3 1 13 Topic 19 Chapter 17 Retroviruses Retrovirus Transcription Transcription the process by which genetic information represented by a sequence of DNA nucleotides is copied into newly synthesized molecules of RNA with the DNA serving as a template Retrovirus any group of RNA viruses that insert a DNA copy of their genome into the host cell in order to replicate ex HIV Retroviruses Retrovirus is a RNA virus that replicates in a host cell Has two copies of RNA genome packaged in a protein capsid Uses own reverse transcriptase enzyme to produce DNA from its RNA genome This is reverse of the usual pattern where RNA is produced using DNA o Retro backwards o The new complementary DNA cDNA is then incorporated into the host s genome becomes a provirus required to assemble new copies of the virus o virus The cell then treats the viral DNA as part of its own instructions where it produces the proteins It undergoes usual transcription translation process to express the genes carried by the RNA DNA RNA Protein Retroviral Life Cycle o Begins with outer proteins of envelope binding to host cell Envelope is composed of lipids from host plasma membrane during budding process as well as glycoprotein encoded by the env gene Proteins are recognized signaling for the RNA to be released o Retrovirus fuses with host cell and RNA inside capsid is inserted o RNA virus then undergoes reverse transcriptase to create cDNA o cDNA is then moved into host nucleus and integrated into host genome Integration occurs via a transposition like mechanism o Transcription occurs creating viral genomic RNA o Translation occurs on the viral genomic RNA creating more virus proteins which will then be used to assemble new copies of the virus o the new assembled copies then exit the host cell by exocytoses to become new retrovirus that will infect other host cells Retroviral Integration o the integrated virus is the provirus it is the cDNA that is reverse transcribed from RNA o Consequences of Retrovirual Integration If integrated into germline retrovirus is transmitted to offspring present in all cells If host sequences recombine w integrated retroviral sequence become part of retroviral genome Recombination of retrovirus with host genome is how retroviruses evolve Retroviral Genome o Has a 5 7 methylguanosine cap and a 3 poly A tail o R region terminal noncoding regions example of plus strand virus explained below Short 10 80nt sequence which forms a direct repeat at both ends of genome Important for RNA replication o U5 short sequence after R A unique non coding region which is the first of the genome to be reverse transcribed Forms the 3 end of the provirus dsDNA genome o Typically consists of 3 4 genes each gene codes for multiple proteins gag group specific antigen protein component of nucleoprotein core of virion o virion a complete infective form of a virus outside a host cell with a core of RNA or DNA and a capsid pol polymerase nucleic acid synthesis and recombination env envelope infection o responsible for synthesis of viral DNA integration into host DNA after components of the envelope that sequester cellular membrane o plays a role in association and entry of virion into host cell o critical for retrovirus to be infectious o U3 short sequence before R A unique non coding region which forms the 5 end of the provirus after reverse transcriptase Contains the promoter elements responsible for transcription of the provirus Retroviral Proteins o Each gene codes multiple proteins frame shifting and processing gag encodes 3 proteins code for the structural components of capsid and core gag proteins are major components of viral capsid pol encodes 3 proteins env Encodes 2 proteins codes for reverse transcriptase protease and integrase codes for the retroviral coat proteins o Viruses can be Single Stranded or Double Stranded Single Stranded RNA Viruses can be plus or minus Plus Strand RNA virus ex poliovirus o strand viral RNA is similar to mRNA same sense thus can be immediately translated by host cell Minus Strand RNA Virus ex influenza virus o strand viral RNA is complementary to mRNA thus must be converted to positive sense RNA by RNA polymerase before translation RNA polymerase works on RNA only RNA dependent o strand is inactive so cannot infect until converted into plus strand Ex Flu Double Stranded RNA Virus RNA virus is double stranded Must be transcribed by RNA polymerase into plus minus strand o Translating gag pol proteins Stop codons located between each gene gag and pol produce three proteins each the three proteins are three domains connected by polypeptide bonds that are then cut by protease also a mechanism that allows stop codon to be skipped a special tRNA glutamyl tRNA anti termination that recognizes stop codon is inserted allowing system to keep going can also skip by causing ribosome frameshift where the entire reading frame is changed thus removing the stop codon However this read through is inefficient because it ends up making 20x more gag then pol proteins o Translating env proteins Env is translated from a spliced RNA Gag and pol are spliced out only leaving beginning of gag and env is then transcribed Env creates two proteins or two domains that are connected by polypeptide bonds that are then cut by protease Appearance of RNA and DNA forms of Viruses o When RNA is reverse transcribed into cDNA an additional U3 region is added to 5 end of sequence o How it is added 1 Synthesis of 1 st DNA Strand the tRNA anneals to U5 Primer Binding Site reverse transcriptase then adds primer in 5 3 direction starting from tRNA when enzyme reaches end of template RNA strand the 5 end of template is degraded leaving a 3 over hang this entire new primer sequence then shifts down the sequence until it reaches the RNA R repeat region o the 3 over hang pairs with the R region and the U5 region will hang off synthesis then proceeds across entire RNA until reaches the 5 R region the tRNA is then removed the old bottom RNA strand is then degraded leaving only a few bases to act as primer from fragment DNA synthesis proceeds to the end of the strand the newly synthesis DNA sequence then jumps to other end of strand o the 5 U3 region is now the new 3 U3 overhang DNA synthesis then proceeds across entire DNA producing mature DNA copy of virus with the new additional U3 region added to 5 end 2 Synthesis of 2 nd DNA Strand 1st DNA Strand 2nd DNA Strand U3 Reverse Transcriptase o There are three main activities 1