Disruption in the tropomodulin1 (Tmod1) gene compromises cardiomyocyte development in murine embryonic stem cells by arresting myofibril maturation.
Tropomodulins (Tmods) comprise a family of capping proteins for actin filament pointed ends. To decipher the significance of Tmod1 functions during de novo myofibrillogenesis, we generated Tmod1 null embryonic stem (ES) cells and studied their differentiation into cardiomyocytes. Strikingly, in vitro cardiomyocyte differentiation of wild type (WT) ES cells faithfully recapitulates in vivo cardiomyocyte differentiation, allowing us to evaluate the phenotypes of Tmod1 knockout (KO) myofibrils irrespective of embryonic lethality of Tmod1 KO mice. Immunofluorescence and electron microscopy studies revealed that Tmod1 null cardiac myocytes were round, morphologically immature, and contained underdeveloped myofibrils that were shorter, narrower, and had fewer thin filaments than those in WT cells. Unexpectedly, clear gaps in the staining pattern for F-actin at the H-zone were detected in most KO cells, indicating the presence of filaments at uniform lengths. This indicates that additional mechanisms other than capping proteins are responsible for thin filament length maintenance in cardiac myocytes. Also unexpectedly, approximately 40% of the KO cardiac myocytes exhibited contractile activity. Our data indicate that differentiating ES cells are a powerful system to investigate the functional properties of contractile proteins and that Tmod1 functions are critical for late stages of myofibrillogenesis, and for the maturation of myofibrils.
Ono
Yasuko
Y
Department of Cell Biology and Anatomy, University of Arizona, 1501 N. Campbell Avenue-LSN 455, Tucson, AZ 85724, USA.
Schwach
Catherine
C
Antin
Parker B
PB
Gregorio
Carol C
CC
D000818:Animals; D015139:Blotting, Southern; D002352:Carrier Proteins / Q000235:genetics*; D002454:Cell Differentiation / Q000235:genetics* / Q000502:physiology; D017931:DNA Primers; D004622:Embryo, Mammalian / Q000166:cytology*; D019084:Fluorescent Antibody Technique, Indirect; D040461:Gene Components; D018390:Gene Targeting; D005822:Genetic Vectors / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000235:genetics*; D008854:Microscopy, Electron; D032383:Myocytes, Cardiac / Q000502:physiology* / Q000648:ultrastructure; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000502:physiology*; D051348:Tropomodulin
D002352:Carrier Proteins; D017931:DNA Primers; D008840:Microfilament Proteins; C495831:Tmod1 protein, mouse; D051348:Tropomodulin