Oncogenic dna virus

This presentation includes information about DNA oncogenic viruses and latent and lytic replication of Herpes virus.

1. Femina Anjum PhD Scholar (Animal Biotech.) Deptt . Of Veterinary microbio . &Biotech. Rajasthan university of Veterinary and Animal Sciences Bikaner Oncogenic DNA viruses 1

2. An oncovirus is a virus that can cause cancer and which induce malignant transformation of cells on culture. This term originated from studies of acutely transforming retroviruses in the 1950–60s . Now refers to any virus with a DNA or RNA genome causing cancer. The vast majority of human and animal viruses do not cause cancer . 2

3. Cancers are the result of a disruption of the normal restraints on cellular proliferation. There are two classes of these genes in which altered expression can lead to loss of growth control:  Those genes that are stimulatory for growth and which cause cancer when hyperactive. Those genes that inhibit cell growth and which cause cancer when they are turned off. Viruses are involved in cancers because they can either carry a copy of one of these genes or can alter expression of the cell's copy of one of these genes. 3

4. How do oncogenes and tumor suppressors work? oncogene Tumor suppressor

5. Oncogenesis - R esult of genetic changes that alter the expression or function of proteins that play critical roles in the control of cell growth and division Proto- oncogenes - normal (pre-mutation) (pre-diseased) genes - present in normal cells - conserved in their genomes - code for proteins which regulate cell growth &differentiation Oncogenes - mutated versions of proto-oncogenes

6. Activation of Cellular Oncogenes Oncogene activation via insertional mutagenesis -the presence upstream from a c-onc gene of an integrated provirus , with it’s strong promoter and enhancer elements may enhance the expression of c-gene. e.g. integrated avian leukosis provirus increase the synthesis of c-myc oncogene product 30-100 fold. Oncogene activation via transposition – Transposition of c- onc may result in their enhanced expression by bringing them under the control of strong promoter and enhancer elements. E.g. Burkitt’s lymphoma Oncogene activation via gene amplification- Increase in gene copy number leads to corresponding amount of oncogene products , thus producing cancer. E.g. c- myc gene , c- ras gene Oncogene activation via mutation- Mutation alter the function of corresponding oncoprotein . it may be induced by physical or chemical means or in course of recombination with integrated retroviral DNA.

7. Conversion of Proto-oncogene to Oncogene

8. CELLULAR ONCOGENES Present in cancer cells Contains introns characteristic of eukaryotic cells Encodes proteins triggering transformation of normal cells VIRAL ONCOGENES Present in viruses Host cell origin Do not possesss introns Also called ‘cancer genes’ Encodes proteins triggering transformation of normal cells into cancer cells

9. MECHANISM OF ONCOGENECITY Introduction of new Alteration of expression of ‘Transforming gene’ preexisting cellular gene into the cell Loss of normal growth regulation processes Affection of DNA repair mechanisms Genetic instability Mutagenic phenotype DIRECT ACTING INDIRECT ACTING

10. Worldwide , the WHO International Agency for Research on Cancer estimated that in 2002, 17.8% of human cancers were caused by infection, with 11.9% being caused by one of seven different viruses . The importance of this is that these cancers might be easily prevented through vaccination (e.g., papillomavirus vaccines), diagnosed with simple blood tests, and treated with less-toxic antiviral compounds. 10

11. 11

12. 12

13. Oncovirus DNA oncogenic viruses RNA oncogenic viruses 13

14. TUMOR VIRUSES DNA VIRUSES RNA VIRUSES VIRAL ONCOPROTEIN EXPRESSION VIRAL ONCOGENE PROTO ONCOGENE CONVERSION PROVIRAL INSERTION NEAR CELLULAR ONCOGENE TRANSFORMING NON TRANSFORMING INACTIVATION OF TUMOR SUPPRESSOR GENES

15. DNA tumor virus have a DNA genome that is transcribed into RNA which is translated into protein. They  have two life-styles : In permissive cells, all parts of the viral genome are expressed. In cells that are non-permissive for replication, viral DNA is usually, but not always, integrated into the cell chromosomes at random sites. DNA Oncogenic viruses 15

16. DNA oncoviruses typically impair two families of tumor suppressor proteins: tumor proteins p53 and the retinoblastoma proteins ( Rb ). While several DNA oncoviruses have been discovered, three have been studied extensively.  Adenoviruses  can lead to tumors in rodent models but do not cause cancer in humans. Simian virus 40 (SV40), a  polyomavirus , can cause tumors in rodent models but is not oncogenic in humans .   The  Human Papillomavirus-16 (HPV-16) has been shown to lead to cervical cancer and other cancers, including head and neck cancer .   All three of these DNA oncoviruses are able to integrate their DNA into the host cell, and use this to transcribe it and transform cells by bypassing the G1/S checkpoint of the cell cycle. 16

17. Integration of viral DNA The DNA is believed to be inserted during transcription or replication, when the two annealed strands are separated .  This event is relatively rare and generally unpredictable; there seems to be no deterministic predictor of the site of integration .   After integration, the host’s cell cycle loses regulation from Rb and p53, and the cell begins cloning to form a tumor. 17

18. The G1/S Checkpoint Rb and p53 regulate the transition between G1 and S phase, arresting the cell cycle before DNA replication until the appropriate checkpoint inputs, such as DNA damage repair, are completed .   p53 regulates the p21 gene, which produces a protein which binds to the Cyclin D-Cdk4/6 complex .  This prevents Rb phosphorylation and prevents the cell from entering S phase .   In mammals, when Rb is active ( unphosphorylated ), it inhibits the E2F family of transcription factors, which regulate the  Cyclin E-Cdk2 complex, which inhibits Rb , forming a positive feedback loop, keeping the cell in G1 until the input crosses a threshold .   18

19. Inactivation of p53 The  adenovirus E1B protein (55K) prevents p53 from regulating genes by binding to the site on p53 which binds to the genome .   In SV40, the large T antigen (LT) is an analogue; LT also binds to several other cellular proteins, such as p107 and p130, on the same residues .  LT binds to p53’s binding domain on the DNA (rather than on the protein), again preventing p53 from appropriately regulating genes . The HPV protein E6 binds to a cellular protein called the E6-associated protein (E6-AP, also known as UBE3A), forming a complex which causes the rapid and specific  ubiquitination  of p53 . 19

20. Inactivation of Rb Rb is inactivated (thereby allowing the G1/S transition to progress unimpeded) by different but analogous viral oncoproteins . The adenovirus early region 1A  (E1A) is an oncoprotein which binds to Rb and can stimulate transcription and transform cells .   SV40 uses the same protein for inactivating Rb , LT, to inactivate p53 .   HPV contains a protein, E7, which can bind to Rb in much the same way .   20

21. Variations There may be many different mechanisms which have evolved separately; in addition to those described above, for example, the Hepatitis B virus (an RNA virus) inactivates p53 by sequestering it in the cytoplasm . SV40 has been well studied and does not cause cancer in humans, but a recently discovered analogue called Merkel cell polyomavirus  has been associated with Merkel cell carcinoma, a form of skin cancer .  The Rb binding feature is believed to be the same between the two viruses . 21

22. Classification of DNA Oncogenic viruses Small DNA tumor viruses Complex DNA tumor viruses S. No. Family Members 1 Papillomaviridae Papilloma virus 2 Polyomaviridae Polyomaviruses Mouse polymavirus Simian virus 40 Human polyomavirus 3 Adenoviridae Adenovirus S. No. Family Members 1 Herpesviridae Herpesviruses Epstein barr viruses (Human Herpes virus 4) Kaposi’s sarcoma Herpes Viruses (Human Herpes Viruses 8) Human cytomegalovirus (Human Herpes Virus 5) 2 Hepadnaviridae Hepatitis B virus 22

23. Oncogenic Papillomaviruses Papillomas are hyperplastic epithelial outgrowths that generally regress spontaneously. Occasionally, however, infections by some papillomavirus types may cause malignant cellular transformation, resulting in the development of cancer. In warts, the papillomavirus DNA remains episomal . As the pattern of integration is clonal within cancers, each cancer cell carries at least one, and often many incomplete copies of the viral genome. The site of virus integration is random, and there is no consistent association with cellular proto- oncogenes . For some papillomaviruses , integration disrupts one of the early genes,  E2 , which is a viral repressor. These oncogenes alter normal cell growth and division and the overexpression of E6 and E7 -critical step in malignant transformation by a human papillomavirus . However, bovine papillomavirus type 1 -cause equine sarcoids predominantly through changes in cell proliferation -mediated by the E5 oncoprotein . SMALL DNA TUMOR VIRUSES 23

24. Polyomavirus - or adenovirus -transformed cells do not produce virus. Most of the integrated viral genomes are complete in the case of the polyomaviruses , but defective in the case of the adenoviruses. Only certain early viral genes are transcribed, albeit at an unusually high rate. Their products, demonstrable by immunofluorescence , used to be known as  tumor (T) antigens . Virus can be rescued from polyomavirus -transformed cells—that is, virus can be induced to replicate by irradiation, treatment with certain mutagenic chemicals, or cocultivation with certain types of permissive cells. This cannot be done with adenovirus-transformed cells, as the integrated adenovirus DNA contains substantial deletions. 24

25. Oncogenic Herpesviruses Marek’s disease virus of chickens ( gallid herpesvirus 2) transforms T lymphocytes, causing them to proliferate to produce a generalized polyclonal T lymphocyte neoplasm. The disease is preventable by vaccination with live-attenuated virus vaccines that lack the retrovirus v- onc  genes that are present in Marek’s disease virus. The best characterized oncogene is the  Meq  protein, which inhibits tumor suppressor genes and stimulates expression of proteins important for cell growth (IL-2, Bcl-2, CD30). It also binds to the promoter of and stimulates the expression of micro RNA21, which subsequently causes expression of metalloproteinases required for tissue invasion by tumor cells. COMPLEX TUMOR VIRUSES 25

26. Oncogenic Hepadnaviruses Mammalian, but not avian, hepadnaviruses are associated strongly with naturally occurring hepatocellular carcinomas in their natural hosts. Woodchucks that are chronically infected with woodchuck hepatitis virus almost inevitably develop hepatocellular carcinoma, even in the absence of other carcinogenic factors. Ground squirrel and woodchuck hepatitis viruses activate cellular oncogenes, the mode of action of human hepatitis B virus is uncertain, as it apparently has no consistent site of integration or oncogene association. 26

27. 27 Herpesviridae E H V 2 H V S HHV8 EBV HSV1 HSV2 E H V 1 P R V HH V 6 V Z V A L P H A H E R P E S V I RU S E S H C M V BET A H E R P E S V I RU S E S HH V 7

28. 28

29. Introduction Enveloped , spherical to pleomorphic , 150-200 nm in diameter, T=16 icosahedral symmetry . Capsid consists of 162 capsomers and is surrounded by an amorphous asymmetrical tegument . Glycoproteins complexes are embed in the lipid envelope . 29

30. Double Stranded Genome of Herpesviridae Monopartite , linear, dsDNA genome of 120-240 kb. The genome contains terminal and internal reiterated sequences. 30

31. GENE EXPRESSION Each viral transcript usually encodes a single protein and has a promoter/regulatory sequence, a TATA box, a transcription initiation site, a 5' leader sequence of 30-300 bp , a 3 ' sequence of 10-30 bp , and a poly A signal. There are many gene overlaps. There are only few spliced genes. Some of the expressed ORFs are antisense to each other. Some ORFs can be accessed from more than one promoter. Certain proteins are downregulated translationaly by a leaky scanning from an upstream ORF. 31

32. replication of circular viral episome in tandem with the host cell DNA using the host cell replication machinery. L atent replication 32

33. 33

34. Host-virus interaction Adaptive immune response inhibition Herpesviruses have evolved different strategies to inhibit the host adaptive immune response. For example, Herpes simplex protein US12 binds specifically to transporters associated with antigen processing (TAP), blocking peptide-binding to TAP and subsequent loading of peptides onto MHC class I molecules. HCMV instead encodes a protein termed US3 that directly binds and inhibits host tapasin . 34

35. Apoptosis modulation Apoptosis is very often modulated by herpesviridae . The mechanisms used can be  caspase -dependent such as HCMV vICA that prevents host caspase-8 activation, or can involved the inhibition other cellular proteins involved in apoptosis such as EBV protein BHRF1, a viral homologue of the Bcl-2 that protect the infected cell against apoptosis . Autophagy modulation Several herpesvirus are able to inhibit host cell autophagy  process, such as HHV-1 ICP34.5 that interacts with Beclin-1 and stop autophagosomes development. 35

36. Cell-cycle modulation The UL24 protein that is present in all herpesvirus subfamilies induces a cell cycle arrest at G2/M transition through inactivation of the host cyclinB /cdc2 complex  . Innate immune response inhibition Herpes viruses inhibit the cascade leading to production of interferon-beta by mainly targeting the host IRF3 protein. Thus, herpes simplex virus, varicella virus, or HCMV all possess proteins to prevent IRF3 activation . Host splicing inhibition HSV-1 ICP27 is an alternative splicing regulator of host mRNA. This protein is conserved in several herpesviridae genera. It has been shown to act as a splicing silencer at the 3' splice site of the PML intron 7a  . 36

37. Human Herpesvirus (HHV) classification Name Synonym Subfamily Primary Target Cell Pathophysiology Site of Latency Means of Spread HHV‑1 Herpes simplex virus-1 (HSV-1) α (Alpha) Mucoepithelial Oral  or  genital herpes (predominantly orofacial ), as well as other herpes simplex infections Neuron Close contact (oral or sexually transmitted infection) HHV-2 Herpes simplex virus-2 (HSV-2) α Mucoepithelial Oral or genital herpes (predominantly genital), as well as other herpes simplex infections Neuron Close contact (oral or sexually transmitted disease) HHV-3 Varicella zoster virus  (VZV) α Mucoepithelial Chickenpox and shingles Neuron Respiratory and close contact (including sexually transmitted disease) 37

38. Name Synonym Subfamily Primary Target Cell Pathophysiology Site of Latency Means of Spread HHV-4 Epstein–Barr virus (EBV),  lymphocryptovirus γ (Gamma) B cells and epithelial cells Infectious mononucleosis,  Burkitt's lymphoma, CNS lymphoma in AIDS patients, post-transplant lymphoproliferative syndrome (PTLD), nasopharyngeal carcinoma, HIV-associated hairy leukoplakia B cell Close contact, transfusions, tissue transplant, and congenital HHV-5 Cytomegalovirus (CMV) β (Beta) Monocytes and  epithelial cells Infectious mononucleosis-like syndrome, retinitis Monocyte , and ? Saliva, urine, blood, breast milk HHV-6A and 6B Roseolovirus , Herpes lymphotropic virus β T cells and ? Sixth disease  ( roseola infantum  or  exanthem subitum ) T cells and ? Respiratory and close contact? 38

39. Name Synonym Subfamily Primary Target Cell Pathophysiology Site of Latency Means of Spread HHV-7 β T cells and ? drug-induced hypersensitivity syndrome, encephalopathy, hemiconvulsion - hemiplegia -epilepsy syndrome, hepatitis infection, postinfectious myeloradiculoneuropathy, pityriasis rosea , and the reactivation of HHV-4, leading to "mononucleosis-like illness" T cells and ? ? HHV-8 Kaposi's sarcoma-associated herpesvirus (KSHV), a type of  rhadinovirus γ Lymphocyte and other cells Kaposi's sarcoma, primary effusion lymphoma, some types of multicentric   Castleman's disease B cell Close contact (sexual), saliva? 39

40. H er p esvirus e s in wh i c h V a cc i ne s a r e o f S ig n ifi c a nce Herpesvirus Abbr Host Species Commercial Vaccine Marek disease virus MDV Chicken MLV, killed Bovine herpesvirus 1 BHV-1 Cattle MLV, killed Suid herpesvirus 1 (pseudorabies virus) SHV-1 (PRV) Swine MLV, killed Equine herpesvirus 1 EHV-1 Horse MLV, killed Feline herpesvirus FHV-1 Cat MLV, killed

41. E vo lut i o n o f H er p es vi r u s V a c c i n e s 1 st G ener a t i o n  - C o n v e n t i onal k i lle d - - M o di f ie d- li v e e. g . P R - V a c , P s e ud o v ax 2 n d G e n e r a t i o n  3 rd G e n e r a t i o n  4 th Ge n e r a t i o n  ? ? G e n e -d e le t ed , i . e . vir ul e n c e g e n es e .g. O m n i V a c - 1 G ene -d e le t ed , i . e . d i f f er e nt i al m a r k e r e .g. P R V-m ar k e r , To l v id O m ni v ac I I M u l t ipl e m ar k er ge n es e .g. P R V- G ol d

42. Globally, almost 20% of cancers are related to infection agents. Several viruses with oncogenic potential stimulate cell proliferation and cause tumors and cancer in animals and humans. They act with different mechanisms depending on different factors. The tumor viruses with small genomes integrate into host cell chromosomal DNA and cause mutations and chromosomal rearrangements that predispose to cancer. The oncogenic DNA and RNA viruses that are carrying oncogenes encode transforming proteins to stimulate tumor formation. 42

43. Murat ŞEVİK, Laboratory of Molecular Microbiology, Veterinary Control Institute, Konya – TURKEY, Oncogenic viruses and mechanism of oncogenesis , 2012; 36(4): pg. no.323-329. Parkin , Donald Maxwell (2006). "The global health burden of infection-associated cancers in the year 2002". International Journal of Cancer, 118 (12): pg. no.3030–44. Oncovirus , from wikipedia , the free encyclopedia. Arbuthnot P, Kew M. 2001  Int. J. Exp. Path.  82: 77–100 Astori G,  Lavergne D, Benton C,  Hockmayr B,  Egawa K,  Garbe C, de Villiers E-M. 1998  J. Invest. Dermatol .  110: 752–755 Bachmann A,  Hanke B,  Zawatzky R, Soto U, van Riggelen J,  zur Hausen H,  Rösl F. 2001  J. Virol .  in press Pathogenesis of Viral Infections and Diseases In  Fenner's Veterinary Virology (Fifth Edition), 2017 Dr Richard Hunt (Professor, Department of Pathology, Microbiology and Immunology), Virology - Chapter Six  (Part One), Oncogenic Viruses -DNA Tumor Viruses, University of South Carolina School of Medicine 43

44. 44