Expression of acetylcholinesterase gene(s) in the human brain: molecular cloning evidence for cross-homologous sequences.
The regulation of acetylcholinesterase (AChE) in the human brain has been approached at the level of the genome. A human DNA fragment of the length of 2 600 nucleotides was isolated from a human genomic library. This DNA fragment, designated Huache 1R, bears sequence homology to a DNA fragment from the vicinity of the Drosophila Ace region, that controls AChE biosynthesis (Soreq et al., 1985). Polyadenylated RNA from human brain was hybridized with Huache 1R DNA, eluted and microinjected into Xenopus oocytes in the absence or presence of 35S-methionine. The hybrid-selected RNA induced the biosynthesis of active AChE in the oocytes. Immunoprecipitation of labeled oocyte proteins with monoclonal antibodies against human AChE (Fambrough et al., 1982) resulted in the selective precipitation of an 85 000 Mr induced protein, with a similar size to that of the subunit of human brain AChE. These findings show that the Huache 1R DNA hybridizes with human brain AChEmRNA. The Huache 1R fragment was employed to select a collection of 12 homologous phage-cloned human genomic DNA fragments with different restriction patterns. A cDNA library in pBR322 plasmids was prepared from polyadenylated RNA isolated from embryonic brain. This library was also screened using labeled Huache 1R DNA as a probe. Forty-two out of 37 000 colonies were found positive. Several of these were selected for further analyses. Hybrid-selection experiments using DNA from two of the positive plasmid clones showed that these cDNAs also hybridize with AChEmRNA from human brain. DNA blot hybridization revealed homologies between these cDNA chains and the original Huache 1 fragment.(ABSTRACT TRUNCATED AT 250 WORDS)
D000110:Acetylcholinesterase / Q000235:genetics*; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics; D004622:Embryo, Mammalian; D004625:Embryo, Nonmammalian; D005786:Gene Expression Regulation; D006801:Humans; D009693:Nucleic Acid Hybridization; D009865:Oocytes / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism*; D014981:Xenopus
D012333:RNA, Messenger; D004247:DNA; D000110:Acetylcholinesterase