title: Regulation of nonmuscle myosin II during 3-methylcholanthrene induced dedifferentiation of C2C12 myotubes.
abstract: 3-Methylcholanthrene (3MC) induces tumor formation at the site of injection in the hind leg of mice within 110 days. Recent reports reveal that the transformation of normal muscle cells to atypical cells is one of the causes for tumor formation, however the molecular mechanism behind this process is not well understood. Here, we show in an in vitro study that 3MC induces fragmentation of multinucleate myotubes into viable mononucleates. These mononucleates form colonies when they are seeded into soft agar, indicative of cellular transformation. Immunoblot analysis reveals that phosphorylation of myosin regulatory light chain (RLC20) is 5.6±0.5 fold reduced in 3MC treated myotubes in comparison to vehicle treated myotubes during the fragmentation of myotubes. In contrast, levels of myogenic factors such as MyoD, Myogenin and cell cycle regulators such as Cyclin D, Cyclin E1 remain unchanged as assessed by real-time PCR array and reverse transcriptase PCR analysis, respectively. Interestingly, addition of the myosin light chain kinase inhibitor, ML-7, enhances the fragmentation, whereas phosphatase inhibitor perturbs the 3MC induced fragmentation of myotubes. These results suggest that decrease in RLC20 phosphorylation may be associated with the fragmentation step of dedifferentiation.
authors.affiliation: Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 32, India.
authors.forename: Sumit K
authors.initials: SK
authors.lastname: Dey
authors.affiliation: Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 32, India.
authors.forename: Shekhar
authors.initials: S
authors.lastname: Saha
authors.affiliation: Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 32, India.
authors.forename: Provas
authors.initials: P
authors.lastname: Das
authors.affiliation: Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 32, India.
authors.forename: Mahua R
authors.initials: MR
authors.lastname: Das
authors.affiliation: Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 32, India. Electronic address: bcssj@iacs.res.in.
authors.forename: Siddhartha S
authors.initials: SS
authors.lastname: Jana
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects*; D007124:Immunoenzyme Techniques; D008748:Methylcholanthrene / Q000494:pharmacology*; D051379:Mice; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D024462:Myosin Type II / Q000235:genetics / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D012333:RNA, Messenger; D008748:Methylcholanthrene; D009219:Myosin-Light-Chain Kinase; D024462:Myosin Type II
keywords: 3-Methylcholanthrene; Dedifferentiation; Fragmentation; Myosin regulatory light chain; Myotubes; Nonmuscle myosin II