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Presence and tyrosine phosphorylation of c-met receptor in human sperm
1Department of Obstetrics and Gynecology, Medical College of Ohio, Toledo 43614-5806, USA.
Abstract:
The c-met receptor is a p190MET tyrosine kinase proto-oncoprotein that through its binding to its ligand, designated hepatocyte growth factor (HGF), induces mitogenic, motogenic, and morphogenic activities in a variety of cell types. The present study was conducted to examine whether or not the c-met receptor is expressed and tyrosine phosphorylated in the human sperm cell. The Western blot analysis, using a monoclonal antibody (MAb2) directed against the extracellular domain of the c-met receptor, showed a specific band of 195 kDa corresponding to the intact c-met receptor in the detergent-solubilized human sperm preparation (HSP). This protein band was not recognized by the control myeloma lg (immunoglobin). In the immunoprecipitation procedure, a similar specific band of 195 kDa and a 145-kDa band corresponding to the beta-subunit of c-met receptor were seen. In the indirect immunofluorescence technique, the c-met receptor was localized predominantly in the acrosomal region of the sperm cell. The c-met receptor was tyrosine phosphorylated/autophosphorylated during capacitation and in the cell-free in vitro kinase assay. Incubation of human sperm with hepatocyte growth factor (HGF) or MAb2 to c-met receptor enhanced the degree of tyrosine phosphorylation/autophosphorylation of the c-met receptor up to 5.1-fold. These findings indicate that the c-met receptor is present in the acrosomal region of human sperm cell and is tyrosine phosphorylated, which is enhanced by HGF and the receptor antibody. The c-met system may have an important role in sperm function.
Insights
The c-met receptor, a key protein in cell signaling, is present in human sperm. Its tyrosine phosphorylation, enhanced by hepatocyte growth factor (HGF), suggests a role in sperm function.
Area of Science:
- Reproductive Biology
- Cell Signaling
- Molecular Endocrinology
Background:
- The c-met receptor tyrosine kinase, activated by hepatocyte growth factor (HGF), regulates crucial cellular processes.
- Proto-oncogene c-MET (MET) and its ligand HGF are implicated in various physiological and pathological functions.
- The presence and role of the c-met receptor in human sperm remain largely unexplored.
Purpose of the Study:
- To investigate the expression and tyrosine phosphorylation of the c-met receptor in human sperm.
- To determine the localization of the c-met receptor within human sperm cells.
- To assess the impact of HGF and a specific antibody on c-met receptor phosphorylation in sperm.
Main Methods:
- Western blot analysis to detect c-met receptor protein in human sperm preparations.
- Immunoprecipitation to identify c-met receptor and its beta-subunit.
- Indirect immunofluorescence microscopy to localize c-met receptor expression.
- In vitro kinase assays to study tyrosine phosphorylation during capacitation and with HGF stimulation.
Main Results:
- A 195 kDa band, corresponding to the intact c-met receptor, was identified in human sperm using Western blot and immunoprecipitation.
- Immunofluorescence localized the c-met receptor predominantly to the acrosomal region of sperm.
- Tyrosine phosphorylation of the c-met receptor was observed during sperm capacitation and significantly enhanced by HGF or MAb2.
- HGF and MAb2 increased c-met receptor tyrosine phosphorylation by up to 5.1-fold.
Conclusions:
- The c-met receptor is expressed and tyrosine phosphorylated in the human sperm acrosomal region.
- Hepatocyte growth factor (HGF) and receptor antibody stimulation enhance c-met receptor tyrosine phosphorylation.
- The c-met signaling system likely plays a significant role in human sperm function.