The Src signaling pathway regulates osteoclast lysosomal enzyme secretion and is rapidly modulated by estrogen.
To investigate the role of the pp60src signaling pathway in osteoclast activity, we have used dominant negative pp60src, c-ras, and c-raf expression vectors to individually disrupt their functions in osteoclasts. Osteoclasts were transiently transfected and secretions of cathepsin B/K and tartrate-resistant acid phosphatase (TRAP) were monitored. Expression of these constructs increased secretion of lysosomal enzymes. In contrast, constitutively active pp60src expression caused decreased lysosomal enzyme secretion. Osteoclasts respond to 17-beta estradiol (17betaE2) treatment with decreased lysosomal enzyme secretion. Therefore, we investigated the effects of E2 on pp60src kinase activity and observed an E2 time- and dose-dependent decrease in cytoskeletal membrane-associated pp60src tyrosine kinase activity. We have shown that estrogen decreases lysosomal enzyme gene expression and secretion; so we have examined the effects of the expression constructs on estrogen regulation of enzyme secretion. Constitutively active pp60src blocked E2 effects on secretion whereas expression of dominant negative pp60src, c-Ras, or c-Raf enhanced E2 effects. These data support that the kinase domain of cytoskeletal-associated pp60src is likely to be involved in the regulation of lysosomal enzyme secretion.
Pascoe
D
D
Department of Biology, University of Minnesota, Duluth 55812, USA.
Oursler
M J
MJ
D000135:Acid Phosphatase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002401:Cathepsin B / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D002645:Chickens; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000494:pharmacology; D004967:Estrogens / Q000378:metabolism* / Q000494:pharmacology; D000077267:Fulvestrant; D011905:Genes, ras; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology*; D010010:Osteoclasts / Q000187:drug effects / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000187:drug effects / Q000378:metabolism; D051076:Proto-Oncogene Proteins c-fyn; D019908:Proto-Oncogene Proteins c-raf / Q000187:drug effects / Q000378:metabolism; D051575:Proto-Oncogene Proteins c-yes; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D000071681:Tartrate-Resistant Acid Phosphatase; D014162:Transfection; D019061:src-Family Kinases / Q000378:metabolism*
D004965:Estrogen Antagonists; D004967:Estrogens; D007527:Isoenzymes; D011518:Proto-Oncogene Proteins; D000077267:Fulvestrant; D004958:Estradiol; D051076:Proto-Oncogene Proteins c-fyn; D051575:Proto-Oncogene Proteins c-yes; D016392:Proto-Oncogene Proteins pp60(c-src); D019061:src-Family Kinases; D019908:Proto-Oncogene Proteins c-raf; D000135:Acid Phosphatase; D000071681:Tartrate-Resistant Acid Phosphatase; D002401:Cathepsin B