Cloning of human Ca2+ homoeostasis endoplasmic reticulum protein (CHERP): regulated expression of antisense cDNA depletes CHERP, inhibits intracellular Ca2+ mobilization and decreases cell proliferation.
A monoclonal antibody which blocks InsP(3)-induced Ca(2+) release from isolated endoplasmic reticulum was used to isolate a novel 4.0 kb cDNA from a human erythroleukaemia (HEL) cell cDNA expression library. A corresponding mRNA transcript of approx. 4.2 kb was present in all human cell lines and tissues examined, but cardiac and skeletal muscle had an additional transcript of 6.4 kb. The identification in GenBank(R) of homologous expressed sequence tags from many tissues and organisms suggests that the gene is ubiquitously expressed in higher eukaryotes. The gene was mapped to human chromosome 19p13.1. The cDNA predicts a 100 kDa protein, designated Ca(2+) homoeostasis endoplasmic reticulum protein (CHERP), with two putative transmembrane domains, multiple consensus phosphorylation sites, a polyglutamine tract of 12 repeats and regions of imperfect tryptophan and histadine octa- and nona-peptide repeats. In vitro translation of the full-length cDNA produced proteins of M(r) 128000 and 100000, corresponding to protein bands detected by Western blotting of many cell types. CHERP was co-localized in HEL cells with the InsP(3) receptor by two-colour immunofluorescence. Transfection of HEL cells with antisense cDNA led to an 80% decline in CHERP within 5 days of antisense induction, with markedly decreased intracellular Ca(2+) mobilization by thrombin, decreased DNA synthesis and growth arrest, indicating that the protein has an important function in Ca(2+) homoeostasis, growth and proliferation.
Laplante
J M
JM
Department of Pharmacology, The University of Connecticut Health Center, Farmington 06032, USA.
O'Rourke
F
F
Lu
X
X
Fein
A
A
Olsen
A
A
Feinstein
M B
MB
D000595:Amino Acid Sequence; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D003001:Cloning, Molecular; D016373:DNA, Antisense / Q000235:genetics / Q000494:pharmacology; D018076:DNA, Complementary / Q000032:analysis; D004268:DNA-Binding Proteins; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D015723:Gene Library; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D017136:Ion Transport; D008565:Membrane Proteins / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D016601:RNA-Binding Proteins; D014162:Transfection
C410225:CHERP protein, human; D016373:DNA, Antisense; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D008565:Membrane Proteins; D016601:RNA-Binding Proteins; D002118:Calcium