An inherited allelic polymorphism controls the relative expression of these isoforms in PBL and other tissues. This study has analysed an independent and overriding tissue specific regulation of CD46 splicing. Salivary gland and kidney produce RNA transcripts that preferentially include exon 8, giving rise to the 66 kDa protein species, while exon 8 is spliced out in brain tissue to give the 56 kDa protein. The cytoplasmic tail of CD46 is encoded by either exon 13 (CYT 1) or exon 14 (CYT 2). There is a preferential deletion of exon 13 from transcripts in salivary gland, kidney and brain to encode a protein containing cytoplasmic tail CYT 2. This preferential production of the CYT 2 tail is contrary to that seen on peripheral blood lymphocytes where equivalent expression of both CYT 1 and CYT 2 is observed. Our results suggest that while the splicing of exons within most cells is controlled by nucleotide sequences within or close to the CD46 gene (i.e. cis-regulation), splicing in tissues such as salivary gland, kidney and brain is regulated by trans-splicing factors encoded by another gene(s).

How polarity shapes the destiny of T cells

Publisher: The Company of Biologists

Date: 15-01-2008

DOI: 10.1242/JCS.021253

Abstract: The differentiation, activation and expansion of T cells are dictated by their integrated response to a complex array of extracellular signals. Recent studies provide insight into how these signals are integrated and demonstrate a key role for cell shape in many aspects of T-cell signalling. T cells polarise during migration, antigen presentation and cell ision to give rise to daughter cells that can have different cell fates. In each case, the polarity of the T cell facilitates this activity. This raises the possibility that adoption of a polarised state acts as a positive feedback mechanism to enhance responses to specific signals. Similarly, in asymmetric ision of other cell types, the distribution of different molecules into each daughter can have profound consequences for proliferation, death and differentiation. The mechanisms of polarity regulation are far better understood in cells such as epithelial cells, neurons and neuronal precursors, and the fertilised zygote. With the emerging parallels between polarity in these cells and T cells, we should now be able to elucidate how polarity affects signalling and cell fate determination in T cells.

Second harmonic generation imaging via nonlinear endomicroscopy

Publisher: Optica Publishing Group

Date: 11-01-2010

DOI: 10.1364/OE.18.001255

Polarized Cells, Polarized Views: Asymmetric Cell Division in Hematopoietic Cells

Publisher: Frontiers Media SA

Date: 2014

DOI: 10.3389/FIMMU.2014.00026

Heterodimerization of the IL-2 receptor β- and γ-chain cytoplasmic domains is required for signalling

Publisher: Springer Science and Business Media LLC

Date: 05-1994

DOI: 10.1038/369330A0

Abstract: The interaction of interleukin-2 (IL-2) and IL-2 receptors critically regulates the T-cell immune response following antigen activation. IL-2 can signal through high or intermediate affinity receptors which contain IL-2R alpha (refs 3, 4) +beta (refs 5-8) +gamma (ref. 9) or beta+gamma chains, respectively. IL-2R gamma is a common gamma chain, gamma c, also shared by the IL-7 (ref. 10) and IL-4 (refs 11, 12) receptors, which when mutated results in X-linked severe combined immunodeficiency. Using chimaeric receptor constructs together with monoclonal or bispecific antibodies we demonstrate here that IL-2 signalling requires ligand-induced extracellular-domain-mediated heterodimerization of the beta- and gamma c-chain cytoplasmic domains. Anti-IL-2R alpha monoclonal antibodies trigger proliferation of cells transfected with chimaeric constructs in which the extracellular domains of IL-2R beta and gamma c are replaced by that of IL-2R alpha. Other experiments using chimaeric constructs indicated that IL-2 binds monomerically and monovalently to IL-2R alpha and that the beta-transmembrane domain is not required for receptor chain interactions. Finally, we provide a method for mapping residues in the gamma c cytoplasmic domain even in cells that constitutively express gamma c.

Mediating signaling response to actin‐mediated forces: Cas‐L is causal in the T‐cell response to forces triggered by antigen presentation

Publisher: Wiley

Date: 20-09-2016

DOI: 10.1038/ICB.2016.79

Liquid Thermoelectrics: Review of Recent And Limited New Data of Thermogalvanic Cell Experiments

Publisher: Informa UK Limited

Date: 11-2013

DOI: 10.1080/15567265.2013.776149

Chitosan-coated amyloid fibrils increase adipogenesis of mesenchymal stem cells

Publisher: Elsevier BV

Date: 10-2017

DOI: 10.1016/J.MSEC.2017.05.050

Abstract: Mesenchymal stem cells (MSCs) have the potential to revolutionize medicine due to their ability to differentiate into specific lineages for targeted tissue repair. Development of materials and cell culture platforms that improve differentiation of either autologous or allogenic stem cell sources into specific lineages would enhance clinical utilization of MCSs. In this study, nanoscale amyloid fibrils were evaluated as substrate materials to encourage viability, proliferation, multipotency, and differentiation of MSCs. Fibrils assembled from the proteins lysozyme or β-lactoglobulin, with and without chitosan coatings, were deposited on planar mica surfaces. MSCs were cultured and differentiated on fibril-covered surfaces, as well as on unstructured controls and tissue culture plastic. Expression of CD44 and CD90 proteins indicated that multipotency was maintained for all fibrils, and osteogenic differentiation was similarly comparable among all tested materials. MSCs grown for 7days on fibril-covered surfaces favored multicellular spheroid formation and demonstrated a >75% increase in adipogenesis compared to tissue culture plastic controls, although this benefit could only be achieved if MSCs were transferred to TCP for the final differentiation step. The largest spheroids and greatest tendency to undergo adipogenesis was evidenced among MSCs grown on fibrils coated with the positively-charged polysaccharide chitosan, suggesting that spheroid formation is prompted by both topography and cell-surface interactivity and that there is a connection between multicellular spheroid formation and adipogenesis.

A Functional Interaction between CD46 and DLG4

Publisher: Elsevier BV

Date: 02-2002

DOI: 10.1074/JBC.M108479200

A Chemical Screening Approach to Identify Novel Key Mediators of Erythroid Enucleation

Publisher: Public Library of Science (PLoS)

Date: 16-11-2015

DOI: 10.1371/JOURNAL.PONE.0142655

Stepwise progression of β-selection during T cell development involves histone deacetylation

Publisher: Life Science Alliance, LLC

Date: 25-10-2022

DOI: 10.26508/LSA.202201645

Abstract: During T cell development, the first step in creating a unique T cell receptor (TCR) is genetic recombination of the TCRβ chain. The quality of the new TCRβ is assessed at the β-selection checkpoint. Most cells fail this checkpoint and die, but the coordination of fate at the β-selection checkpoint is not yet understood. We shed new light on fate determination during β-selection using a selective inhibitor of histone deacetylase 6, ACY1215. ACY1215 disrupted the β-selection checkpoint. Characterising the basis for this disruption revealed a new, pivotal stage in β-selection, bookended by up-regulation of TCR co-receptors, CD28 and CD2, respectively. Within this “DN3b Pre ” stage, CD5 and Lef1 are up-regulated to reflect pre-TCR signalling, and their expression correlates with proliferation. These findings suggest a refined model of β-selection in which a coordinated increase in expression of pre-TCR, CD28, CD5 and Lef1 allows for modulating TCR signalling strength and culminates in the expression of CD2 to enable exit from the β-selection checkpoint.

Scribble and E-cadherin cooperate to control symmetric daughter cell positioning by multiple mechanisms

Publisher: The Company of Biologists

Date: 15-01-2023

DOI: 10.1242/JCS.260547

Abstract: The fate of the two daughter cells is intimately connected to their positioning, which is in turn regulated by cell junction remodelling and orientation of the mitotic spindle. How multiple cues are integrated to dictate the ultimate positioning of daughters is not clear. Here, we identify novel mechanisms of regulation of daughter positioning in single MCF10A cells. The polarity protein, Scribble cooperates with E-cadherin for sequential roles in daughter positioning. First Scribble stabilises E-cadherin at the mitotic cortex as well as the retraction fibres, to mediate spindle orientation. Second, Scribble re-locates to the junction between the two daughters to allow a new E-cadherin-based-interface to form between them, influencing the width of the nascent daughter–daughter junction and subsequent cell positioning. Thus, E-cadherin and Scribble dynamically relocate to different intracellular sites during cell ision to orient the mitotic spindle and control placement of the daughter cells after cell ision. This article has an associated First Person interview with the first author of the paper.

Developing T cells form an immunological synapse for passage through the β−selection checkpoint

Publisher: Cold Spring Harbor Laboratory

Date: 12-08-2019

DOI: 10.1101/732511

Abstract: The β-selection checkpoint of T cell development tests whether the cell has recombined its genomic DNA to produce a functional T Cell Receptor β (TCRβ) receptor. Passage through the β-selection checkpoint requires the nascent TCRβ protein to mediate signaling through a pre-TCR complex. In this study, we show that developing T cells at the β-selection checkpoint establish an immunological synapse in in vitro & in situ, resembling that of the mature T cell. The immunological synapse is dependent on two key signaling pathways known to be critical for the transition beyond the β-selection checkpoint, Notch and CXCR4 signaling. In vitro and in situ analyses indicate that the immunological synapse promotes passage through the β-selection checkpoint. Collectively, these data indicate that developing T cells regulate pre-TCR signaling through the formation of an immunological synapse. This signaling platform integrates cues from Notch, CXCR4, and MHC on the thymic stromal cell, to allow transition beyond the β-selection checkpoint. T cell development requires testing whether genomic rearrangement has produced a T cell receptor capable of transmitting signals. Most T cells fail this test. Here, we show that passage through the β-selection checkpoint requires assembly of a platform to support TCR signaling.

Polarity and asymmetric cell division in the control of lymphocyte fate decisions and function

Publisher: Elsevier BV

Date: 04-2016

DOI: 10.1016/J.COI.2016.02.004

Abstract: Polarity is important in several lymphocyte processes including lymphocyte migration, formation of the immunological synapse, and asymmetric cell ision (ACD). While lymphocyte migration and immunological synapse formation are relatively well understood, the role of lymphocyte ACD is less clear. Recent advances in measuring polarity enable more robust analyses of asymmetric cell ision. Use of these new methods has produced crucial quantification of ACD at precise phases of lymphocyte development and activation. These developments are leading to a better understanding of the drivers of fate choice during lymphocyte activation and provide a context within which to explain the effects of ACD.

A Network of PDZ-Containing Proteins Regulates T Cell Polarity and Morphology during Migration and Immunological Synapse Formation

Publisher: Elsevier BV

Date: 06-2005

DOI: 10.1016/J.IMMUNI.2005.04.009

Abstract: T cell shape is dictated by the selective recruitment of molecules to different regions of the cell (polarity) and is integral to every aspect of T cell function, from migration to cytotoxicity. This study describes a mechanism for the regulation of T cell polarity. We show that T cells contain a network of asymmetrically distributed proteins with the capacity to dictate the subcellular localization of both cell surface receptors and morphological determinants in T cells. Proteins from the Scribble, Crumbs3, and Par3 complexes, previously shown to regulate epithelial polarity, were polarized in T cells containing either uropods or immunological synapses. Reduction in Scribble expression prevented the polarization of cell surface receptors and prevented morphological changes associated with uropod formation, migration, and antigen presentation. By dynamically coordinating molecular distribution throughout the T cell, this network provides a mechanism by which T cell function and polarity are linked.

Expression of interleukin‐2 receptor γ on human monocytes: characterization of lineage specific post‐translational modifications

Publisher: Wiley

Date: 1995

DOI: 10.1002/EJI.1830250148

Abstract: Functional interleukin-2 receptors (IL-2R) on lymphocytes contain both IL-2R beta and gamma chains. Whereas constitutive expression of IL-2R beta has been found on monocytes, the expression of IL-2R gamma on these phagocytes has not been examined. We performed reverse-transcription-polymerase chain reaction with Southern blot analysis on RNA derived from purified human monocytes and discovered that they constitutively produce IL-2R gamma mRNA. Western immunoblotting revealed 58- and 64-kDa forms of IL-2R gamma on YT-1 and human monocytes, whereas 58-, 64-, and 69-kDa bands were detected using peripheral blood mononuclear cells and non-adherent lymphocytes. These different forms resulted from variable N-linked glycosylation since culture of the cells in tunicamycin resulted in detection of a single 39-kDa band which corresponds to the molecular weight predicted from the deduced amino acid sequence. By co-immunoprecipitation, the IL-2R beta subunit associates with only the 64-kDa IL-2R gamma protein band in monocytes.

Ligation of the cell surface receptor, CD46, alters T cell polarity and response to antigen presentation

Publisher: Proceedings of the National Academy of Sciences

Date: 05-12-2006

DOI: 10.1073/PNAS.0602458103

Abstract: Lymphocyte function in vivo is dictated by multiple external cues, but the integration of different signals is not well understood. Here, we show that competition for the axis of polarization dictates functional outcomes. We investigated the effect of ligation of the immunoregulatory cell surface receptor, CD46, on lymphocyte polarity during antigen presentation and cytotoxic effector function. Ligation of CD46 on human T cells prevented recruitment of the microtubule organizing center, CD3, and perforin to the interface with the antigen-presenting cell and caused a reduction in IFN-γ production. In human NK cells, similar changes in polarity induced by CD46 ligation inhibited the recruitment of the microtubule organizing center and perforin to the interface with target cells and correlated with reduced killing. These data indicate that external signals can alter lymphocyte polarization toward antigen-presenting cells or target cells, inhibiting lymphocyte function.

E-cadherin in developing murine T cells controls spindle alignment and differentiation during β-selection

Publisher: Cold Spring Harbor Laboratory

Date: 15-06-2022

DOI: 10.1101/2022.06.14.496211

Abstract: A critical stage of T cell development is β-selection at this stage the TCRβ chain is generated and the developing T cell starts to acquire antigenic specificity. Progression through β-selection is assisted by a low affinity interaction between the nascent TCRβ chain and peptide presented on stromal MHC and external cues provided by the niche, including Notch and CXCR4. In this study, we reveal the importance of a new cue within the murine developing T cell niche which is critical for T cell development. E-cadherin mediates cell-cell interactions and influences cell fate in many developmental systems. In developing T cells E-cadherin contributed to the formation of an immunological synapse and the alignment of the mitotic spindle with the polarity axis during ision, which facilitated subsequent T cell development. Collectively, these data highlight a new aspect of the developing T cell niche and provide insights into the role of E-cadherin in the β-selection stage of T cell development.

Super‐resolution imaging and statistical analysis of CdSe/CdS Core/Shell semiconductor nanocrystals

Publisher: Wiley

Date: 17-06-2010

DOI: 10.1002/JBIO.201000032

Abstract: Here we present a multifunctional algorithm. Firstly a super-resolution method is presented for optically imaging the spatial distribution of semiconductor nanocrystals with nanometre localisation. Secondly highly resolved multiple photoluminescence trajectories of hundreds of single semiconductor nanocrystals are obtained simultaneously.

The common γ chain (γc) and its involvement in X-linked SCID

Publisher: Elsevier BV

Date: 06-1994

DOI: 10.1016/0197-1859(94)90049-3

The Reorientation of T-Cell Polarity and Inhibition of Immunological Synapse Formation by CD46 Involves Its Recruitment to Lipid Rafts

Publisher: Hindawi Limited

Date: 2011

DOI: 10.1155/2011/521863

Abstract: Many infectious agents utilize CD46 for infection of human cells, and therapeutic applications of CD46-binding viruses are now being explored. Besides mediating internalization to enable infection, binding to CD46 can directly alter immune function. In particular, ligation of CD46 by antibodies or by measles virus can prevent activation of T cells by altering T-cell polarity and consequently preventing the formation of an immunological synapse. Here, we define a mechanism by which CD46 reorients T-cell polarity to prevent T-cell receptor signaling in response to antigen presentation. We show that CD46 associates with lipid rafts upon ligation, and that this reduces recruitment of both lipid rafts and the microtubule organizing centre to the site of receptor cross-linking. These data combined indicate that polarization of T cells towards the site of CD46 ligation prevents formation of an immunological synapse, and this is associated with the ability of CD46 to recruit lipid rafts away from the site of TCR ligation.

Asymmetric T Lymphocyte Division in the Initiation of Adaptive Immune Responses

Publisher: American Association for the Advancement of Science (AAAS)

Date: 23-03-2007

DOI: 10.1126/SCIENCE.1139393

Abstract: A hallmark of mammalian immunity is the heterogeneity of cell fate that exists among pathogen-experienced lymphocytes. We show that a iding T lymphocyte initially responding to a microbe exhibits unequal partitioning of proteins that mediate signaling, cell fate specification, and asymmetric cell ision. Asymmetric segregation of determinants appears to be coordinated by prolonged interaction between the T cell and its antigen-presenting cell before ision. Additionally, the first two daughter T cells displayed phenotypic and functional indicators of being differentially fated toward effector and memory lineages. These results suggest a mechanism by which a single lymphocyte can apportion erse cell fates necessary for adaptive immunity.

TACTICS, an interactive platform for customized high-content bioimaging analysis

Publisher: Oxford University Press (OUP)

Date: 22-01-2013

DOI: 10.1093/BIOINFORMATICS/BTT035

Abstract: Summary: We describe a modular MATLAB® Toolbox named TACTICS for time-lapse image analysis that meets several requirements not generally offered by currently available software packages: (i) the ability to assess quality of extracted imaging information by directly linking data end points to the original position, (ii) massively parallel analysis of each parameter, for flow cytometry-like assessment of possible relationships between parameters within sub-populations of the images, (iii) options for user control of the tracking such as an interface to restrict the analysis region, (iv) manual correction of automated processes and (v) user interfaces for post-tracking analysis that is linked to the original images, including options to view cell pedigrees and normalized polarization ratios based on fluorescence ratiometric measurements. Availability and implementation: We provide TACTICS source code as well as video tutorials, data s le and comprehensive user guide on the TACTICS toolbox website (www.tactics-toolbox.com). Installation of TACTICS requires MATLAB 7.6 (R2008a) with the presence of Image Processing Toolbox™ and Statistics Toolbox™. Contact: sarah.russell@petermac.org.

Scribble acts as an oncogene in Eμ-myc-driven lymphoma

Publisher: Springer Science and Business Media LLC

Date: 18-05-2015

DOI: 10.1038/ONC.2015.167

Abstract: Scribble complex proteins maintain apicobasal polarity, regulate cell fate determination and function as tumour suppressors in epithelial tissue. Despite evidence that the function of Scribble is maintained in the lymphocyte lineage, we still understand little about its role as a tumour suppressor in haematological malignancies. Using the Eμ-myc model of Burkitt's lymphoma we investigated the role of Scribble in lymphomagenesis. We found that contrary to its well-documented tumour suppressor role in epithelial tissue, loss of Scribble expression delayed the expansion of peripheral B cells and delayed the onset of Eμ-myc-driven lymphoma. This was despite upregulated ERK phosphorylation levels in Scribble-deficient tumours, which are associated with loss of Scribble expression and the development of more aggressive Burkitt's lymphoma. Interestingly, the developmental stage of lymphoma was unaffected by Scribble expression challenging any role for Scribble in fate determination in the haematopoetic lineage. These data provide evidence for oncogenic properties of Scribble in Myc-driven B-cell lymphomagenesis, reinforcing recent findings that overexpression of a mutant form of Scribble can act as an oncogene in epithelial cells. Our results support the growing appreciation that the tumour regulatory functions of Scribble, and other polarity protein family members, are context dependent.

Asymmetric segregation and self-renewal of hematopoietic stem and progenitor cells with endocytic Ap2a2

Publisher: American Society of Hematology

Date: 15-03-2012

DOI: 10.1182/BLOOD-2011-11-393272

Abstract: The stem cell–intrinsic model of self-renewal via asymmetric cell ision (ACD) posits that fate determinants be partitioned unequally between daughter cells to either activate or suppress the stemness state. ACD is a purported mechanism by which hematopoietic stem cells (HSCs) self-renew, but definitive evidence for this cellular process remains open to conjecture. To address this issue, we chose 73 candidate genes that function within the cell polarity network to identify potential determinants that may concomitantly alter HSC fate while also exhibiting asymmetric segregation at cell ision. Initial gene-expression profiles of polarity candidates showed high and differential expression in both HSCs and leukemia stem cells. Altered HSC fate was assessed by our established in vitro to in vivo screen on a subcohort of candidate polarity genes, which revealed 6 novel positive regulators of HSC function: Ap2a2, Gpsm2, Tmod1, Kif3a, Racgap1, and Ccnb1. Interestingly, live-cell videomicroscopy of the endocytic protein AP2A2 shows instances of asymmetric segregation during HSC rogenitor cell cytokinesis. These results contribute further evidence that ACD is functional in HSC self-renewal, suggest a role for Ap2a2 in HSC activity, and provide a unique opportunity to prospectively analyze progeny from HSC asymmetric isions.

The tumour-suppressor Scribble dictates cell polarity during directed epithelial migration: regulation of Rho GTPase recruitment to the leading edge

Publisher: Springer Science and Business Media LLC

Date: 09-10-2007

DOI: 10.1038/SJ.ONC.1210016

Abstract: Altered expression of human Scribble is associated with invasive epithelial cancers, however, its role in tumour development remains unclear. Mutations in Drosophila Scribble result in loss of polarity, overproliferation and 3D-tumourous overgrowth of epithelial cells. Using complementation studies in Drosophila we recently demonstrated that expression of human Scribble can also regulate polarity and restrict tissue overgrowth. Here, we have undertaken a detailed study of human Scribble function in the polarized mammary cell line, MCF10A. We show that although Scribble does not seem to be required for apical-basal polarity or proliferation control in MCF10A cells, Scribble is essential for the control of polarity associated with directed epithelial cell migration. Scribble-depleted MCF10A cells show defective in vitro wound closure and chemotactic movement. The cells at the wound edge fail to polarize, show reduced lamellipodia formation and impaired recruitment of Cdc42 and Rac1 to the leading edge. Furthermore, we show that this function is relevant in vivo as Scribble mutant mice show defective epidermal wound healing. This data identifies an essential role for mammalian Scribble in the regulation of the polarity specifically involved in directed epithelial migration.

Maps of variability in cell lineage trees

Publisher: Public Library of Science (PLoS)

Date: 12-02-2019

DOI: 10.1371/JOURNAL.PCBI.1006745

University Of Melbourne

Location: Australia

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Swinburne University Of Technology

Location: Australia

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Peter MacCallum Cancer Centre

Location: Australia

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ARC Future Fellowships - Grant ID: FT0990405

Start Date: 03-2010

End Date: 2015

Amount: $891,200.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE220100185

Start Date: 2022

End Date: 12-2022

Amount: $675,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0770031

Start Date: 2008

End Date: 12-2009

Amount: $263,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0451224

Start Date: 01-2004

End Date: 12-2007

Amount: $290,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0347356

Start Date: 2002

End Date: 12-2003

Amount: $238,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE150100163

Start Date: 2015

End Date: 12-2016

Amount: $560,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE180100001

Start Date: 2018

End Date: 12-2018

Amount: $345,475.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE110100092

Start Date: 04-2011

End Date: 03-2013

Amount: $300,000.00

Funder: Australian Research Council