CpG islands are important in the protection of adjacent housekeeping genes from de novo DNA methylation and for keeping them in a transcriptionally active state. However, little is known about their capacity to protect heterologous genes and assure position-independent transcription of adjacent transgenes or retroviral vectors. To tackle this question, we have used the mouse aprt CpG island to flank a Rous sarcoma virus (RSV)-derived reporter vector and followed the transcriptional activity of integrated vectors. RSV is an avian retrovirus which does not replicate in mammalian cells because of several blocks at all levels of the replication cycle. Here we show that our RSV-derived reporter proviruses linked to the mouse aprt gene CpG island remain undermethylated and keep their transcriptional activity after stable transfection into both avian and nonpermissive mammalian cells. This effect is most likely caused by the protection from de novo methylation provided by the CpG island and not by enhancement of the promoter strength. Our results are consistent with previous finding of CpG islands in proximity to active but not inactive proviruses and support further investigation of the protection of the gene transfer vectors from DNA methylation.

• MeSH
• Adenine Phosphoribosyltransferase genetics   MeSH
• Cell Line virology   MeSH
• CpG Islands * MeSH
• Defective Viruses genetics   MeSH
• DNA, Viral chemistry   genetics   MeSH
• DNA (Cytosine-5-)-Methyltransferases metabolism   MeSH
• Sarcoma, Experimental genetics   virology   MeSH
• Fibroblasts virology   MeSH
• Transcription, Genetic * MeSH
• Genetic Vectors genetics   physiology   MeSH
• Virus Integration MeSH
• Terminal Repeat Sequences MeSH
• Cricetinae MeSH
• Mesocricetus MeSH
• Chick Embryo MeSH
• DNA Methylation MeSH
• Mice MeSH
• Proviruses genetics   MeSH
• Gene Expression Regulation, Viral * MeSH
• Virus Replication MeSH
• Genes, Reporter MeSH
• Gene Silencing * MeSH
• Avian Sarcoma Viruses genetics   physiology   MeSH
• Animals MeSH
• Check Tag
• Cricetinae MeSH
• Chick Embryo MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• Adenine Phosphoribosyltransferase MeSH
• DNA, Viral MeSH
• DNA (Cytosine-5-)-Methyltransferases MeSH

The LTR, v-src, LTR provirus, which arose by the reverse transcription and integration of src mRNA in the H-19 hamster tumor, has been successfully rescued by fusion with chicken fibroblasts infected with Rous-associated virus RAV-1. One rescued virus, E6, acquired 1 kilobase of the 5' end of the gag gene structure. Recombination took place in the region of 15-nucleotide homology exactly between v-src exon (position 7054) and gag (position 1417). This recombination resulted in the alteration of src splice acceptor site sequences, but this site is maintained as a functional splice acceptor site. The nucleotide structure of the long terminal repeat of recombinant E6 virus suggests that it arose by the intermolecular jump of reverse transcription from RAV-1 to src mRNA and then the switch of templates between already depicted regions of homology. The second jump of reverse transcription was apparently an intramolecular event. The acquisition of 1 kilobase of the 5' gag by E6 resulted in maintaining the balance of unspliced and spliced E6 RNAs and assured the replication advantage of rescued E6 virus over rescued F6 virus, the genome of which corresponds to that present in ancestral H-19 cells.

• MeSH
• Genes, gag MeSH
• RNA, Messenger genetics   MeSH
• Molecular Sequence Data MeSH
• Oncogenes * MeSH
• Proviruses genetics   MeSH
• Recombination, Genetic MeSH
• Repetitive Sequences, Nucleic Acid MeSH
• RNA, Viral genetics   MeSH
• Base Sequence MeSH
• Cell Transformation, Viral genetics   MeSH
• Avian Leukosis Virus genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, U.S. Gov't, P.H.S. MeSH
• Names of Substances
• RNA, Messenger MeSH
• RNA, Viral MeSH