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Updated: May 28, 2026
Automated Detection and Analysis of Exocytosis
Published on: September 11, 2021
Insight into the role of Ca2+-binding protein 5 in vesicle exocytosis
Izabela Sokal1, Françoise Haeseleer
1Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195, USA.
Purpose:
CaBP5 is a neuronal calmodulin-like Ca(2+)-binding protein that is expressed in the retina and in the cochlea. Although CaBP5 knockout mice displayed reduced sensitivity of retinal ganglion cell light responses, the function of CaBP5 in vivo is still unknown. To gain further insight into CaBP5 function, the authors screened for CaBP5-interacting partners.
Methods:
Potential retinal interacting partners for CaBP5 were identified using affinity chromatography followed by mass spectrometry and by yeast two-hybrid screening of a bovine retina cDNA library. Interacting partners were further analyzed using coimmunoprecipitation. Immunohistochemistry and subcellular fractionation were performed to determine their colocalization in the retina. The effect of CaBP5 on dopamine release and neurite outgrowth of PC12 cells was analyzed using ELISA and fluorescent labeling.
Results:
Using affinity chromatography, the authors identified Munc18-1 and myosin VI as interacting partners for CaBP5. Munc18-1 was also identified using the yeast two-hybrid system. Colocalization and coimmunoprecipitation of CaBP5 with these two proteins in retinal tissue further established their physiological interactions. Furthermore, CaBP5 expression in NGF-stimulated PC12 cells stimulates neurite outgrowth and dopamine exocytosis.
Conclusions:
This study shows that CaBP5 interacts with Munc18-1 and myosin VI, two proteins involved in the synaptic vesicle cycle. Together with the effect of CaBP5 in stimulating neurite outgrowth and vesicle exocytosis in PC12 cells, these results suggest that CaBP5 plays a role in neurotransmitter release.
Insights
Calcium-binding protein 5 (CaBP5) interacts with Munc18-1 and myosin VI, proteins crucial for synaptic vesicle cycling. This suggests CaBP5 plays a role in neurotransmitter release and neurite outgrowth.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Calcium-binding protein 5 (CaBP5) is a neuronal calmodulin-like protein found in the retina and cochlea.
- CaBP5 knockout mice show diminished light response sensitivity in retinal ganglion cells, indicating a potential in vivo role.
- The precise function of CaBP5 in vivo remains largely uncharacterized.
Purpose of the Study:
- To identify CaBP5-interacting proteins in the retina to elucidate its function.
- To investigate the physiological relevance of identified interactions within retinal tissue.
- To assess the impact of CaBP5 on cellular processes like neurite outgrowth and neurotransmitter release.
Main Methods:
- Affinity chromatography coupled with mass spectrometry and yeast two-hybrid screening were employed to identify CaBP5-binding partners.
- Coimmunoprecipitation and immunohistochemistry were used to validate and localize protein interactions in retinal tissue.
- PC12 cells were utilized to analyze the effects of CaBP5 on dopamine release and neurite outgrowth via ELISA and fluorescent labeling.
Main Results:
- Munc18-1 and myosin VI were identified as novel interacting partners of CaBP5 using both biochemical and genetic screening methods.
- Coimmunoprecipitation and colocalization studies confirmed the physiological interaction of CaBP5 with Munc18-1 and myosin VI in the retina.
- CaBP5 expression promoted neurite outgrowth and dopamine exocytosis in nerve growth factor-stimulated PC12 cells.
Conclusions:
- CaBP5 interacts with Munc18-1 and myosin VI, key components of the synaptic vesicle cycle.
- These findings suggest a functional role for CaBP5 in regulating neurotransmitter release.
- CaBP5's influence on neurite outgrowth and vesicle exocytosis highlights its importance in neuronal function.
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