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The Src signaling pathway regulates osteoclast lysosomal enzyme secretion and is rapidly modulated by estrogen
1Department of Biology, University of Minnesota, Duluth 55812, USA.
Abstract:
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.
Insights
Estrogen signaling regulates osteoclast activity by modulating the pp60src pathway. This study reveals how pp60src kinase activity influences lysosomal enzyme secretion in osteoclasts.
Area of Science:
- Cell Biology
- Molecular Biology
- Endocrinology
Background:
- Osteoclasts are crucial for bone remodeling.
- The pp60src signaling pathway plays a role in cellular functions.
- Estrogen (17betaE2) influences bone metabolism and osteoclast activity.
Purpose of the Study:
- To investigate the role of the pp60src signaling pathway in osteoclast activity.
- To determine how estrogen affects pp60src kinase activity.
- To elucidate the interplay between estrogen and pp60src in regulating lysosomal enzyme secretion.
Main Methods:
- Osteoclasts were transfected with dominant-negative and constitutively active pp60src, c-Ras, and c-Raf expression vectors.
- Lysosomal enzyme secretion (cathepsin B/K, TRAP) was monitored.
- pp60src tyrosine kinase activity was measured in response to estrogen treatment.
Main Results:
- Disrupting pp60src, c-Ras, or c-Raf function increased lysosomal enzyme secretion.
- Constitutively active pp60src decreased enzyme secretion.
- Estrogen decreased pp60src kinase activity and lysosomal enzyme secretion.
- Constitutively active pp60src blocked estrogen's effects, while dominant-negative constructs enhanced them.
Conclusions:
- The kinase domain of cytoskeletal-associated pp60src is involved in regulating lysosomal enzyme secretion in osteoclasts.
- Estrogen exerts its effects on osteoclast function, in part, through modulation of the pp60src signaling pathway.
- These findings provide insights into the molecular mechanisms of estrogen's action on bone resorption.