Background

I.  Human Adenoviruses:  Clinical Aspects - 30 strains classified into 5 serogroups

    A.  Serogroup A

        1.  found in feces of healthily individuals

        2.  high oncogenicity in hamsters

    B.  Serogroup B

        1.  upper respiratory tract infections

        2.  weak oncogenecity

    C.  Serogroup C

        1.  isolated from respiratory and GI tracts of kids with mild respiratory disease

        2.  can remain latent in lymphoid tissue

        3.  not oncogenic

    D.  Serogroup D

        1.  Ad8 and Ad19 cause acute epidemic keratoconjunctivitis and pharyngoconjunctival fever

        2.  remaining members of this serotype are not associated with disease

        3.  not oncogenic

    E.  Serogroup E

        1.  severe acute respiratory disease

        2.  not oncogenic

II.  Virion Structure

    A.  Icosahedron with fibers and knobs at each of the vertices and made up of 15 proteins

        1.  Hexons-240 per facet, each made up[ of three copies of 120 kDa protein

        2.  Pentons- one at each of the vertices (12), each made up of 5 copies of a 70 kDa protein

        3.  Fibers-  62 kDa protein

        4.  Internal proteins

            a.   basic protein binds and neutralizes negative charge on DNA phosphate groups

            b.   one organizes DNA into a multi-lobed form and binds the DNA to the 12 pentons

            c.   one is the terminal protein

    B.  DNA 36,000 bp- encodes 20 proteins

        1.  has inverted terminal redundancy - ss lollipop configuration

        2.  has covalently linked terminal protein at each 5' terminus

III.  Adenovirus multiplication

    A.  Adsorption-  slow, maxes out after several hours

    B.  Penetration- receptor mediated endocytosis.   Virion escapes endocytic vesicle.

    C.  Uncoating - in cytoplasm

        1.  pentons and surrounding hexons removed

        2.  hexons an associated proteins removed

        3.  naked virus core enters nucleus through nuclear pore

    D.  Early mRNA synthesized beginning at about 3 hours post infection

    E.  DNA replication begins in the nucleus at about 6-8 hours post-infection

        1.  reaches a maximum at 18-20 hr p.i.

        2.  stops 22-24 hr p.i.

        3.  Major pathway:   Initiation of DNA replication may occur at either termini

            a.   No RNA primer is required.  Primer is a 55 kDa protein (TP) at 5' termini

            b.   TP is 80 kDA protein when covalently linked to 5' ends.  Then cleaved by protease.

            d.  The 80 kDA protein interacts with the 55 kDa TP allowing the C to hydrogen bond to G and generate
                  primer structure.

            e.  Chain growth proceeds from 3'-OH of dCTP and the 80 kDa is cleaved

            f.  In the major pathway, type I intermediates are formed.  Growth of the daughter strand continues to
                the other end of  the molecule (5'-3' continuous, leading strand replication).  The daughter strand
                displaces the other parental strand.

            g.   Complementary strand synthesis begins at the 3' end of the displaces strand, producing type II
                  intermediates.  Initiation probably occurs on a panhandle structure.  The ds terminus of such as
                  structure would have all the same  features as the ds terminus on the original ds DNA.

There is no lagging strand synthesis as a result of this type of DNA replication.   Replication of each strand involves only leading strand, continuous DNA synthesis.

        4.  Minor Pathway:  initiation at both termini (probably does not occur often)

            a.  replication proceeds until the middle of the molecule is reached

            b.  at that time, the two replicating molecules would fall apart into 2 type II replication intermediates

    G.  Late transcription

        1.  Begins shortly after DNA replication- which it requires

        2.  Occurs primarily on the r strand.   All mRNA are generated from one long primary transcript that is
             spliced into many  mRNA molecules.  Each mRNA consists of the leader sequence (nontranslated DNA
             found upstream of the coding  region and which contains regulatory elements) spliced to
             non-contiguous coding region.

            a.  promoter is at map unit 16.4 and the transcripts span most of the genome

            b.  processing regulates the amount of each of the late mRNAs and thus the amount of each protein.  
                 Different splice  sites are preferentially used

            c.  Early transcription is still occurring but only 30% are expressed

    H.  Assembly

        1.  DNA and Rna are made in the nucleus and virions are assembled there.  Late structural proteins are
             produced on cytoplasmic ribosomes and transported back into the nucleus.

        2.  Assembly begins 2-4 hr after capsid protein synthesis begins

IV.  Contrast between permissive and nonpermissive replication

    A.  In nonpermissive cells there is no DNA replication and no late transcription.  Infecting DNA becomes
          integrated into the  host chromosome and is replicated concomitantly with cell DNA replication. 
          Expression of the integrated genome causes cells to acquire the transformed phenotype.

        1.  Integrated adeno DNA always contains a functional copy of E1A.  Thus it is supposed that one of its
             two products  must be a tumor antigen.

        2.  The adenovirus genome is "destroyed" upon integration-most of it is lost, leaving the E1A early region
             intact.