Structure and regulation of acetylcholinesterase gene.

摘要

cDNA clones encoding mouse acetylcholinesterase and butyrylcholinesterase were isolated from a mouse brain cDNA library. Transfection of clones encoding both enzymes in COS cells under the control of a SV40 or CMV promoter produced active catalytic subunits which are similar to those of the hydrophilic species of acetylcholinesterase in Torpedo and butyrylcholinesterase in human.; The genes encoding mouse and human acetylcholinesterases have been cloned from genomic libraries. Restriction analysis and a comparison of sequence with the cDNA defined exon-intron boundaries. mRNA protection studies how that the cDNA encoding the hydrophilic catalytic subunits represents the dominant mRNA species in mammalian nerve and muscle while divergent mRNA species are evident in cells of hematopoietic origin (bone marrow cells and an erythroleukemia cell line).; Analyses of mRNA species and the genomic sequence have enabled me to define two alternative exons in addition to the one found in the cDNAs; these exons encode unique carboxyl-terminal sequences in acetylcholinesterase. One mRNA consists of a direct extension through the intervening sequence between the common exon and the 3{dollar}spprime{dollar} exon in cDNA. Another mRNA encodes glycophospholipid-linked species of the acetylcholinesterase by using an alternatively spliced exon. A cDNA for the putative glycophospholipid-linked form was constructed by loop-out mutagenesis. An active enzyme expressed with the loop-out construction in COS cells could be released from cellular membranes by phosphatidylinositol-specific phospholipase C.; Alternative processing of the mRNA of acetylcholinesterase at the 5{dollar}spprime{dollar} terminus of AChE mRNA was also analyzed by Northern blot analysis, RNase protection and primer extension. The results indicate that alternative splicing of the 5{dollar}spprime{dollar} non-coding region of AChE mRNA is related to the alternative usages of different promoters. The promoter region, which appeared to be most extensively used in most tissues, was sequenced and characterized with respect to cis-elements and function.