Calreticulin negatively regulates the surface expression of Cav1.3 L-type calcium channel
Eddy Karnabi1, Yongxia Qu, Yunkun Yue
1Cardiovascular Research Program, Department of Physiology and Pharmacology, State University of New York Downstate Medical Center, Brooklyn, USA.
Background:
The neuroendocrine Cav1.3 L-type Ca channels have been recently found in the Human fetal heart and shown to play a vital role in Ca entry from the sarcolemma into the cell and in Ca homeostasis. Calreticulin, a Ca binding endoplasmic reticulum (ER) resident protein, has been recently shown to translocate to the cell surface where its role and function are just emerging. Here, we demonstrated a novel mechanism of Cav1.3 and calreticulin interaction resulting in downregulation of Cav1.3 channel densities in native Human fetal cardiac cells and Human Embryonic Kidney cell lines (tsA201).
Methods And Results:
Cell surface and cytoplasmic staining of calreticulin was demonstrated first in cultured human fetal cardiomyocytes (HFC), gestational age 18-24 weeks, using confocal microscopy thereby establishing that calreticulin is present at the cell surface in HFC. Co-immunoprecipitation from HFC using anti-Cav1.3 Ca channel antibody, and probing with anti-calreticulin antibody revealed a 46 kDa band corresponding to calreticulin suggesting that Cav1.3 Ca channel and calreticulin co-assemble in a macromolecular complex. Co-expression of Cav1.3 and calreticulin in tsA201 cells resulted in a decrease in surface expression of Cav1.3 Ca channels. These findings were consistent with the electrophysiological studies showing that co-transfection of Cav1.3 Ca channel and calreticulin resulted in 55% reduction of Cav1.3 Ca current densities recorded from tsA201 cells.
Conclusions:
The results show the first evidence that calreticulin: (1) is localized outside the ER on the cell surface of HFC; (2) coimmunoprecipitates with Cav1.3 L-type Ca channel; (3) negatively regulates Cav1.3 surface expression thus resulting in decreased Cav1.3 Ca current densities. The data demonstrate a novel mechanism of modulation of Cav1.3 Ca channel by calreticulin, which may be involved in pathological settings such as autoimmune associated congenital heart block where Cav1.3 Ca channels are downregulated.
Insights
Calreticulin, a protein, interacts with Cav1.3 calcium channels on the cell surface of human fetal heart cells. This interaction reduces Cav1.3 channel activity, offering insights into heart conditions.
Area of Science:
- Cardiology
- Cell Biology
- Molecular Biology
Background:
- Cav1.3 L-type Ca channels are crucial for calcium entry and homeostasis in the human fetal heart.
- Calreticulin, an endoplasmic reticulum protein, is increasingly recognized for its cell surface functions.
Purpose of the Study:
- To investigate the novel interaction between Cav1.3 channels and calreticulin in human fetal cardiac cells.
- To elucidate the functional consequences of this interaction on Cav1.3 channel activity and surface expression.
Main Methods:
- Confocal microscopy to detect cell surface calreticulin in human fetal cardiomyocytes (HFC).
- Co-immunoprecipitation assays to confirm Cav1.3 and calreticulin complex formation.
- Co-expression studies in tsA201 cells and electrophysiological recordings to assess Cav1.3 current densities.
Main Results:
- Calreticulin is present on the cell surface of HFC.
- Cav1.3 Ca channels and calreticulin co-assemble in a macromolecular complex.
- Co-expression of calreticulin with Cav1.3 channels reduces Cav1.3 surface expression and current densities by 55%.
Conclusions:
- Calreticulin localizes to the cell surface of HFC and co-immunoprecipitates with Cav1.3 channels.
- Calreticulin negatively regulates Cav1.3 surface expression, decreasing channel activity.
- This novel mechanism may play a role in conditions like congenital heart block where Cav1.3 channels are downregulated.
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