CDNA CLONING, EXPRESSION AND CHROMOSOMAL LOCALIZATION OF THE HUMAN SARCO ENDOPLASMIC RETICULUM CA2+-ATPASE-3 GENE/

cDNA and genomic clones encoding human sarco/endoplasmic reticulum Ca2 +-ATPase 3 (SERCA3) were isolated. The composite nucleotide sequence o f the 4.6 kb cDNA, as well as the partial structure of 25 kb of genomi c DNA encoding all but the 5' region of the gene, was determined. The nucleotide sequence coding for the last six amino acids of the pump an d the 3'-untranslated region were identified within the sequence of th e last exon. Northern blot hybridization analysis using cDNA probes de rived from this exon detected a 4.8 kb transcript in several human tis sues. Using a cDNA probe derived from the 5'-coding region an unexpect ed mRNA distribution pattern, consisting of two mRNA species of 4.8 an d 4.0 kb, was detected in thyroid gland and bone marrow only, This is the first indication of an alternative splicing mechanism operating on the SERCA3 gene transcript, which most likely generates SERCA3 isofor ms with altered C-termini. Human SERCA3 expressed in platelets and in COS cells transfected with the corresponding cDNA was detected with th e previously described antibody N89 (directed against the N-terminal r egion of rat SERCA3) and with a new SERCA3-specific antiserum C91, dir ected against the extreme C-terminus of the human isoform. A monoclona l antibody PL/IM430, previously assumed to recognize SERCA3 in human p latelets, does not react with the 97 kDa human SERCA3 transiently expr essed in COS cells. Therefore the 97 kDa isoform detected by PL/IM430 more likely represents a novel SERCA pump, as recently suggested [Kova cs, Corvazier, Papp, Magnier, Bredoux, Enyedi, Sarkadi and Enouf (1994 ) J. Biol. Chem. 269, 6177-6184]. Finally, by fluorescence in situ hyb ridization and chromosome G-banding analyses, the SERCA3 gene was assi gned to human chromosome 17p13.3.