This proposal describes experiments that continue our work on mobile, dispersed, repeated DNA elements, their RNAs and their possible functioning in mammalian cells. The majority of the proposal focuses on two repeat families, the KpnI family and the Alu superfamily. The KpnI family has dispersed it members to approxiamtely 45,000 different loci in mammalian DNAs via an RNA intermediate having the structure of a messenger RNA. All currently known mammalian mobile DNA elements have transcripts that contain within them the sequences required for transcription, and hence newly dispersed family members are transcription competent and can thus serve as sources for the generation of still more new family members. By analogy, we suspect this might be true for KpnI transcripts, and we will attempt to determine whether this is so. We will also examine the nature of non-polysomal KpnI RNA-containing structures we found in the cytoplasm of cultured human cells. 4.5sRNA is an Alu sequence-related low molecular weight RNA with a short half line and low abundance initially isolated because it hydrogen bonds with poly(A)-terminated RNAs. We will clone from different rodent species genes encoding the 4.5s RNA and identify sequence involved in its transcription. We will use 4.5s RNA transcripts to identify and clone mRNAs with which it hydrogen bonds to obtain probes to examine their synthesis, turnover, nuclear/cytoplasmic levels and association with polyribosomes under different conditions of cell growth and differentiation. We will continue experiments we have initiated to examine the ability of Alu family members to serve as origins of DNA replication. We will attempt to observe transposition events in cultured mammalian cells using plasmids that replicate independent of host cell DNA as """"""""landing pads"""""""" for mobile DNA elements and by analyzing mutations in the chromosomal HGPRT gene that are caused by DNA insertions.
| Schoeniger, L O; Jelinek, W R (1986) 4.5S RNA is encoded by hundreds of tandemly linked genes, has a short half-life, and is hydrogen bonded in vivo to poly(A)-terminated RNAs in the cytoplasm of cultured mouse cells. Mol Cell Biol 6:1508-19 |
| Johnson, E M; Jelinek, W R (1986) Replication of a plasmid bearing a human Alu-family repeat in monkey COS-7 cells. Proc Natl Acad Sci U S A 83:4660-4 |
| Kaplan, G; Jelinek, W R; Bachvarova, R (1985) Repetitive sequence transcripts and U1 RNA in mouse oocytes and eggs. Dev Biol 109:15-24 |
