FoxP1 Represses MEF2A in Striated Muscle

Abstract

Myocyte enhancer factor 2 (MEF2) proteins are involved in multiple developmental, physiological, and pathological processes in vertebrates. Protein-protein interactions underlie the plethora of biological processes impacted by MEF2A, necessitating a detailed characterization of the MEF2A interactome. A nanobody based affinity-purification/mass spectrometry strategy was employed to achieve this goal. Specifically, the MEF2A protein complexes were captured from myogenic lysates using a GFP-tagged MEF2A protein immobilized with a GBP-nanobody followed by LC-MS/MS proteomic analysis to identify MEF2A interactors. After bioinformatic analysis, we further characterized the interaction of MEF2A with a transcriptional repressor, FOXP1. FOXP1 coprecipitated with MEF2A in proliferating myogenic cells which diminished upon differentiation (myotube formation). Ectopic expression of FOXP1 inhibited MEF2A driven myogenic reporter genes (derived from the creatine kinase muscle and myogenin genes) and delayed induction of endogenous myogenin during differentiation. Conversely, FOXP1 depletion enhanced MEF2A transactivation properties and myogenin expression. The FoxP1:MEF2A interaction is also preserved in cardiomyocytes and FoxP1 depletion enhanced cardiomyocyte hypertrophy. FOXP1 prevented MEF2A phosphorylation and activation by the p38MAPK pathway. Overall, these data implicate FOXP1 in restricting MEF2A function in order to avoid premature differentiation in myogenic progenitors and also to possibly prevent re-activation of embryonic gene expression in cardiomyocyte hypertrophy.

Keywords:

• Myogenesis
• phosphorylation
• differentiation
• cardiomyocyte
• MEF2
• FOXP1

Acknowledgments

The authors thank Gwillym Declan Williams for assistance with the proteomic data submission. The authors acknowledge the SPARC BioCentre, Hospital for Sick Children, Toronto, Canada for performing the mass spectrometry analysis.

Data availability statement

The materials and reagents described herein are freely available to the research community to use for noncommercial purpose and will be provided through contact with the corresponding author. The mass spectrometry data have been deposited with the ProteomeXchange Consortium via the PRIDE⁶² partner repository with the dataset identifier PXD050042.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

This research was funded by a Project Grant (PJT-159644) to JCM from the Canadian Institutes of Health Research (CIHR). JCM is a recipient of the McLaughlin Research Chair in the Faculty of Science, York University.