Primary structure of rat plasma membrane Ca(2+)-ATPase isoform 4 and analysis of alternative splicing patterns at splice site A.
We have determined the primary structure of the rat plasma membrane Ca(2+)-ATPase isoform 4 (PMCA4), and have analysed its mRNA tissue distribution and alternative splicing patterns at splice site A. Rat PMCA4 (rPMCA4) genomic clones were isolated and used to determine the coding sequences and intron/exon organization of the 5'-end of the gene, and the remaining coding sequence was determined from PCR-amplified cDNA fragments. Pairwise comparisons reveal that the amino acid sequence of rPMCA4 has diverged substantially from those of rPMCA isoforms 1, 2 and 3 (73-76% identity) and from that of human PMCA4 (87%). Despite the high degree of sequence divergence between the two species, comparisons of intron and untranslated mRNA sequences with the corresponding human sequences confirm the identity of this rat isoform as PMCA4. Northern blot studies demonstrate that the PMCA4 mRNA is expressed in all rat tissues examined except liver, with the highest levels in uterus and stomach. A combination of PCR analysis of alternative splicing patterns and sequence analysis of the gene demonstrate that a 36 nt exon at site A is included in PMCA4 mRNAs of most tissues but is largely excluded in heart and testis. Alternative splicing of both the 36 nt exon and a previously characterized 175 nt exon at splice site C, each of which can be either included or excluded in a highly tissue-specific manner, leads to the production of four different PMCA4 variants ranging in size from 1157 to 1203 amino acids.
Keeton
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TP
Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, OH 45267-0524.
Shull
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GE
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D000252:Calcium-Transporting ATPases / Q000235:genetics / Q000302:isolation & purification*; D002462:Cell Membrane / Q000201:enzymology*; D003001:Cloning, Molecular; D015698:Genomic Library; D007527:Isoenzymes; D008969:Molecular Sequence Data; D009928:Organ Specificity; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D016415:Sequence Alignment
D007527:Isoenzymes; D012333:RNA, Messenger; D000252:Calcium-Transporting ATPases