Severe myopathy in mice lacking the MEF2/SRF-dependent gene leiomodin-3
Bercin K. Cenik, … , Eric N. Olson, Ning Liu
Bercin K. Cenik, … , Eric N. Olson, Ning Liu
Published March 16, 2015
Citation Information: J Clin Invest. 2015;125(4):1569-1578. https://doi.org/10.1172/JCI80115.
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Research Article Muscle biology

Severe myopathy in mice lacking the MEF2/SRF-dependent gene leiomodin-3

Abstract

Maintenance of skeletal muscle structure and function requires a precise stoichiometry of sarcomeric proteins for proper assembly of the contractile apparatus. Absence of components of the sarcomeric thin filaments causes nemaline myopathy, a lethal congenital muscle disorder associated with aberrant myofiber structure and contractility. Previously, we reported that deficiency of the kelch-like family member 40 (KLHL40) in mice results in nemaline myopathy and destabilization of leiomodin-3 (LMOD3). LMOD3 belongs to a family of tropomodulin-related proteins that promote actin nucleation. Here, we show that deficiency of LMOD3 in mice causes nemaline myopathy. In skeletal muscle, transcription of Lmod3 was controlled by the transcription factors SRF and MEF2. Myocardin-related transcription factors (MRTFs), which function as SRF coactivators, serve as sensors of actin polymerization and are sequestered in the cytoplasm by actin monomers. Conversely, conditions that favor actin polymerization de-repress MRTFs and activate SRF-dependent genes. We demonstrated that the actin nucleator LMOD3, together with its stabilizing partner KLHL40, enhances MRTF-SRF activity. In turn, SRF cooperated with MEF2 to sustain the expression of LMOD3 and other components of the contractile apparatus, thereby establishing a regulatory circuit to maintain skeletal muscle function. These findings provide insight into the molecular basis of the sarcomere assembly and muscle dysfunction associated with nemaline myopathy.

Authors

Bercin K. Cenik, Ankit Garg, John R. McAnally, John M. Shelton, James A. Richardson, Rhonda Bassel-Duby, Eric N. Olson, Ning Liu

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Figure 5

Regulation of Lmod3 promoter by MEF2 and SRF transcription factors.

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Regulation of Lmod3 promoter by MEF2 and SRF transcription factors. (A) ...
(A) The Lmod3 promoter region contains consensus sites for MEF2 and SRF (CArG box). The MEF2 site is also a TATA box. Numbering of nucleotide positions is relative to the ATG (+1) start codon. (B) LacZ staining was performed on E12.5 F0 embryos containing the 1691- or 1069-bp enhancer fragment of the Lmod3 promoter cloned into an hsp68-LacZ construct. WT Lmod3 Tg embryos showed strong muscle-specific expression that was diminished in the SRF/MEF2 Tg mutants. Scale bar: 1 mm. (C) COS-7 cells were transfected with the upstream fragments of the Lmod3 gene cloned into the luciferase reporter plasmid pGL3. Luciferase reporter assays showed activation of the Lmod3 promoter by SRF in response to its coactivator MRTF-A. (D) Luciferase activity was measured in COS-7 cells transfected with the 1691-bp Lmod3 reporter plasmid with or without mutations in the MEF2 site and/or the CArG box. Cells were cotransfected with MEF2C. (E) Luciferase reporter activity was measured in COS-7 cells transfected with the 1691-bp Lmod3 reporter plasmid with or without mutations in the CArG box and/or the MEF2 site. Cells were cotransfected with MRTF-A. Experiments were performed at least twice, with all transfections done in triplicate. All results are reported as percentage values ± SEM (WT construct with MEF2C and MRTF-A was set to 100%). Luc, luciferase.