Phosphorylation of the Myogenic Factor Myocyte Enhancer Factor-2 Impacts Myogenesis In Vivo

Abstract

Activity of the myogenic regulatory protein myocyte enhancer factor-2 (MEF2) is modulated by post-translational modification. We investigated the in vivo phosphorylation of Drosophila MEF2, and identified serine 98 (S98) as a phosphorylated residue. Phospho-mimetic (S98E) and phospho-null (S98A) isoforms of MEF2 did not differ from wild-type in their activity in vitro, so we used CRISPR/Cas9 to generate an S98A allele of the endogenous gene. In mutant larvae we observed phenotypes characteristic of reduced MEF2 function, including reduced body wall muscle size and reduced expression of myofibrillar protein genes; conversely,S98A homozygotes showed enhanced MEF2 function through muscle differentiation within the adult myoblasts associated with the wing imaginal disc. In adults, S98A homozygotes were viable with normal mobility, yet showed patterning defects in muscles that were enhanced when the S98A allele was combined with a Mef2 null allele. Overall our data indicate that blocking MEF2 S98 phosphorylation in myoblasts enhances its myogenic capability, whereas blocking S98 phosphorylation in differentiating muscles attenuates MEF2 function. Our studies are among the first to assess the functional significance of MEF2 phosphorylation sites in the intact animal, and suggest that the same modification can have profoundly different effects upon MEF2 function depending upon the developmental context.

Keywords:

• MEF2
• Drosophila
• muscle
• myoblast
• transcription
• phosphorylation

ACKNOWLEDGEMENTS

We thank Dr A Veraksa for providing pUAST-CTAP(SG) and UAS-NTAP-Krz, and Dr Ruth Lehman for sharing rabbit anti-ZFh1 antibody. We acknowledge technical support from the Molecular Biology Facility.

DATA AVAILABILITY STATEMENT

The authors confirm that the data supporting the findings of this study are available at: https://figshare.com/articles/figure/Supporting_data_for_Vishal_et_al_2023/22294432

Additional information

Funding

This work was supported by R01 (GM124498) awarded by the NIH to RMC. AAD was supported by NIH grants R25 GM075149 and R25 HG007630. We acknowledge the Cell Biology Facility at the Department of Biology, University of New Mexico, supported by NIH grant P20 (GM103452) from the Institute Development Award (IdeA) Program of NIGMS.