title: Platelet Ca(2+)ATPases : a plural, species-specific, and multiple hypertension-regulated expression system.
abstract: Gaining insight into nonmuscle Ca(2+) signaling requires basic knowledge of the major structures involved. We investigated the expression of platelet Ca(2+)ATPases in normal and hypertension-associated abnormal Ca(2+) signaling. First, overall identification of normotensive Wistar-Kyoto rat Ca(2+)ATPases was attempted by looking for newly described human platelet 3'-end alternatively spliced sarco/endoplasmic reticulum Ca(2+)ATPases (SERCA) 3b mRNA and plasma membrane Ca(2+)ATPase (PMCA) 1b and 4b proteins, in addition to SERCA2b and SERCA3a isoforms. For SERCAs, comparative analyses of human and Wistar-Kyoto rat SERCA3 platelet mRNA by reverse transcription-polymerase chain reaction (RT-PCR) followed by sequencing established that human platelets coexpressed SERCA3b and a third SERCA3c, while rat cells were devoid of them but expressed a still unknown splice variant that we termed rSERCA3b/3c. Its identification using 3'-end SERCA3 gene and rapid amplification of cDNA ends (RACE)-PCR studies showed that it results from an additional SERCA3 alternative splicing process, which uses a second alternative polyadenylation site located in the last intron. For PMCAs, with the use of gene-specific RT-PCR followed by sequencing and Western blotting using 5F10 monoclonal antibody, expression of human and rat platelet PMCA1b and PMCA4b was similar. Second, comparative analysis of these newly identified Ca(2+)ATPases and SERCA3a in age-matched spontaneously hypertensive rat platelets demonstrated (1) a marked downregulation of rSERCA3b/3c, which became null, and a 1.71-fold increase in SERCA3a and (2) an opposite regulation of the 2 PMCAs, namely, a 3.3-fold decrease in PMCA1b mRNA and a 3.7-fold increase in PMCA4b mRNA. Hence, platelets coexpress multiple, diverse, and species-specific Ca(2+)ATPases, including a novel fourth SERCA3. Moreover, expression of PMCA (1b and 4b), SERCA3a, and rSERCA3b/3c was modulated in rat hypertension. Hence, Ca(2+)ATPases should be regarded as constituting a new rational basis for the understanding of nonmuscle cell Ca(2+) signaling.
authors.affiliation: U348 INSERM, IFR Circulation Lariboisière, Hôpital Lariboisière, Paris, France.
authors.forename: V
authors.initials: V
authors.lastname: Martin
authors.forename: R
authors.initials: R
authors.lastname: Bredoux
authors.forename: E
authors.initials: E
authors.lastname: Corvazier
authors.forename: B
authors.initials: B
authors.lastname: Papp
authors.forename: J
authors.initials: J
authors.lastname: Enouf
mesh_terms: D000328:Adult; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001792:Blood Platelets / Q000201:enzymology*; D020013:Calcium Signaling; D000252:Calcium-Transporting ATPases / Q000097:blood* / Q000235:genetics*; D002462:Cell Membrane / Q000201:enzymology; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000201:enzymology; D006367:HeLa Cells; D006801:Humans; D006973:Hypertension / Q000201:enzymology* / Q000235:genetics*; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000097:blood / Q000235:genetics; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000097:blood / Q000235:genetics; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D000252:Calcium-Transporting ATPases