The Ly-5 system of the mouse composes a group of transmembrane glycoprotein isoforms that range in size from Mr 200 to 220 kDa and are expressed selectively by different lineages and differentiational stages of hematopoietic cells, including T and B lymphocytes. The main objective is to relate alternative Ly-5 exons to particular Ly-5 structural epitopes as a means to relate the differential structure of Ly-5 isoform to their functions in different Ly-5+ cell lineages. Use of exon-specific probes, derived from genomic DNA as before, to determine the representation of selective exons and isoforms epitopes in particular cell sets and lineages, will be extended to include further Ly-5+ cells populations and differentiational stages. It is proposed to use already obtained cDNA and genomic DNA in conjunction with retrovirus vector to construct cDNAs which on transfection may give rise to soluble secreted Ly-5 isoforms representing chosen selective exons and exon combinations; one application of such purified soluble isoforms, among several, would be competitive inhibition in functional assays, as another means of relating particular isoform epitopes to particular functions, immunological and non-immunological. Definition of two putative promoters and identification of regulatory sequences controlling Ly-5 expression are planned on the basis of the CAT assay system in vivo and transcriptional assay in vitro. As a first approach to elucidating control of Ly-5 exon selection and alternative splicing, construction of a variety of minigenes from the critical region of Ly-5 genomic DNA is proposed, each to transfected into a range of cell lines representing the non- expressor (Ly-5-) phenotype and various distinct isoform expressor (Ly-5+) phenotypes, for evaluation and interpretation of the details of resulting Ly-5 expression.
| Saga, Y; Furukawa, K; Rogers, P et al. (1990) Further data on the selective expression of Ly-5 isoforms. Immunogenetics 31:296-306 |
| Saga, Y; Lee, J S; Saraiya, C et al. (1990) Regulation of alternative splicing in the generation of isoforms of the mouse Ly-5 (CD45) glycoprotein. Proc Natl Acad Sci U S A 87:3728-32 |
