IDENTIFICATION OF A PUTATIVE DNA-REPLICATION ORIGIN IN THE GAMMA-AMINOBUTYRIC-ACID RECEPTOR SUBUNIT BETA-3 AND ALPHA-5 GENE-CLUSTER ON HUMAN-CHROMOSOME 15Q11-Q13, A REGION ASSOCIATED WITH PARENTAL IMPRINTING AND ALLELE-SPECIFIC REPLICATION TIMING

The region containing the GABA(A) receptor beta 3 and alpha 5 subunit- encoding genes is subject to parental imprinting and is organized in d ifferent allele-specific replication timing domains. A 60-kb domain di splaying a maternal early/paternal late pattern of allele-specific rep lication timing asynchrony is nested within a larger region displaying the opposite pattern. The proximal portion of this maternal early rep licating domain is incorporated into phage clone lambda 84. In order t o identify DNA structures which may be associated with the boundary be tween the replication domains, phage lambda 84 has been subcloned into smaller fragments and several of these have been analyzed by nucleoti de sequencing. A plot of helical stability for 13 kb of contiguous seq uence reveals several A+T-rich regions which display potential DNA unw inding. The plasmid subclones from phage lambda 84 have been analyzed for bent DNA and one of these, p82, contains bent DNA and overlaps wit h the region of highest potential helical instability. Of the seven pl asmids tested, only p82 shows strong autonomous replication activity i n an in vitro replication assay, with replication initiating within th e genomic insert. These results suggest that a putative origin of DNA replication contained within p82 may play a role in establishing the a llele-specific replication timing domains in the GABA(A) receptor subu nit gene cluster.