Calcium plays a pivotal role in the establishment of the differentiated phenotype in myogenic cells but the involved molecular mechanisms are still matter of debate. Here we studied the effects of exposing L6-C5 myogenic cells to high extracellular Ca2þ concentration ([Ca2þ]o), which induces an increase of intracellular calcium ([Ca2þ]i) without involving Ca2þ release from the intracellular stores but exclusively due to plasma membrane influx (Naro et al., 2003). Exposure of L6-C5 cells to [Ca2þ]o up to 20mM for 30 min, before shifting them into a differentiative medium, induced the appearance of multinucleated, myosin-positive myotubes, much larger than in control cells with an increased protein/DNA ratio. These large myotubes showed nuclear accumulation of the hypertrophy marker GATA-2. The hypertrophic growth of these cells was blocked by cyclosporin A (CsA), FK506, or overexpression of a calcineurin-dominant negative protein, suggesting the involvement in this process of the Ca2þ responsive phosphatase calcineurin. Furthermore, transient exposure of L6-C5 cells to high [Ca2þ]o increased the expression of luciferase reporter driven by myoglobin (Mb) and b-MHC promoters but not IIB-MHC and MCK promoters. Luciferase transcription driven by CK promoter was, instead, enhanced by mobilizing Ca2þ from the intracellular stores. These data indicate that a transient increase of [Ca2þ]i due to plasma-membrane influx is sufficient to induce a hypertrophic phenotype and an increased expression of slow-fiber genes but not fast-fiber genes.

Hypertrophy and transcriptional regulation induced in myogenic cell line L6-C5 by an increase of extracellular calcium.

CANATO, MARTA;REGGIANI, CARLO;
2005

Abstract

Calcium plays a pivotal role in the establishment of the differentiated phenotype in myogenic cells but the involved molecular mechanisms are still matter of debate. Here we studied the effects of exposing L6-C5 myogenic cells to high extracellular Ca2þ concentration ([Ca2þ]o), which induces an increase of intracellular calcium ([Ca2þ]i) without involving Ca2þ release from the intracellular stores but exclusively due to plasma membrane influx (Naro et al., 2003). Exposure of L6-C5 cells to [Ca2þ]o up to 20mM for 30 min, before shifting them into a differentiative medium, induced the appearance of multinucleated, myosin-positive myotubes, much larger than in control cells with an increased protein/DNA ratio. These large myotubes showed nuclear accumulation of the hypertrophy marker GATA-2. The hypertrophic growth of these cells was blocked by cyclosporin A (CsA), FK506, or overexpression of a calcineurin-dominant negative protein, suggesting the involvement in this process of the Ca2þ responsive phosphatase calcineurin. Furthermore, transient exposure of L6-C5 cells to high [Ca2þ]o increased the expression of luciferase reporter driven by myoglobin (Mb) and b-MHC promoters but not IIB-MHC and MCK promoters. Luciferase transcription driven by CK promoter was, instead, enhanced by mobilizing Ca2þ from the intracellular stores. These data indicate that a transient increase of [Ca2þ]i due to plasma-membrane influx is sufficient to induce a hypertrophic phenotype and an increased expression of slow-fiber genes but not fast-fiber genes.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11577/1481793
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