Transforming growth factor-beta1 and bone morphogenetic protein-2 downregulate CaV3.1 channel expression in mouse C2C12 myoblasts
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Published on: May 31, 2016
Traudy Avila1, Arturo Andrade, Ricardo Felix
1Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute (Cinvestav-IPN), Mexico City, Mexico.
Abstract:
In the developing skeletal muscle, fusion of myoblasts and myotube formation is a process that involves Ca2+ influx through T-type (CaV3) channels. Treatment of myoblasts with transforming growth factor-beta1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) decreases the number of CaV3 channels in the plasma membrane and reduces myotube formation. In the current report, we examined whether the inhibitory actions of TGF-beta1 and BMP-2 involve alterations in CaV3 mRNA expression in the myoblast C2C12 cell line. Using RT-PCR, we found that CaV3.1 but not CaV3.2 and CaV3.3 transcripts are present in either undifferentiated or fusion competent C2C12 myoblasts. Semi-quantitative analysis revealed a significant decrease of CaV3.1 mRNA expression in cells treated with TGF-beta1 and BMP-2. In contrast, patch-clamp recordings on HEK-293 cells stably expressing recombinant CaV3.1 channels showed that T-type currents were not affected by chronic exposure to the growth factors. These results suggest that muscle T-channel downregulation by TGF-beta1 and BMP-2 may be mediated by reduced transcription rather than through post-transcriptional modifications of CaV3.1 channels.
Insights
Transforming growth factor-beta1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) reduce skeletal muscle T-type calcium channels (CaV3.1) by decreasing their mRNA expression. This suggests transcriptional regulation, not post-transcriptional modification, impacts myoblast fusion.
Area of Science:
- Molecular Biology
- Cell Biology
- Muscle Physiology
Background:
- Skeletal muscle development involves myoblast fusion and myotube formation, processes dependent on calcium influx via T-type (CaV3) channels.
- Transforming growth factor-beta1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) inhibit myotube formation by reducing CaV3 channel numbers in the plasma membrane.
Purpose of the Study:
- To investigate if TGF-beta1 and BMP-2 inhibit myotube formation by altering CaV3 mRNA expression in C2C12 myoblasts.
- To determine the specific CaV3 channel subtypes involved in TGF-beta1 and BMP-2 mediated inhibition.
Main Methods:
- Reverse transcription polymerase chain reaction (RT-PCR) to analyze CaV3 mRNA expression in C2C12 myoblasts.
- Semi-quantitative analysis of CaV3.1 mRNA levels following treatment with TGF-beta1 and BMP-2.
- Patch-clamp recordings on HEK-293 cells expressing CaV3.1 channels to assess T-type currents after growth factor exposure.
Main Results:
- CaV3.1 transcripts were detected in C2C12 myoblasts, while CaV3.2 and CaV3.3 were absent.
- TGF-beta1 and BMP-2 significantly decreased CaV3.1 mRNA expression in myoblasts.
- Chronic exposure to TGF-beta1 and BMP-2 did not affect T-type currents in HEK-293 cells expressing CaV3.1 channels.
Conclusions:
- TGF-beta1 and BMP-2 downregulate muscle T-type calcium channels (CaV3.1) primarily through reduced mRNA transcription.
- These findings suggest that the inhibitory effects of TGF-beta1 and BMP-2 on myotube formation are mediated by transcriptional downregulation of CaV3.1 channels.
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