SEQUENCE-BASED EXON PREDICTION AROUND THE SYNAPTOPHYSIN LOCUS REVEALSA GENE-RICH AREA CONTAINING NOVEL GENES IN HUMAN PROXIMAL XP

The human Xp11.23-p11.22 interval has been implicated in several inher ited diseases including Wiskott-Aldrich syndrome; three forms of X-lin ked hypercalciuric nephrolithiaisis; and the eye disorders retinitis p igmentosa 2, congenital stationary night blindness, and Aland Island e ye disease. In constructing YAC contigs spanning Xp11.23-p11.22, we ha ve previously shown that the region around the synaptophysin (SYP) gen e is refractory to cloning in YACs, but highly stable in cosmids. Prel iminary analysis of the latter suggested that this might reflect a hig h density of coding sequences and we therefore undertook the complete sequencing of a SYP-containing cosmid. Sequence data were extensively analyzed using computer programs such as CENSOR (to mask repeats), BLA ST (for homology searches), and GRAIL and GENE-ID (to predict exons). This revealed the presence of 29 putative exons, organized into three genes, in addition to the 7 exons of the complete SYP coding region, a ll mapping within a 44-kb interval. Two genes are novel, one (CACNA1F) showing high homology to alpha 1 subunits of calcium channels, the ot her (LMO6) encoding a product with significant similarity to LIM-domai n proteins. RT-PCR and Northern blot studies confirmed that these loci are indeed transcribed. The third locus is the previously described, but not previously localized, A4 differentiation-dependent gene. Given that the intron-exon boundaries predicted by the analysis are consist ent with previous information where available, we have been able to su ggest the genomic organization of the novel genes with some confidence . The region has an elevated GC content (>53%), and we identified CpG islands associated with the 5' ends of SYP, A4, and LMO6. The order of loci was Xpter-A4-LMO6-SYP-CACNA1F-Xcen, with intergenic distances ra nging from similar to 300 bp to similar to 5 kb. The density of transc ribed sequences in this area (>80%) is comparable to that found in the highly gene-rich chromosomal band Xq28. Further studies may aid our u nderstanding of the long-range organization surrounding such gene-enri ched regions. (C) 1997 Academic Press.