replication of viral dna ii - microbe.tv
TRANSCRIPT
![Page 1: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/1.jpg)
1
![Page 2: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/2.jpg)
2
![Page 3: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/3.jpg)
3
![Page 4: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/4.jpg)
DNA Replication
• Viruses must replicate their genomes to make new progeny
• This always requires expression of at least one virus protein, sometimes many (hence always delayed after infection)
• DNA is always synthesized 5’ – 3’
• Replication initiates at a defined origin (Ori) using a primer
• The host provides other proteins
4
![Page 5: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/5.jpg)
The 5’ end problem
elongate
3’ 5’
5’ 3’
RNA primers
DNA template
3’ 5’
5’ 3’
3’ 5’
5’ 3’
excise primers, elongate, ligate
Now what?5
![Page 6: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/6.jpg)
Two Basic Modes of Replication
6
![Page 7: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/7.jpg)
Where Does the DNA Polymerase Come From?
• Small DNA viruses do not encode an entire genome replication system-encode proteins that orchestrate the host-Papillomaviridae, Polyomaviridae, Parvoviridae
• Large DNA viruses encode most of their own replication systems-Herpesviridae, Adenoviridae, Poxviridae
7
![Page 8: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/8.jpg)
8
![Page 9: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/9.jpg)
Protein Priming
ITRs Single-stranded DNA Template
Displaced ss Template DNA
9
![Page 10: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/10.jpg)
Lessons from bacteriophage ϕ29
10
![Page 11: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/11.jpg)
Adenoviral DBP
11
![Page 12: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/12.jpg)
Replication of adenovirus genome
• An example of strand displacement synthesis
• Utilizes a protein primer
• Origin is at both ends
• DNA polymerase is viral
• Other viral proteins involved: terminal protein, ssDNA binding protein
• Viral early proteins (E1a) induce quiescent cells into S phase
12
![Page 13: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/13.jpg)
Adenovirus DNA replication requires cellular proteins
13
![Page 14: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/14.jpg)
14
![Page 15: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/15.jpg)
Herpes simplex virus
• HSV has 2 identical oriS and a unique oriL that is active in terminally differentiated neurons – role in transition from latent to productive infection?
• DNA enters as linear molecule converts to circle
• Replicates as a rolling circle
15
![Page 16: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/16.jpg)
16
![Page 17: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/17.jpg)
Viral proteins for Herpes simplex virus genome replication
• UL5, 8 and 52 - form primase, helicase
• UL42 - a processivity protein
• UL9 - Origin Binding Protein
• UL29 - ssDNA Binding Protein
• UL30 - DNA polymerase
• 5 enzymes of nucleic acid metabolism, such as TK
• Necessary but not sufficient!17
![Page 18: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/18.jpg)
Poxviruses
• All viruses discussed replicate in nucleus
• Poxvirus - cytoplasmic factories - DNA synthesis is independent of cellular proteins
18
![Page 19: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/19.jpg)
Vaccinia DNA factories
DNA stained blue; viral DNA binding protein (I3) stained red
DNA only I3 only merge
19
![Page 20: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/20.jpg)
Vaccinia DNA factories
DNA stained blue; viral DNA polymerase (E9) stained red
20
![Page 21: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/21.jpg)
Poxvirus DNA replication
From “Fundamentals of Molecular Virology” by Nicholas H. Acheson, John Wiley & sons, 2007
21
![Page 22: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/22.jpg)
Poxvirus DNA replication enzymes
22
![Page 23: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/23.jpg)
Delayed synthesis of virion structural proteins prevents
• When new viral DNAs are encapsidated they cannot be used as templates for additional DNA synthesis
• Consequently genome packaging into particles is delayed – transcriptional control
23
![Page 24: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/24.jpg)
Inhibition of cellular DNA synthesis
• When viral DNA replication is carried out mostly by viral proteins, cell DNA synthesis is often inhibited
• Increases availability of substrates
• Adenovirus, herpesvirus, poxvirus
• Mechanisms not understood
24
![Page 25: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/25.jpg)
Viral DNAs are synthesized in specialized intracellular
25
![Page 26: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/26.jpg)
Mechanism of exponential viral DNA replication
Cellular retinoblastoma (rb) gene: Rb protein controls entry into S from G1
Loss is associated with tumors = tumor suppressor gene26
![Page 27: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/27.jpg)
Adenovirus E1A protein binds Rb
27
![Page 28: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/28.jpg)
Limited replication of viral DNA
• Most DNA viruses: exponential replication of genomes
• Some establish long-term relationships with cells, number of genomes is limited
• Various mechanisms effect genome copy number
28
![Page 29: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/29.jpg)
Rep78/68 a site & strand-specific endonuclease
Parvovirus DNA Replication
29
![Page 30: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/30.jpg)
Parvovirus DNA replication
• DNA replicates only in cells coinfected with helper adenovirus
• Adenoviral helper proteins allow synthesis of large quantities of Rep 78/68
30
![Page 31: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/31.jpg)
Parvovirus DNA replication
• When no helper adenovirus is present, Rep 78/68 level is low
• Little viral DNA synthesis occurs
• Genome integrates into host cell DNA
31
![Page 32: replication of viral DNA II - Microbe.TV](https://reader031.vdocuments.us/reader031/viewer/2022012205/61de7a6bdc0dfb5577586186/html5/thumbnails/32.jpg)
Papillomaviruses: Controlled and exponential replication from a single
Ori
32