Leiomodin-3 dysfunction results in thin filament disorganization and nemaline myopathy
Michaela Yuen, … , Kathryn N. North, Nigel F. Clarke
Michaela Yuen, … , Kathryn N. North, Nigel F. Clarke
Published September 24, 2014
Citation Information: J Clin Invest. 2014;124(11):4693-4708. https://doi.org/10.1172/JCI75199.
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Research Article

Leiomodin-3 dysfunction results in thin filament disorganization and nemaline myopathy

Abstract

Nemaline myopathy (NM) is a genetic muscle disorder characterized by muscle dysfunction and electron-dense protein accumulations (nemaline bodies) in myofibers. Pathogenic mutations have been described in 9 genes to date, but the genetic basis remains unknown in many cases. Here, using an approach that combined whole-exome sequencing (WES) and Sanger sequencing, we identified homozygous or compound heterozygous variants in LMOD3 in 21 patients from 14 families with severe, usually lethal, NM. LMOD3 encodes leiomodin-3 (LMOD3), a 65-kDa protein expressed in skeletal and cardiac muscle. LMOD3 was expressed from early stages of muscle differentiation; localized to actin thin filaments, with enrichment near the pointed ends; and had strong actin filament-nucleating activity. Loss of LMOD3 in patient muscle resulted in shortening and disorganization of thin filaments. Knockdown of lmod3 in zebrafish replicated NM-associated functional and pathological phenotypes. Together, these findings indicate that mutations in the gene encoding LMOD3 underlie congenital myopathy and demonstrate that LMOD3 is essential for the organization of sarcomeric thin filaments in skeletal muscle.

Authors

Michaela Yuen, Sarah A. Sandaradura, James J. Dowling, Alla S. Kostyukova, Natalia Moroz, Kate G. Quinlan, Vilma-Lotta Lehtokari, Gianina Ravenscroft, Emily J. Todd, Ozge Ceyhan-Birsoy, David S. Gokhin, Jérome Maluenda, Monkol Lek, Flora Nolent, Christopher T. Pappas, Stefanie M. Novak, Adele D’Amico, Edoardo Malfatti, Brett P. Thomas, Stacey B. Gabriel, Namrata Gupta, Mark J. Daly, Biljana Ilkovski, Peter J. Houweling, Ann E. Davidson, Lindsay C. Swanson, Catherine A. Brownstein, Vandana A. Gupta, Livija Medne, Patrick Shannon, Nicole Martin, David P. Bick, Anders Flisberg, Eva Holmberg, Peter Van den Bergh, Pablo Lapunzina, Leigh B. Waddell, Darcée D. Sloboda, Enrico Bertini, David Chitayat, William R. Telfer, Annie Laquerrière, Carol C. Gregorio, Coen A.C. Ottenheijm, Carsten G. Bönnemann, Katarina Pelin, Alan H. Beggs, Yukiko K. Hayashi, Norma B. Romero, Nigel G. Laing, Ichizo Nishino, Carina Wallgren-Pettersson, Judith Melki, Velia M. Fowler, Daniel G. MacArthur, Kathryn N. North, Nigel F. Clarke

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Figure 5

LMOD3 expression in human muscle tissue and primary cells.

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LMOD3 expression in human muscle tissue and primary cells. LMOD3 express...
LMOD3 expression during muscle cell differentiation and muscle development in (A) human skeletal muscle tissue and (B) differentiating primary human myoblasts. (A) In skeletal muscle, LMOD3 was detected at all ages tested (assessed age range: 14 weeks gestation to 58 years), but expression before birth was lower than that in mature muscle. Developmental MHC (dMHC) and cardiac actin (c actin) were expressed in fetal muscle biopsies as expected. Sarcomeric actin (s actin), α-actinin-2, GAPDH, and MHC (Coomassie-stained MHC [cMHC]) served as loading controls. (B) LMOD3 was detected weakly in undifferentiated human myoblasts (day 0 [d0]), and levels increased during differentiation into myotubes similar to other muscle proteins, such as α-actinin-2, developmental MHC, and cardiac actin. β-Tubulin served as a loading control. PHM Diff, primary human myoblasts differentiation. (C) LMOD2 and LMOD3 expression in mature human cardiac muscle and skeletal muscle. LMOD3 expression was higher in skeletal muscle than in heart, while the reverse was true for LMOD2. (D) Localization of LMOD3 in primary human myoblasts at day 10 of differentiation by confocal imaging. (E) Magnified view of white squares in D. Square 1 (S1) shows a striated area, demonstrating that LMOD3 was most abundant at the thin filament pointed end and/or M line, which colocalize in unstretched sarcomeres, and was absent from the Z-disc (yellow arrows). In square 2 (S2), LMOD3 staining appears granular in areas where striations have not formed. Scale bar: 7.5 μm (D and E). C, human heart; S, postnatal human skeletal muscle.