Expression and localization of myotonic dystrophy protein kinase in human skeletal muscle cells determined with a novel antibody: Possible role of the protein in cytoskeleton rearrangements during differentiation

Pascale Beffy, Cristina Barsanti, Renata Del Carratore, Silvana Simi, Pier Alberto Benedetti, Luca Benzi, Alessandro Prelle, Patrizia Ciscato, Marcella Simili

Research output: Contribution to journalArticle

Abstract

Myotonic dystrophy is a multisystemic disorder, due to a CTG triplet expansion at the 3'UTR of the DM1 gene encoding for myotonic dystrophy protein kinase. Recent studies indicate that decreased DMPK levels could account for part of the symptoms suggesting a role of this protein in skeletal muscle differentiation. To investigate this aspect, polyclonal antibodies were raised against two peptides of the catalytic domain and against the human full-length DMPK (DMFL). In western blots, anti-hDMFL antibody was able to detect low amounts of purified human recombinant protein and recognized the splicing isoforms in heart and stomach of overexpressing mice. In human muscle extracts, this antibody specifically recognized a protein of apparent molecular weight of 85 kDa and it specifically stained neuromuscular junctions in skeletal muscle sections. In contrast, both anti-peptide antibodies demonstrated low specificity for either denatured or native DMPK, suggesting that these two epitopes are probably cryptic sites. Using anti-hDMFL, the expression and localization of DMPK was studied in human skeletal muscle cells (SkMC). Western blot analysis indicated that the antibody recognizes a main protein of apparent MW of 75 kDa, which appears to be expressed during differentiation into myotubes. Immunolocalization showed low levels of DMPK in the cytoplasm of undifferentiated cells; during differentiation the staining became more intense and was localized to the terminal part of the cells, suggesting that DMPK might have a role in cell elongation and fusion.

Original languageEnglish
Pages (from-to)742-753
Number of pages12
JournalCell Biology International
Volume29
Issue number9
DOIs
Publication statusPublished - Sep 2005

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Cytoskeleton
Muscle Cells
Skeletal Muscle
Antibodies
Anti-Idiotypic Antibodies
Proteins
Western Blotting
Protein Splicing
Myotonic Dystrophy
Cell Fusion
Neuromuscular Junction
Skeletal Muscle Fibers
3' Untranslated Regions
Recombinant Proteins
Epitopes
Catalytic Domain
Stomach
Protein Isoforms
Cytoplasm
Molecular Weight

Keywords

  • Antibodies
  • Differentiation
  • Myotonic dystrophy protein kinase
  • Skeletal muscle cells

ASJC Scopus subject areas

  • Cell Biology

Cite this

Expression and localization of myotonic dystrophy protein kinase in human skeletal muscle cells determined with a novel antibody : Possible role of the protein in cytoskeleton rearrangements during differentiation. / Beffy, Pascale; Barsanti, Cristina; Del Carratore, Renata; Simi, Silvana; Benedetti, Pier Alberto; Benzi, Luca; Prelle, Alessandro; Ciscato, Patrizia; Simili, Marcella.

In: Cell Biology International, Vol. 29, No. 9, 09.2005, p. 742-753.

Research output: Contribution to journalArticle

Beffy, Pascale ; Barsanti, Cristina ; Del Carratore, Renata ; Simi, Silvana ; Benedetti, Pier Alberto ; Benzi, Luca ; Prelle, Alessandro ; Ciscato, Patrizia ; Simili, Marcella. / Expression and localization of myotonic dystrophy protein kinase in human skeletal muscle cells determined with a novel antibody : Possible role of the protein in cytoskeleton rearrangements during differentiation. In: Cell Biology International. 2005 ; Vol. 29, No. 9. pp. 742-753.
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