Adenoviruses

Introduction:

Adenoviruses are a frequent cause of acute upper respiratory tract (URT) infections, i.e. "colds". In addition, they also cause a number of other types of infection. They were first isolated in 1953 by investigators trying to establish cell-lines from adenoidal tissue of children removed during tonsillectomy and from military recruits with febrile illness.
In 1962, some Adenoviruses were shown to cause tumours in rodents - this caused a considerable panic ! (N.B. Adenovirus oncogenesis appears to be associated with abortive infections and has never been observed in humans.)
During investigation of the Adenovirus genome and gene expression, many techniques were developed which were subsequently used to examine other viruses/cellular genes - these viruses are an important model system for the understanding of many other viruses. Some characteristic features of Adenoviruses are:

Taxonomy:

Haemagglutination:
Sub-Group: Types: Oncogenic Potential: Rhesus:Rat:
A12, 18, 31HIGH-+ / -
B3, 7, 11, 14, 21, 34, 35Weak+-
C1, 2, 5, 6None-+ / -
D8-10, 13, 15, 17, 19, 20, 22-30, 32, 33, 36-39None+ / -+
E4None-+ / -
F - G40, 41?-+ / -

Morphology:


There are at least 10 proteins in the Adenovirus capsid:
Name: Location: Known Functions:
IIHexon monomerStructural
IIIPenton basePenetration
IIIaAssociated with penton basePenetration
IVFibreReceptor binding; haemagglutination
VCore: associated with DNA & penton baseHistone-like; packaging?
VIHexon minor polypeptideStabilization/assembly of particle?
VIICoreHistone-like
VIIIHexon minor polypeptideStabilization/assembly of particle?
IXHexon minor polypeptideStabilization/assembly of particle?
TPGenome - Terminal ProteinGenome replication
N.B. There is no protein I (!)

All Adenovirus particles are similar: non-enveloped, 60-90nm diameter. They have icosahedral symmetry easily visible in the electron microscope by negative staining and are composed of 252 capsomers: 240 "hexons" + 12 "pentons" at vertices of icosahedron (2-3-5 symmetry).
Individual protomers can be isolated by progressive chemical disruption of purified virus particles. The hexons consist of a trimer of polypeptide II with a central pore; VI, VIII and IX are minor polypeptides also associated with the hexon, thought to be involved in stabilization and/or assembly of the particle. The pentons are more complex; the base consists of a pentamer of peptide III, 5 molecules of IIIa are also associated with the penton base. The pentons have a toxin-like activity, purified pentons causing c.p.e. in the absence of any other virus components (a unique property). A thin glycoprotein fibre (IV) protrudes form the centre of each penton - responsible for haemagglutination (the RBC receptor molecule is not known).

To view a negatively-stained electron micrograph of adenovirus particles, click here. The thin fibres protruding from each vertex of the icosahedral particle are just visible (look closely!) and the triangular faces of the icosahedral particle can be made out.
Image reconstruction of a type 2 adenovirus particle.

The core of the particle contains at least 4 proteins:

Genome:

 Linear, non-segmented, d/s DNA, 30-38kbp (size varies from group to group) which has the theoretical capacity to encode 30-40 genes. Genome structure (cross-hybridization, restriction map) is one of the characters used to assign viruses to groups (70-95% homology within groups, 5-20% homology between groups).

Replication:

 Replication of all Adenoviruses is similar and occurs in the NUCLEUS:

Replication is divided into EARLY and LATE phases, the latter defined as beginning with the onset of DNA replication (N.B. this division is characteristic of the replication of DNA viruses!). Attachment to cells is rather slow, taking several hours to reach a maximum.

Uptake of the adenovirus particle is a two stage process involving an initial interaction of the fibre protein with a range of cellular receptors, which include the MHC class I molecule & the coxsackievirus-adenovirus receptor. The penton base protein then binds to the integrin family of cell surface heterodimers allowing internalisation via receptor-mediated endocytosis. Most cells express primary receptors for the adenovirus fibre coat protein, however internalisation is more selective.

PENETRATION involves phagocytosis into phagocytic vacuoles, after which the toxic activity of the pentons is responsible for rupture of the phagocytic membrane and release of the particle into the cytoplasm.

Uncoating follows an ordered sequence, first the pentons, releasing a spherical, partially uncoated particle into the cytoplasm. The core migrates to the nucleus where the DNA enters through nuclear pores, whereupon it is converted into a virus DNA-cell histone complex.

Gene Expression:


Before and independently of genome replication, immediate early and early mRNAs are transcribed from the input DNA. Transcription of the Adenovirus genome is regulated by virus-encoded trans-acting regulatory factors. Products of the immediate early genes regulate expression of the early genes. Early genes are encoded at various locations on both strands of the DNA (l="leftward strand" and r="rightward strand"). Multiple protein products are made from each gene by alternative splicing of mRNA transcripts - splicing was first discovered in Adenoviruses (Sharp, 1977).

Phase: Genes Transcribed:
Immediate earlyE1A
EarlyE1B, E2A, E2B, E3, E4, some virion proteins
LateLate genes, mostly virion proteins

The first mRNA/protein to be made (~1h after infection) is E1A. This protein is a trans-acting transcriptional regulatory factor whose precise mode of action is not known (not a DNA-binding "transcription factor") but is necessary for transcriptional activation of early genes. The protein is also capable of activating transcription from a variety of other viral and cellular promoters and shows no sequence-specificity, rather a modification of the cellular environment.
The second protein made is E1B. E1A + E1B together (and independently of other virus proteins) are capable of transforming primary cells in vitro (especially Ad5, Ad12).
Transformation is: "A CHANGE IN THE MORPHOLOGICAL, BIOCHEMICAL OR GROWTH PARAMETERS OF THE CELL", which may or may not result in cells which are able to produce tumours in experimental animals ( = NEOPLASTIC transformation).

The activities of the two proteins can be dissected out by molecular techniques:

E1A: Can immortalize primary cells in vitro.
E1B: Does not transform cells on its own, but "co-operates" with E1A to stably transform cells.
E1A + E1B: Necessary for full transformation and tumour formation in animals.
E1A has also been shown to bind a cellular protein, p105-RB, the product of the retinoblastoma gene (retinoblastomas result when this gene is deleted or damaged, hence it is an "anti-oncogene" or "tumour suppressor"). E1B binds to p53, another tumour suppressor involved in the control of the cell cycle. N.B. Binding of DNA virus nuclear proteins to cellular tumour suppressors is a shared mechanism of cell transformation found in several virus families.
Together, these observations indicate that Adenoviruses, in the course of sequestering cellular machinery and altering the intracellular environment to favour viral replication, have profound effects on cellular functions. Viewed in this light, transformation is just an accidental (and rare) outcome of infection. The basis for oncogenesis (c.f. immortalization of cells in vitro - above) is not clear, but it is known that Ad12 E1A turns off class I MHC expression, possibly allowing tumours to escape destruction by CTLs.

DNA Replication:


Adenovirus DNA replication has been studied extensively both in vivo (t.s. mutants in infected cells) and in vitro (nuclear extracts). At least 3 virus-encoded proteins are known to be involved in DNA replication: In addition, many cellular proteins in the nucleus also participate in replication of the genome (e.g. NFI, NFII, topoisomerase I).

Late Transcription:

At the onset of DNA replication, the pattern of transcription changes radically from the early to the late genes. There is cis-acting control of this switch, i.e. only newly replicated DNA is used for late gene transcription, but the mechanism controlling this is not understood. The late genes are transcribed from the major late promoter; at least 13 species of mRNA are produced by alternative splicing.
Assembly occurs in the nucleus, but begins in the cytoplasm when individual monomers form into hexon and penton capsomers. Empty, immature capsids are assembled from these protomers in the nucleus, where the core is formed from genomic DNA + associated core proteins. Although host cell macromolecular synthesis ceases earlier in the infection, infected cells remain intact and do not lyse (disruption of cytoskeleton changes shapes of cell - rounds up). Virus particles tend to accumulate in the nucleus and are visible in the microscope as eosinophilic crystals - INCLUSION BODIES. These are thought to be the basis of latent infections - reactivation is caused by accidental lysis of infected cells, releasing virus particles from the nuclei - effectively a re-infection. More properly, this type of mechanism of persistence is known as "occult" (hidden) infection, rather than "latent" (c.f. Herpesviruses).
Adenoviruses are known to interact with other viruses, notably defective Parvoviruses (Adeno Associated Viruses). SV40 (Papovavirus) has also been shown to act as a helper virus for Adenoviruses - co-infection overcomes a late block to Adenovirus replication in certain cell types which are normally non-permissive. This may involve functional substitution of SV40 T-antigen for Ad DBP (?). Infectious Adeno-SV40 hybrids have been isolated by rescuing Adenovirus deletion mutants by super-infecting with SV40, indicating functional overlap between the two families.

Pathogenesis:

 Certain types of Adenovirus are commonly associated with particular clinical syndromes:

Disease: At Risk:
Acute Respiratory Illness Military recruits, boarding schools, etc.
Pharyngitis Infants
Gastroenteritis Infants
Conjunctivitis All
Pneumonia Infants, military recruits
Keratoconjunctivitis All
Acute Haemorrhagic Cystitis Infants
Hepatitis Infants, liver transplant patients

Note that this list includes relatively common infections (at the top) and some rare infections (bottom). Most Adenovirus infections involve either the respiratory or gastrointestinal tracts or the eye.
Adenovirus infections are very common, most are asymptomatic. Most people have been infected with at least 1 type at age 15. Virus can be isolated from the majority of tonsils/adenoids surgically removed, indicating latent infections. It is not known how long the virus can persist in the body, or whether it is capable of reactivation after long periods, causing disease (it is hard to isolate this occult virus as it may be present in only a few cells). It is known that virus is reactivated during immunosuppression, e.g. in AIDS patients.

Therapy:

None. Inactivated vaccines have been developed and are routinely used for military recruits in some countries (e.g. USA). This is because adolescents and others in close daily contact are at risk for epidemic spread of respiratory infections - risk to general population is so low that vaccination is not a viable proposition.
In recent years, there has been considerable interest in developing Adenoviruses as defective vectors to carry and express foreign genes for therapeutic purposes. One reason for this is that the Adenovirus genome is relatively easily manipulated in vitro (c.f. Retroviruses) and the genes coupled to the MLP are efficiently expressed in large amounts.

Return to BS335 HomePage

© AJC 1997