Abstract
We recently discovered a novel gene on chromosome 19p13.1 and its product, an integral endoplasmic reticulum (ER) membrane protein, termed CHERP (calcium homoeostasis endoplasmic reticulum protein). A monoclonal antibody against its C-terminal domain inhibits Ins(1,4,5) P (3)-induced Ca(2+) release from ER membrane vesicles of many cell types, and an antisense-mediated knockdown of CHERP in human erythroleukemia (HEL) cells greatly impaired Ca(2+) mobilization by thrombin. In the present paper, we explore further CHERP's function in Jurkat T-lymphocytes. Confocal laser immunofluorescence microscopy showed that CHERP was co-localized with the Ins(1,4,5) P (3) receptor throughout the cytoplasmic and perinuclear region, as previously found in HEL cells. Transfection of Jurkat cells with a lac I-regulated mammalian expression vector containing CHERP antisense cDNA caused a knockdown of CHERP and impaired the rise of cytoplasmic Ca(2+) (measured by fura-2 acetoxymethyl ester fluorescence) caused by phytohaemagglutinin (PHA) and thrombin. A 50% fall of CHERP decreased the PHA-induced rise of the cytoplasmic free Ca(2+) concentration ([Ca(2+)](i)), but Ca(2+) influx was unaffected. Greater depletion of CHERP (>70%) did not affect the concentration of Ins(1,4,5) P (3) receptors, but diminished the rise of [Ca(2+)](i) in response to PHA to