Retroviruses and AIDS 2

Discovery of retroviruses Retroviruses possess a unique enzyme known as RT (reverse transcriptase) RT uses viral RNA as a template for making DNA copy which integrate in to the chromosome of the host cell and serves either as basis for viral replication or as oncogene. Howard Temin and David Baltimore received Nobel Prize for discovery of RT enzyme.

Discovery of HIV In 1981 new clinical syndrome characterized by profound immunodeficiency was recorded in male homosexual and termed AIDS. Unusual prevalence of Pneumocystis carinii pneumonia in in a group of young previously healthy male homosexual. Kaposi’s sarcoma (rare cancer) in previously healthy male homosexual??

Discovery of HIV First isolation of HIV-1 made by Luc Montagnier and Barre-Sinoussi at Pasteur institute in Paris in 1983. This observation is confirmed by Robert Gallo in the USA

Discovery of HIV HIV-2 isolated from mildly immunosuppressed patient n west Africa. 5000 cases of HIV-1 cases per Day? 41 million people have been infected world wide. HIV-2 account for 4.5% of HIV cases.

Retroviridae

Lentiviruses

Primates infected with lentiviruses > 30 species of African primates naturally infected with SIV SIV infections: natural acquired not known Natural infections: >50% of adults nonpathogenic Chimpanzee the only ape

Photograph by Karl Ammann

Photograph by Karl Ammann

Properties of HIV Classification The family Retroviridae is named for RT. (Retro= Backwards) Seven genera is now recognised (only two of them cause disease in human):

Properties of HIV Lentivirus: containing HIV-1 and HIV-2, characterised by: Cone shaped Nucleocapsid absence of oncogenicity and the lengthy and insidious onset of clinical signs.

Properties of HIV BLV-HTLV retroviruses: contain HTLV-I and II: characterised by ability to cause tumours rather than immunosuppression. Spumavirus : Causes characteristic foamy appearance in infected primate cell culture. (they are not pathogenic).

Morphology of HIV HIV particle is 100-150 nm in diameter. Outer envelope of lipid penetrated by 72 glycoprotein spike (the lipid envelope protein) The envelope protein is composed of two subunits: the outer glycoprotein knob (gp120) and transmembrane protein (gp41) The receptor binding site for CD4 is present on gp120 as well as very important antigen such as V3 loop.

Morphology of HIV The inner surface of virus lipid envelope is lined by matrix protein (p17)?. There is also abundant cellular proteins in the lipid envelope (MHC class I and II) antigens. In HIV-1 the lipid envelope encloses an icosahedral shell of protein (p17), within which is a vase or cone shaped protein core (p24, p7, and p9) containing two molecules of positive sense ssRNA The RNA genome is associated with several copies of RT, integrase, and protease.

HIV genome tat: transactivation. vif: viral infectivity Positive sense ssRNA genome The genome is approximately 10kb in size The genome contain control genes which can enhance viral replication: rev: regulator of virus tat: transactivation. vif: viral infectivity repressor genes: nef: negative factor

HIV genome The genome is flanked at each end by LTR (Long Terminal Repeats) 3’ LTR has the polyadenylation signal and 5’LTR has the enhancer promotor sequence for viral transcription. The pol gene code for reverse transcriptase RT, integrase and protease.

The HIV-1 genome rev vif nef tat 5’ LTR 3’ LTR vpu vpr gag pol env p17 matrix antigen p24 capsid antigen p6/7 nucleocapsid reverse transcriptase protease integrase envelope glycoprotein (gp120) transmembrane glycoprotein (gp41)

Binding HIV binds to specific receptor on the surface of CD4+ T lymphocytes (T-helper cells) It also infects: B lymphocytes Macrophages dendritic cells brain cells. Second subsidiary receptor belongs to chemokine receptor family CXCR4 on the T-cells and CCR5 on the surface of macrophages.

HIV lifecycle maturation virus binding fusion virion assembly and release ssRNA (+) reverse transcription translation of viral proteins dsDNA nuclear transport integration transcription nucleus cytoplasm

HIV life cycle After attachment the virus penetrate the cell by fusion from without (Mediated by gp120 and gp41) Synthesis of viral cDNA starts when the virion enters the cell cytoplasm. The viral RT enzyme directs the synthesis of cDNA strand (the minus strand) using host positive RNA as a primer and the viral genomic RNA as a template. Viral RNAse enzymatically remove the viral RNA while the RT synthesize the second DNA strand (plus strand).

HIV life cycle Viral dsDNA will enter the Nucleolus of the host cell as a pre-integration complex (compose of viral protein M, Vpr, integrase, and dsDNA) Can exist extra-chromosomally as linear or circular form the integration of dsDNA to the host chromosome occurs (forming pro-viral DNA) by the enzyme integrase After integration viral and cellular factors are needed to activate HIV transcription. Initial expression of viral RNA is stimulated by vpr and further stimulated by cellular transcription factors .

HIV life cycle Assembly of new virion can begin by proteolytic cascade by viral proteases. Different viral structural proteins begin to assemble with the p24 as a core and p7 enclosing viral RNA. Viral genome assemble in the cytoplasm. Retroviruses including HIV are release from the infected cells by budding from the infected cells. The pro viral DNA may reside quietly in the chromosome for years.

Genetic Variability RT has NO proof-reading mechanism therefore mutations (point point mutations and deletions/insertions) occur Quasispecies = swarm of genetically distinct yet related viruses

Effects of Variability Immune escape by changing/masking antigenic determinants Resistance to anti-retroviral drugs Point mutations in enzymatic proteins RT - resistance to nucleoside and non-nucleoside analogues Protease - resistance to protease inhibitors Altered cytopathogenicity Env and particularly V3 mutations alter co-receptor usage Different cell tropism, eg. Macrophages, T-cells, glial cells, langerhans cells etc. Different tissue tropisms, e.g. brain

Summary HIV member of the Retroviridae family (reverse transcriptase) Entry mediated by CD4 plus co-receptor Reverse transcription leads to errors Virus can become integrated into chromosome (can be latent) Transcription – short (spliced) then long RNAs New virus buds at surface.