Flaviviruses are the agents of many mosquito-transmitted infections such as encephalitis and dengue. Hepatitis C virus is a member of the same virus family. Using Australian flavivirus Kunjin as a model and advanced techniques in molecular biology, biochemistry and electron micriscopy, the research at SASVRC has established international leadership in the area of flavivirus RNA replication and ultrastructure of virus-infected cells. The objectives of this application are to advance further our u ....Flaviviruses are the agents of many mosquito-transmitted infections such as encephalitis and dengue. Hepatitis C virus is a member of the same virus family. Using Australian flavivirus Kunjin as a model and advanced techniques in molecular biology, biochemistry and electron micriscopy, the research at SASVRC has established international leadership in the area of flavivirus RNA replication and ultrastructure of virus-infected cells. The objectives of this application are to advance further our understanding of how the flavivirus RNA replication complex synthesizes RNA and how this RNA is specifically packaged to produce infectious virus. To achieve these goals we will employ state-of-the-art molecular biology techniques based on manipulations with infectious complementary DNA copy of Kunjin virus RNA. The intimate understanding of these mechanisms in flavivirus replication should facilitate the design of efficient antiviral drugs by specifically targeting unique events in RNA replication and-or packaging. This may assist in the development of antiviral drugs for treatment of infections caused by other higly pathogenic flaviviruses in Australia, such as dengue, Japanese encephalitis and Murray Valley encephalitis, and in the rest of the wirld such as New York strain of West Nile virus as well as the related heptitis C virus. Understanding the mechanisms of Kunjin virus replication and assembly will also aid in the further development of this virus as a safe vaccine vector against other viruses, e.g. HIV, and diseases such as cancer.Read moreRead less
Molecular Analyses Of Flavivirus RNA Replication, Encapsidation, And Complementation
Funder
National Health and Medical Research Council
Funding Amount
$602,545.00
Summary
Flaviviruses are the agents of many mosquito-transmitted infections such as encephalitis and dengue. Hepatitis C virus is a member of the same virus family. Using Australian flavivirus Kunjin as a model and advanced techniques in molecular biology, biochemistry and electron micriscopy, the Flavivirus Research Unit at SASVRC has established itself as an international leader in the area of flavivirus RNA replication and ultrastructure of virus-infected cells. The objectives of this application are ....Flaviviruses are the agents of many mosquito-transmitted infections such as encephalitis and dengue. Hepatitis C virus is a member of the same virus family. Using Australian flavivirus Kunjin as a model and advanced techniques in molecular biology, biochemistry and electron micriscopy, the Flavivirus Research Unit at SASVRC has established itself as an international leader in the area of flavivirus RNA replication and ultrastructure of virus-infected cells. The objectives of this application are to advance further our understanding of how the flavivirus RNA replication complex is assembled, how it synthesizes RNA and how this RNA is specifically packaged to produce infectious virus. To achieve these goals we will employ state-of-the-art molecular biology techniques based on manipulations with infectious complementary DNA copy of Kunjin virus RNA. The intimate understanding of these mechanisms in flavivirus replication should facilitate the design of efficient antiviral drugs by specifically targeting unique events in RNA replication and-or packaging. This may assist in the development of antiviral drugs for treatment of infections caused by other higly pathogenic flaviviruses in Australia, such as dengue, Japanese encephalitis and Murray Valley encephalitis, as well as of the related heptitis C virus.Read moreRead less
Plasmids are extra mini-chromosomes that are present in many bacteria. They carry information that enables their hosts to survive and prosper in hostile environments. Plasmids are able to spread rapidly between bacteria, ensuring that the information they carry is rapidly disseminated throughout bacterial populations. Many plasmids carry information that increases the virulence of their host bacteria, because it adds to their repertoire of toxins and other adjuncts to invasiveness and colonisati ....Plasmids are extra mini-chromosomes that are present in many bacteria. They carry information that enables their hosts to survive and prosper in hostile environments. Plasmids are able to spread rapidly between bacteria, ensuring that the information they carry is rapidly disseminated throughout bacterial populations. Many plasmids carry information that increases the virulence of their host bacteria, because it adds to their repertoire of toxins and other adjuncts to invasiveness and colonisation, or enables them to survive in the presence of antibiotics. The emergence of multi-drug resistant bacteria and the rapid spread of the ability of bacteria to withstand most antibiotics available to date were mediated by plasmids. Plasmids also carry information that ensures their own survival. The consequence of this is that their bacterial hosts retain the plasmids, even when it is no longer beneficial to do so. For example, plasmids carrying information for resistance to antibiotics are not lost when their bacterial hosts grow in the absence of antibiotics. This is because plasmids have control systems, which ensure that on the one hand, replication of the plasmid keeps pace with the replication of its host, and on the other hand that the plasmid does not produce so many copies of itself that it overwhelms its host. This project examines the intricate regulatory system that a group of antibiotic-resistance plasmids uses to ensure that on average each plasmid molecule is replicated once per bacterial cell cycle. This system uses an antisense RNA, a tertiary RNA structure (pseudoknot) that acts as a translational switch, and a protein that interacts with different sequences on the plasmid to initiate replication. Detailed knowledge of the processes underlying this complex system is required if we are to develop new treatments that will lead to elimination of antibiotic-resistance and virulence-contributing plasmids from populations of pathogenic bacteria.Read moreRead less
Current combination antiviral therapy can't cure an HIV infection because long-lived T-cells carrying latent HIV DNA can rekindle the infection when drugs are removed. We will study elements in HIV genetic code that control expression of HIV proteins from latent HIV. A detailed molecular understanding of the structure and function of these HIV RNA elements and the viral and host cell factors that interact with them will expose new targets for therapy of latent HIV.
Title: Structure of hepadnaviral pre-genomic RNA. We aim to study the replication strategy of human hepatitis B virus (HBV), a member of the hepadnavirus family. Hepadnaviruses infect hepatocytes in the liver and are released in high numbers into the bloodstream. Infection is transmitted by blood or sexual contact. Hepadnaviruses cause acute and chronic infection with varying degrees of liver disease. The HBV DNA genome is formed by copying of a viral pre-genome made of RNA, into DNA. This proce ....Title: Structure of hepadnaviral pre-genomic RNA. We aim to study the replication strategy of human hepatitis B virus (HBV), a member of the hepadnavirus family. Hepadnaviruses infect hepatocytes in the liver and are released in high numbers into the bloodstream. Infection is transmitted by blood or sexual contact. Hepadnaviruses cause acute and chronic infection with varying degrees of liver disease. The HBV DNA genome is formed by copying of a viral pre-genome made of RNA, into DNA. This process is called reverse transcription and is performed by the viral polymerase. Reverse transcription occurs within viral nucleocapsids made of core antigen. After formation of the new viral DNA genome, nucleocapsids are enveloped in surface antigen and are released from the cell. It is assumed that 1 copy of HBV pre-genomic RNA is packaged within each viral nucleocapsid. However, members of the retrovirus family that have common evolutionary origins to hepadnaviruses and also replicate via reverse transcription, contain 2 copies of RNA. The human immunodeficiency virus (HIV), the AIDS virus, is a well-studied example. In HIV infection 2 RNA genomes are packaged into each nucleocapsid and form a dimeric RNA genome. The HIV RNA is able to fold into a series of stem loops that promote formation of dimers. During the reverse transcription step in HIV replication, the polymerase switches templates and forms new combined strains of virus. The project aims to determine if 2 copies of pre-genomic RNA are packaged into HBV nucleocapsids. HBV pre-genomic RNA is able to form stem loop structures similar to those in HIV and has the potential to form dimeric RNA. If 2 copies of HBV pre-genomic RNA are packaged this will allow us to redefine the viral replication strategy and to develop a greater understanding of the relationships between hepadnaviruses and retroviruses. The formation of dimers will also provide a mechanism for recombination between HBV strains.Read moreRead less
Control Of Viral Replication By Non-coding Viral RNA
Funder
National Health and Medical Research Council
Funding Amount
$502,270.00
Summary
In 25 years since identified, HIV-AIDS deaths have exceeded 30 million and 40 million more are now living with HIV. The toll will soon far surpass any other infectious disease epidemic in history, or even military deaths from war in the past century. While effective combination drug therapies are available, multi-drug resistant HIV strains are commonly transmitted, leaving some patients with limited treatment options. New classes of drugs aimed at different steps in virus replication are urgentl ....In 25 years since identified, HIV-AIDS deaths have exceeded 30 million and 40 million more are now living with HIV. The toll will soon far surpass any other infectious disease epidemic in history, or even military deaths from war in the past century. While effective combination drug therapies are available, multi-drug resistant HIV strains are commonly transmitted, leaving some patients with limited treatment options. New classes of drugs aimed at different steps in virus replication are urgently needed. We have discovered that viral RNAs that do not code for protein serve important functions in HIV replication. We will study the molecular mechanisms these non-coding (intron) RNAs previously considered junk use to support of HIV gene expression and assess their potential as drug targets. First, we will investigate the role of these junk RNA loops, or lariat introns, produced in large amounts during the HIV replication cycle. Retroviruses employ RNA splicing to make mRNA for envelope and regulatory accessory genes. The complex alternative RNA splicing pattern of HIV spawns several non-coding lariats, including the lariat-intron that contains much of the removed env coding sequence. We have made the counterintuitive finding that the env-lariat dramatically enhances expression of Env protein. We will examine how this occurs and the involvement of the new class of gene-expression controlling micro-RNAs in this process. We will test for functional activity from the other lariat-introns that are produced by HIV. Second, we will characterise the mRNA-element required for efficient expression of the HIV envelope glycoprotein, Env gp160, which is essential for virus binding and entry during infection. This RNA-element directs the cell protein translation machinery to commence protein synthesis at the start of the Envgp160 rather than at upstream start sites for Vpu and Rev. We will determine how this RNA element works, its structure, and how it might be inactivated.Read moreRead less