Strategies for the identification and analysis of viral immune-evasive genes--cytomegalovirus as an example.

Co-evolution of herpesviruses with their hosts has resulted in multiple interactions between viral genes and cellular functions. Some interactions control genomic maintenance and replication in specific tissues, other affect the immune control at various stages. Few immunomodulatory functions of genes can be predicted by sequence homology. The majority of genes with immunomodulatory properties only become apparent in functional assays. This chapter reviews procedures which have been used for successful identification of immunomodulatory genes in the past and deals with recent methods which may be applicable for the identification of additional immunomodulatory functions unknown so far.

The amyloid precursor-like protein 2 associates with the major histocompatibility complex class I molecule K(d).

Amyloid precursor-like protein 2 (APLP2) is a member of a protein family related to the amyloid precursor protein, which is implicated in Alzheimer's disease. Little is known about the physiological function of this protein family. The adenovirus E3/19K protein binds to major histocompatibility complex (MHC) class I antigens in the endoplasmic reticulum, thereby preventing their transport to the cell surface. In cells coexpressing E3/19K and the MHC K(d) molecule, K(d) is associated with E3/19K and two cellular protein species with masses of 100 and 110 kDa, termed p100/110. Interestingly, p100/110 are released from the complex upon the addition of K(d)-binding peptides, suggesting a role for these proteins in peptide transfer to MHC molecules. Here we demonstrate by microsequencing, reactivity with APLP2-specific antibodies, and comparison of biochemical parameters that p100/110 is identical to human APLP2. We further show that the APLP2/K(d) association does not require the physical presence of E3/19K. Thus, APLP2 exhibits an intrinsic affinity for the MHC K(d) molecule. Similar to the binding of MHC molecules to the transporter associated with antigen processing, complex formation between APLP2 and K(d) strictly depends upon the presence of beta(2)-microglobulin. Conditions that prolong the residency of K(d) in the endoplasmic reticulum lead to a profound increase of the association and a drastic reduction of APLP2 transport. Therefore, this unexpected interplay between these unrelated molecules may have implications for both MHC antigen and APLP2 function.

Cytomegaloviral control of MHC class I function in the mouse.

Cytomegaloviruses (CMVs) represent prototypic viruses of the beta-subgroup of herpesviruses. Murine cytomegalovirus (MCMV) infects mice as its natural host. Among viruses, CMVs have evolved the most extensive genetic repertoire to subvert MHC class I functions. To date three MCMV proteins have been identified which affect MHC I complexes. They are encoded by members of large virus-specific gene families located at either flanking region of the 235 kb MCMV genome. The MHC I subversive genes belong to the early class of genes and code for type I transmembrane glycoproteins. The m152-encoded 37/40 kDa glycoprotein interacts with MHC I transiently and retains class I complexes in the endoplasmic reticulum (ER) Golgi intermediate compartment on its journey to the endolysosome. In contrast, the m06-encoded glycoprotein of 48 kDa complexes tightly with ternary MHC class I molecules in the FR. Due to sorting signals in its cytoplasmic tail, gp48 redirects MHC I to endolysosomal compartments for proteolytic destruction. Likewise, the 34 kDa glycoprotein encoded by m04 binds tightly to MHC class I complexes in the ER but the gp34/MHC I complex reaches the plasma membrane. The CD8+ T-cell-dependent attenuation of a m152 deletion mutant virus proves for the first time that inhibition of antigen presentation is indeed essential for the biological fitness of CMVs in vivo.