Gelsolin, a Ca2+-dependent actin-binding protein in a hamster insulin-secreting cell line.
Using a gel overlay technique we have previously described a 90,000-mol wt actin-binding protein in a number of hormone-secreting tissues and tentatively identified this protein as gelsolin. Gelsolin is a protein that cuts or solates cross-linked actin filaments and can also serve as a nucleating site for actin polymerization. The objective of this study was to isolate this protein from a hamster insulin-secreting (HIT) cell line and compare the immunologic properties and peptide maps of purified rabbit macrophage gelsolin, human platelet gelsolin, and the HIT cell 90,000-mol wt protein. DNase I-Sepharose retained the HIT cell actin-binding proteins in 1 mM CaCl2; some of the 90,000-mol wt protein could then be eluted with 1 mM EGTA. The remaining actin-binding proteins were eluted using a buffer containing SDS. The EGTA peak fractions contained two major protein bands of Mr = 90,000 and 42,000, which suggested that a 90,000-mol wt-actin complex was eluted from the DNase I-Sepharose column. Specific antibodies to the human platelet and rabbit macrophage gelsolins bound to the 90,000-mol wt bands in the eluates, but did not crossreact with other actin-binding proteins. Indirect immunofluorescence using an anti-human platelet gelsolin antibody localized the 90,000-mol wt protein to stress fibers that were also stained with phalloidin, which suggested that gelsolin is associated with actin in vivo. Tryptic peptide maps of all three radioiodinated gelsolins were virtually indistinguishable. Thus, gelsolin is a highly conserved gene product found in at least three diverse cell types, an insulin-secreting beta cell line, macrophages, and platelets, and may link a transient increase in Ca2+ cellular levels with changes in actin polymerization and/or the gel-sol state of these cells.
Nelson
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Boyd
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D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D002352:Carrier Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D002457:Cell Extracts / Q000032:analysis; D002460:Cell Line; D002847:Chromatography, Agarose; D006224:Cricetinae; D003429:Cross Reactions; D003850:Deoxyribonuclease I; D018260:Gelsolin; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D008840:Microfilament Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D014357:Trypsin
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D002457:Cell Extracts; D018260:Gelsolin; D007328:Insulin; D008840:Microfilament Proteins; D010446:Peptide Fragments; C016953:brevin; D003850:Deoxyribonuclease I; D014357:Trypsin; D002118:Calcium