Retroviruses
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Retroviruses Replication cycle Envelope glycoproteins Receptors Entry Integration Expression Endogenous retroviruses HIV reverse transcription and viral evolution Phylogenetic trees The Family Retroviridae Genus Example Group Alpha retroviruses Avian leukosis virus ALV Beta retroviruses Murine mammary tumor virus Simian retroviruses MMTV SRV Gamma retroviruses Murine leukemia virus MLV Delta retroviruses Human T-cell leukemia virus HTLV Epsilon retroviruses Walleye dermal sarcoma virus Lentiviruses Human immunodeficiency virus HIV Spumaviruses Simian foamy virus SFV Simple Complex Simple Retroviruses MLV gag pol e.g. murine leukemia virus (MLV) env Complex Retroviruses gag-pol-env-tat-rev-nef-vif-vpr-vpu gag-pol-env-tat-rev-nef-vif-vpr-vpx HIV-1 HIV-2/SIV gag-pol-env-tax-rex-HBZ HTLV Complex retroviruses have additional genes that are described as regulatory or accessory. From “Retroviruses” CSHL Coffin, Hughes,Varmus A. HIV B. Beta retrovirus, MMTV C. Alpha , ALV Structure of retrovirus particles tRNA membrane TM SU ENV capsid CA reverse transcriptase RT matrix MA POL protease PR nucleocapsid NC integrase IN RNA genome GAG RNA genome Two copies of viral RNA are in each virus particle. 5'-Cap-R-U5---PBS--DLS------GAG-POL-ENV------U3-R-AAAAAA-3' R U5 Cap and poly A R repeats U5, U3 unique regions at each end PBS primer binding site (Psi) packaging signal DLS dimerization site U3 R Replication Cycle 2 1 Virions bind receptors on cell surface Virion and cell membranes fuse and virus infects cell RTC: reverse transcriptase complex 3 4 Reverse transcription to form viral DNA PIC: pre-integration complex 5 Integration into chromosome integrase Proviral DNA 6 mRNAs proteins 7 Virion RNA 8 9 Virus particles bud from cell membranes Virus particles mature and become infectious Receptors for retroviruses CD134 CXCR4 TNF-R family CD4 CCR5/CXCR4 FIV HIV/SIV CAT-1 MLVe Pit-1 FeLV-B GALV DD GLUT-1 HTLV-I HTLV-II TVA LDLR-like ALV-A TVB TNF-R family ALV-B ALV-D ALV-E Retroviral trafficking A viral substructure is released and begins trafficking to sites of integration. The RTCs and PICs exploit microtubules to transport towards the chromosomes for integration. Retroviruses and dividing cells Simple retroviruses e.g. MLV, cannot infect non-dividing differentiated cells. HIV and other lentiviruses can infect non-dividing, differentiated cells e.g. macrophages, with the RTC/PIC trafficking through the nuclear membrane Host proteins involved in early events after HIV entry Host Protein Proposed function FEZ1, dynein & microtubules Transport of RTCs/PICs Cyclophilin A uncoating Pin1 uncoating PDZD8 uncoating Importin-7 Nuclear import Transportin-3 Nuclear import LEDGF/p75 (transcription coactivator) Tethering of integrase to chromatin Integration The retroviral cDNA in the PIC becomes integrated into host DNA. No specific site for integration. Integration catalyzed by INTEGRASE. INTEGRATION 1) Two ends held together in the PIC CATT GTAA AATG TTAC 2) 3’ processing OH CA GTAA AATG AC HO 3) Host DNA attacked ACTG TGAC TGAC ACTG 4) Gap repair CA ACTG GTAA AATG TGAC AC ACTG TGAC TG AC CA ACTG GT TGAC Note:flanking sequences now repeated HIV has 5-base duplication MLV, FeLV have 4-base duplication ALV has 5-base duplication Retrovirus integration sites Virus Host protein anchor Integration sites MLV BET near transcription starts e.g. CpG islands. HIV LEDGF along transcriptionally active genes ALV ? weak preference for active genes and no preference for transcription starts. Synthesis of retroviral mRNA Full length HIV mRNA in virions R U5 U3 R mRNA synthesis U3 AP-1 NF-AT NRF NF-kB SP-1 TAR TATA (IL2/NRE) HIV-1 LTR region/promoter RNA pol II R U5 AAUAAA Simple retrovirus make two mRNAs U3 R U5 GAG PRO POL GAG-POL ENV U3 R U5 Y GAG/POL mRNA ENV mRNA R U5 U3 R Complex Retroviruses HIV rev tat vif gag pro env pol nef vpr vpu Complex retroviruses have additional genes that are described as regulatory or accessory. Tax ---tax transactivates transcription. ---Acts on HTLV LTR, stimulating transcription. ---Binds GC rich regions on the LTR adjacent to CRE sites. Then interacts with CREB transcription factors bound to these adjacent CRE sites. ---However, tax has major effects on other transcription factors e.g. NFB. ---Upregulates NFB by stimulating dissociation with IB Tat CDK9 Cyclin T P-TEFb complex TAR( tat responsive element) Secondary RNA structure at the start of mRNAs TAT RNA pol II mRNA P P P DNA without tat, mRNA transcription is curtailed Rev/Rex ---rev and rex are functionally equivalent. ---rev and rex regulate splicing of viral mRNAs. ---confer nuclear export of mRNAs before they are fully spliced. HIV-1 mRNAs and the rev response element RRE GAG/POL ENV AAAA Early mRNAs NEF Late mRNAs AAAA TAT AAAA REV AAAA NEF RRE GAG/POL VIF VPR VPU ENV AAAA GAG/POL AAAA VIF AAAA VPR AAAA VPU/ENV REV Rev increases the production of unspliced or singly spliced mRNAs that code for late mRNAs. Stem loop II Rev binding site Rev binds to a specific site on the RRE and then oligomerizes around the first bound molecule. rex has the same function for HTLV. HIV Simian beta-retroviruses Stem loop II Rev Constitutive transport Element: CTE Rev binding site Crm1 nucleoporins TAP nucleoporins 5S ribosomal RNA export path mRNA export path Translation of Gag and Gag/Pol precursor proteins 1. Gag is translated as a long precursor protein. 2. 5% of Gags are made as a Gag/Pol precursor. 3. Translational readthrough e.g. MLV 4. Frame shift e.g. HIV Proteins produced: Gag Gag/Pol R U5 GAG POL ENV U3 R Virion maturation Viral protease cleaves the Gag and Gag/Pol precursor proteins into mature MA, CA, NC Gag proteins and RT, IN, PR enzymes. Virion maturation is essential for the virion to be infectious. Endogenous Retroviruses (ERVs) ---Proviruses that become integrated into the chromosomes of germ cells can be passed down vertically via the sperm or egg DNA. ---The human genome is littered with 1000s of proviral sequences and make up about 8% of the genome. ---These endogenous retroviruses (ERVs) represent archived or 'fossil' evidence of past retroviral infections. ERV Classification and Taxonomy Note: Class I gamma-related Class II beta-related Class III spuma-related Endogenous lentiviruses shown in rabbits and lemurs but not humans. ERV Classification and Taxonomy ---No sensible taxonomy system. ---Named after host tRNA involved in reverse transcription e.g. HERV-W (class I), HERV-K (class II), HERV-L (class III) etc. ---Sometimes ERVs are arbitrarily named by amino acid motifs e.g. HERV-FRD, in their sequences (class I). ERV Families Once integrated into the germline, endogeneous retroviruses tend to proliferate. So a single germ line infection can give rise to family of closely related elements Colonization of primates by ERVs from; Bannert and Kurth, Annu. Rev. Genom. Human Genet. 2006. 20-39 HERV K106 ---recent acquisition about 150,000 years ago. probably after the emergence of modern humans. ---defective. Nearly all endogenous retroviruses are dead ---Generally endogenous retroviruses are inactive and open reading frames have become interrupted by acquisition of many stop codons as well as deletions. ---Methylation also plays a role. ---Many are in the form of a single LTR. ---Mice, cats sheep, Koalas and pigs carry endogenous proviruses that can give rise to fully infectious retroviruses. Some ERV genes are open despite ancient origins ---HERV W env (syncytin) ---HERV-FRD env (syncytin 2) ---expressed in the placenta ---believed to be involved in the formation of the syncytiotrophoblast The syncytiotrophoblast is at the junction of maternal and fetal tissues From Gude et al. 2004. Thrombosis Res. 114:397-407 ERVs have been associated with: autoimmune disease Cancers Virus particles budding from a human teratocarcinoma cell line Diseases associated with endogenous retroviruses FeLV and leukemia in cats ---exogenous FeLV-A recombines with endogenous FeLV sequences to give rise to the leukemogenic FeLV-B. ---FeLV-B recombinants use a different receptor and have a broader tropism ---leukemias caused by insertional mutagenesis
