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Carboxypeptidase H processing and secretion in rat clonal beta-cell lines.
1Departments of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110.
Endocrinology
|September 11, 1992
Summary
Carboxypeptidase H (CP-H) is primarily membrane-associated in rat insulinoma cells, differing from primary tissues. Its secretion is stimulated by depolarization and inhibited by magnesium, suggesting a role in regulated hormone processing.
Area of Science:
- Endocrinology
- Molecular Biology
- Cell Biology
Background:
- Carboxypeptidase H (CP-H) processes peptide hormones in endocrine and neural tissues.
- Two rat insulinoma cell lines (RIN 1046-38 and RIN 1046-44) were used to study CP-H.
Purpose of the Study:
- To investigate the characteristics and localization of CP-H in rat insulinoma cell lines.
- To compare CP-H distribution in cell lines versus primary tissues.
- To examine the regulation of CP-H secretion.
Main Methods:
- Cell culture of rat insulinoma lines
- Immunoblotting
- Pulse-chase analysis
- Transmission electron microscopy
- Measurement of enzyme activity
Main Results:
- CP-H is predominantly membrane-associated (75%) in RIN cells, unlike primary tissues.
- A 56 kDa precursor is processed to a mature 54 kDa CP-H form.
- CP-H secretion is stimulated by KCl depolarization and inhibited by MgCl2.
- Dense core secretory vesicles are present in both cell lines.
Conclusions:
- Mature CP-H is mainly membrane-associated in RIN cells, with intact C-termini in all forms.
- CP-H secretion is regulated, similar to insulin secretion, via the regulated secretory pathway.