Polyomaviruses Polyomaviridae Virion Genome Genes and proteins Viruses and hosts Diseases Distinctive characteristics

 Virion  Naked icosahedral capsid (T=7). Diameter 45 nm.  Formed from 72 capsomers, pentamers of VP1. Polyomaviruses Polyomaviridae

 Genome  Circular double-stranded DNA, 5.3 Kb.  DNA is packaged as “minichromosome” with nucleosomes formed from cellular histones. Polyomaviruses Polyomaviridae

 Genes and proteins  Two transcription units: early and late.  Divergent transcription from a central control region containing origin of DNA replication.  Differential splicing produces 3–4 mRNAs from each transcription unit.  Early proteins (T antigens) regulate cell cycle and direct DNA replication.  Late proteins (VP1, 2, 3) make virus capsid. Polyomaviruses Polyomaviridae

 Viruses and hosts  Mouse polyomavirus; simian virus 40; polyomaviruses infecting birds, rodents, cattle;  and two human viruses: BK and JC virus. Polyomaviruses Polyomaviridae

 Diseases  Usually persistent, nonsymptomatic infections.  Progressive multifocal leukoencephalopathy, a rare demyelinating disease caused by human polyomavirus JC. Polyomaviruses Polyomaviridae

 Distinctive characteristics  Polyomaviruses are dependent on cellular RNA synthesis and RNA processing enzymes, and DNA replication systems.  T antigens interact with multiple signaling pathways and activate the cell cycle to facilitate viral DNA replication.  T antigens are oncogenes that can transform nonpermissive cells in vitro.  Polyomaviruses are tumorigenic in animals when administered at high concentrations, but rarely cause tumors in nature. Polyomaviruses Polyomaviridae

Virion  Mouse polyomavirus was discovered as a tumor-producing infectious agent  Simian virus 40 was found as a contaminant of Salk poliovirus vaccine  Polyomaviruses are models for studying DNA virus replication and tumorigenesis  Polyomavirus capsids are constructed from pentamers of the major capsid protein

Virion Fig Computer-generated image of mouse polyomavirus capsid.

Genome  The circular DNA genome is packaged with cellular histones  Circular DNA becomes supercoiled upon removal of histones  Supercoiled DNA can be separated from relaxed or linear DNA molecules

Genome Fig Components of polyomavirus virion. (a)A schematic diagram of a virion in cross-section, showing pentamers of VP1 (orange spheres), with single molecules of VP2 or VP3 (gray ovals) Circular viral DNA is bound to nucleosomes formed from cellular histones. (b) A larger view of a nucleosome, around which DNA is wrapped as a toroidal coil.

Genome Fig Gymnastics of circular DNA molecules.

Genes and proteins  Polyomavirus genes are organized in two divergent transcription units

Genes and proteins Fig Control regions on mouse polyomavirus genome. (a)The intergenic region, a noncoding region of 400 nt, bounded by the divergent promoters for early and late transcription (PE and PL) (b) Region where overlapping 3 ends of mRNAs are located

Genes and proteins  Virions enter cells in caveolae and are transported to the nucleus  The viral minichromosome is transcribed by cellular RNA polymerase II  Four early mRNAs are made by differential splicing of a common transcript

Genes and proteins Fig Alternative splicing of early mouse polyomavirus transcripts generates four different T antigens.

Genes and proteins  T antigens share common N-terminal sequences but have different C-terminal sequences  T antigens bring resting cells into the DNA synthesis (S) phase of the cell cycle  Small T antigen inhibits protein phosphatase 2A and induces cell cycling  Middle T antigen stimulates protein tyrosine kinases that signal cell proliferation and division

Genes and proteins Fig Small T antigen binding to protein phosphorylase 2A stimulates MAP kinase pathway.

Genes and proteins Fig Middle T antigen binding to c-Src activates pathways that stimulate cell metabolism and the cell cycle.

Genes and proteins  Large T antigen activates or suppresses transcription of cellular genes by binding to a number of important cellular regulatory proteins  Retinoblastoma protein (pRb) controls cell-cycle and S-phase gene expression.  The DNA J domain acts as a co-chaperone to dissociate pRb from E2F.  p53 blocks the cell cycle and induces apoptosis in response to virus infection.  p300 and general transcription factors control levels of transcription of viral and cellular genes.

Genes and proteins  Large T antigen hexamers bind to the origin of DNA replication and locally unwind the two DNA strands Fig Functional and protein-binding domains of simian virus 40 large T antigen.

Genes and proteins  Large T antigen hexamers assemble cellular DNA synthesis machinery to initiate viral DNA replication

Genes and proteins Fig Mechanism of bidirectional replication of polyomavirus DNA. (a)Initiation of DNA synthesis at viral replication origin (b)Extension of DNA chains beyond the initiation region by DNA polymerase (c) Joining of growing DNA strands

Genes and proteins  High levels of late transcripts are made after DNA replication begins 1. Repression of early transcription by large T antigen. 2. Activation or induction of new transcription factors. 3. Derepression of late transcription by dilution of a repressor. 4. Increased efficiency of processing and export of lateRNAs. 5. Readthrough RNAs hybridize with early transcripts and lead to their degradation.

Genes and proteins  Three late mRNAs are made by alternative splicing  How do polyomaviruses transform cells in vitro and cause tumors in vivo?  abortive infection  Transform nonpermissive cells (10 -5 )

Key Terms  Abortive  Apoptosis  Caveolae  Co-chaperone  Ethidium bromide  Gangliosides  Intergenic region  Myristate  Nonpermissive  Nucleosomes  Oncogene  Polyadenylation signal  Progressive multifocal  leukoencephalopathy (PML)  Cyclins  Divergent transcription  DNA gyrase  DNA helicase  Protein tyrosine kinase  Sialic acid  Sodium dodecyl sulfate  Spliceosome  Supercoiled  TATA box  Topoisomerase I  Toroidal coil  Transcription factors  Transcriptional enhancer