The endogenous avian provirus ev-1 is widespread in white leghorn chickens. Although it has no major structural defects, ev-1 has not been associated with any phenotype and is ordinarily expressed at a very low level. In this report, we describe a chicken embryo (Number 1836) cell culture containing both ev-1 and ev-6 which spontaneously expressed the ev-1 provirus. This culture released a high level of noninfectious virions containing a full complement of virion structural (gag) proteins but devoid of reverse transcriptase activity or antigen. These virions contained 70S RNA closely related to the genome of Rousassociated virus type 0, but identifiable as the ev-1 genome by oligonucleotide mapping. A fraction of the RNA molecules in the 70S complex were unusual in that they were polyadenylated 100 to 200 nucleotides downstream of the usual polyadenylation site. Eight sibling embryo cultures did not share this unusual phenotype with 1836, indicating that it was not inherited. However, an identical phenotype was inducible in the sibling cultures by treatment with 5-azacytidine, an inhibitor of DNA methylation, and the induced expression was stable for more than 10 generations. Analysis of chromatin structure and DNA methylation of the ev-1 provirus in 1836 cells revealed the presence (in a fraction of the proviruses) of both DNase I hypersensitive sites in the long terminal repeats and in gag and a pattern of cleavage sites for methyl-sensitive restriction endonuclease not found in a nonexpressing sibling. These results lend strong support to the role of DNA methylation in the control of gene expression. Additionally, they explain the lack of phenotype associated with ev-1 as due to a combination of its low expression and defectiveness in pol and env.The endogenous viruses of chickens, which are inherited in chromosomal DNA, are closely related to the exogenous avian leukosis proviruses (for review, see 26). At least 13 distinct endogenous proviruses have been identified by DNA restriction endonuclease mapping (3, 18). Although closely related to one another, proviruses at different endogenous virus (ev) loci differ in DNA content and transcriptional activity. Thus, they may provide a set of genes for the study of elements, both viral and host, that contribute to the control of gene expression.The nondefective endogenous proviruses have a structure similar to integrated proviral DNA found after exogenous infection. That is, they retain the gene order 5'-gag-pol-env-3' of genome RNA and are flanked by two long terminal repeat (LTR) sequences, consisting of sequences derived from the 5' (U5) and 3' (U3) ends of viral RNA, as well as a sequence (R) which is itself repeated at the termini of genome RNA. Thus the overall structure is U3-R-U5-gag-pol-env-U3-R-U5 (17, 28; for review, see 6). The U3 region contains sequences important for regulation of growth rate (39) and appears to include the promoter for viral RNA synthesis (10,21,50).The ev-1 provirus is found in more than 99% of white leghorn chickens (38) and does...
