Laboratory of Molecular Microbiology

Christine A. Kozak, Ph.D.

Chief, Viral Biology Section, LMM

Description of Research Program

The Viral Biology Section has a long-standing interest in the identification and characterization of genes that affect susceptibility to mouse leukemia viruses (MLVs) and to MLV-induced disease. The various inbred strains of laboratory mice and wild mouse species differ in their susceptibility to MLVs, and multiple genes are responsible for these differences. We are specifically interested in those genes that inhibit early stages of the viral replicative cycle. For these studies, we use wild mouse species as well as inbred strains of laboratory mice because the common inbred strains were derived from common progenitors and are therefore not representative of the range of genetic diversity within Mus.

Entry Restrictions

Retroviruses enter cells through interaction with specific cell surface receptors. This virus-receptor interaction defines host range, contributes to pathogenesis, and can provide the basis for the evolution of restriction variants that enable natural populations to evade retrovirus infection. Entry can be blocked by mutations in the receptor gene or by host resistance factors that interfere with receptor function. Our projects have identified the following:

• Five naturally occurring functional variants of the XPR1 receptor for the pathogenic polytropic and xenotropic MLV host range groups
• Several laboratory mouse strains that are susceptible to “xenotropic” MLVs, viruses originally defined by their failure to infect mouse cells
• Critical amino acids that form the XPR1 receptor determinants and critical sites in the ecotropic MLV envelope responsible for altered host range and receptor-induced formation of multicleated syncytia
• Roles for receptor and virus-linked glycans in host range and cytopathicity
• Endogenous retrovirus insertions at two restriction loci, Rmcf and Rmcf2, which block exogenous virus infection by receptor interference

Post-entry Restrictions

The prototype for factors that interfere with post-entry stages of virus replication is the Fv1 gene, discovered 40 years ago in studies on resistance for Friend MLV and shown to be a co-opted endogenous retrovirus. We have determined that Fv1 is present in three of the four Mus subgenera demonstrating that the gene was acquired about seven MYA. Using phylogenetic sequence analysis, we found evidence for positive selection of this gene throughout Mus evolution. We also identified novel restriction alleles and identified specific codons under positive selection. These evolutionarily important codons include two known to contribute to Fv1 resistance in laboratory mice and three codons clustered in an Fv1 segment homologous to the retroviral region known to be involved in capsid multimerization. This suggests that Fv1 inhibition may result from its binding to virus capsids.

More recent studies on the mouse APOBEC3 gene have determined that this antiviral factor is polymorphic for the presence of an inserted sequence that represents the long terminal repeat of the xenotropic MLV. This sequence is found in the more antiviral allele and is associated with elevated APOBEC3 expression levels.  We also showed that APOBEC3 has been under positive selection, and six of the positively selected codons specify residues that lie in two clusters that are opposite each other along the predicted active site groove; one of these clusters corresponds to a substrate recognition loop identified in human APOBEC. Most recently, we have examined the genetic basis for the inclusion of the extra exon 5 in some APOBEC3 alleles.  Inclusion of this exon occurred relatively late in Mus evolution and is associated with reduced antiviral activity suggesting that the cost of maintaining an effective but mutagenic enzyme may outweigh its antiviral function in some mouse populations.

Selected Publications

(View list in PubMed.)

Kozak CA. The mouse “xenotropic” gammaretroviruses and their XPR1 receptor. Retrovirology. 2010 Nov 30;7:101.

Baliji S, Liu Q, Kozak CA. Common inbred strains of the laboratory mouse that are susceptible to infection by mouse xenotropic gammaretroviruses and the human-derived retrovirus XMRV. J Virol. 2010 Dec;84(24):12841-9.

Yan Y, Liu Q, Wollenberg K, Martin C, Buckler-White A, Kozak CA. Evolution of functional and sequence variants of the mammalian XPR1 receptor for mouse xenotropic gammaretrovirsues and the human-derived retrovirus XMRV. J Virol. 2010 Nov;84(22):11970-80.

Sanville B, Dolan MA, Wollenberg K, Yan Y, Martin C, Yeung ML, Strebel K, Buckler-White A, Kozak CA. Adaptive evolution of Mus Apobec3 includes retroviral insertion and positive selection at two clusters of residues flanking the substrate groove. PLoS Pathog. 2010 Jul 1;6:e1000974.

Yan Y, Liu Q, Kozak CA. Six host range variants of the xenotropic/polytropic gammaretroviruses define determinants for entry in the XPR1 cell surface receptor.  Retrovirology. 2009 Oct 7;6:87.

Knoper RC, Ferrarone J, Yan Y, Lafont BA, Kozak CA. Removal of either N-glycan site from the envelope receptor binding domain of Moloney and Friend but not AKV mouse ecotropic gammaretroviruses alters receptor usage. Virology. 2009 Sep 1;391(2):232-9.

Yan Y, Buckler-White A, Wollenberg K, Kozak CA. Origin, antiviral function and evidence for positive selection of the gammaretrovirus restriction gene Fv1 in the genus Mus. Proc Natl Acad Sci U S A.. Proc Natl Acad Sci U S A. 2009 Mar 3;106(9):3259-63.

Stocking C, Kozak CA. Murine endogenous retroviruses. Cell Mol Life Sci. 2008 Nov;65(21):3383-98.

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