Download “Ancient” Viruses

Gene Expression and
Replication in Medium DNA
Viruses
Phage Lambda
Adenovirus
What can the medium sized viruses
do that the small ones can not?
Adenovirus: more sophisticated
gene expression strategies
Lambda: two pathways for infection
Adenovirus Properties
 Adenoviruses are non-enveloped Icosahedral viruses with spike-like
projections at the vertices.
 Contain a linear double stranded DNA genome of about 36 KB that
accounts for about 13% of mass of the virus particle.
 First found in 1957 in the adenoid tissues of US Army recruits.
 More than 40 Adenovirus serotypes infect humans.
; More than 100 Adenoviruses infect mammals and birds.
 Most serotypes produce respiratory infections in humans and cause about
5 - 10 % of the common colds in children.
 Some serotypes cause intestinal infections and conjunctivitis.
 Some human adenoviruses can cause tumors in
newborn hamsters but have not been
implicated in human cancers.
 Can transform cell lines and have provided
valuable information about oncogenesis.
 Studies of adenoviruses have resulted in
numerous findings about DNA replication,
cell cycle control, mRNA splicing, and
other biochemical topics.
 Adenoviruses have also been studied for
gene therapy and cancer control.
Adenovirus Structure
Viruses are unenveloped icosahedra 70100 in diameter.
 Virions are composed of a hexon shell
and fibers at the vertices that are
used for attachment to host receptors.
 Vertices consist of 12 antenna-like fibers
called Pentons composed of pentamers
of protein III.
 Faces of the Hexon are composed of 240
capsomers each of which is a trimer of
protein II, the most abundant virus
protein.
 Capsid stabilization is provided by
Proteins IIIa and IX.
 Proteins VI and VIII bridge the shell
and the core.
 Proteins V, VII and m form the core.
 Proteins V, VII and X are arginine-rich
viral DNA binding proteins.
 The Terminal Protein is covalently
attached to the 5’ end of the genome.
 The virus particle also contains a
proteinase.
Characteristics of the Adenovirus DNA Genome
 Linear, double stranded DNA genome is about 36 kb.
 Terminal inverted repeats (100 to 140 bp) have several
functions
1) Contain origins of replication (Ori) at the 3’ termini.
2) Can mediate circularization during DNA replication.
 A 5’ terminal protein (TP) is covalently linked to the
genome by a serine OH residue.
 The terminal protein binds the genome to the nuclear
matrix to initiate DNA replication (nuclear address).
Note Terminal Repeats (Pink) at the ends of the genome.
Adenovirus Gene Expression
 More
sophisticated cascade of gene
regulation (p. 265 Fig 8.14, pp. 272-275)
 Several regulators
 Gene expression organized in several
phases
Immediate early
Early
Late
 More
than one type of cellular DdRp
involved-different classes of genes
All immediate/early genes scattered
Late genes tend to cluster
Early gene functions
Induce S phase (E1A, E4)
Mount a defense against host antiviral
response (E1A, E1B, E3, VA)
Stimulate synthesis of proteins needed for
virus DNA synthesis (E1A activates AdPol,
DBP, pTP)
Constitutive expression
E1A generates 2 proteins called large and
small or 13S and 12S or E1A and e1A
E1A proteins are expressed before any
other viral proteins from a constitutive viral
promoter/enhancer.
E1A proteins have many regulatory
functions.
E1A Expressed First
Cellular promoter, enhancer
Alternate splicing: 2 forms
CR3 stimulates early transcription-interaction with cell
transcription initiation complex
CR1, CR2 interact with tumor suppressor pRB and with other
transcription components
Allows expression of Ad E2 genes for DNA replication
The E1A proteins are
promiscuous transactivators
Large and small E1A proteins interact with
many other proteins in an infected cell.
The interactions influence host cell
processes and activities
E1A Proteins Have Main
Responsibilty For
1)
2)
3)
Stimulate host cell to move into S phase
Help protect virus from host defense
Boost expression of replication-related
virus genes
1) Stimulation of S Phase
E1A sequesters Rb family proteins to relieve
S Phase blocks.
E1A blocks action of CDK proteins also
causes chromatin remodeling.
E1A activates E4 which in turn inactivates
different S blocks.
e1A can regulate many cellular promoters
2) Protection against host
defense
E1A acts indirectly to decrease p53 function
E1A activates expression of other E genes
E1B acts indirectly to degrade p53
E3 is an antagonist of cell-mediated
immunity
E4 independently inhibits p53
E4 inhibits cellular response to DNA
damage
VA RNAs block IFN response
3) Ad DNA synthesis
E1A activates expression of E2 proteins
E2 products are associated with Ad DNA
replication.
E2A codes DBP
E2B codes pTP and AdPol
E1A activates many genes
E1A activates expression of E proteins:
Targets include E1B, E2, E3, E4-and E1A
itself!
E1A appears to recruit transcription
machinery to viral promoters
CR3 domain
Adenovirus DNA Replication
Adenovirus DNA replication is a viral
protein-primed process that
occurs in the nucleus:
1. A preterminal protein (pTP)/DNA
polymerase (Pol) complex binds
to the 3’ origin of replication
using both E2 proteins.
2 dCTP is recruited to form a
phosphodiester bond with the
pTP serine.
3. Continuous 5’ to 3’ synthesis of
DNA by viral polymerase (Pol)
occurs on only one DNA strand.
4. The displaced DNA strand is
coated by DNA Binding Protein
(DBP).
5. The terminal repeats anneal to form
the origin of replication, so pTP
complexes reforms on the
displaced strand which leads to
further replication
Some adaptations of adenovirus
Still some reliance on host enzymes but some
viral enzymes used in replication too
Reliance on host control mechanisms
Functional clustering of some genes
More elaborate capsids
More elaborate control circuits
Can manipulate host cell environment