The complete mouse nebulin gene sequence and the identification of cardiac nebulin

Steven T. Kazmierski, Parker B Antin, Christian C. Witt, Norbert Huebner, Abigail S. McElhinny, Siegfried Labeit, Carol Gregorio

Research output: Contribution to journalArticle

62 Citations (Scopus)

Abstract

Nebulin is a giant (Mr 750-850kDa), modular sarcomeric protein proposed to regulate the assembly, and to specify the precise lengths of actin (thin) filaments in vertebrate skeletal muscles. Nebulin's potential role as a molecular template is based on its structural and biochemical properties. Its central ∼700kDa portion associates with actin along the entire length of the thin filament, its N-terminal region extends to thin filament pointed ends, and ∼80kDa of its C-terminal region integrates within the Z-line lattice. Here, we determined the exon/intron organization of the entire mouse nebulin gene, which contains 165 exons in a 202kb segment. We identified 16 novel exons, 15 of which encode nebulin-repeat motifs (12 from its central region and 3 from its Z-line region). One novel exon shares high sequence homology to the 20 residue repeats of the tight-junction protein, ZO-1. RT-PCR analyses revealed that all 16 novel exons are expressed in mouse skeletal muscle. Surprisingly, we also amplified mRNA transcripts from mouse and human heart cDNA using primers designed along the entire length of nebulin. The expression of cardiac-specific nebulin transcripts was confirmed by in situ hybridization in fetal rat cardiomyocytes and in embryonic Xenopus laevis (frog) heart. On the protein level, antibodies specific for skeletal muscle nebulin's N and C-terminal regions stained isolated rat cardiac myofibrils at the pointed and barbed ends of thin filaments, respectively. These data indicate a conserved molecular layout of the nebulin filament systems in both cardiac and skeletal myofibrils. We propose that thin filament length regulation in cardiac and skeletal muscles may share conserved nebulin-based mechanisms, and that nebulin isoform diversity may contribute to thin filament length differences in cardiac and skeletal muscle.

Original languageEnglish (US)
Pages (from-to)835-846
Number of pages12
JournalJournal of Molecular Biology
Volume328
Issue number4
DOIs
StatePublished - May 9 2003

Fingerprint

Genes
Exons
Skeletal Muscle
Myofibrils
Myocardium
Zonula Occludens-1 Protein
nebulin
Xenopus laevis
Sequence Homology
Actin Cytoskeleton
Cardiac Myocytes
Anura
Introns
In Situ Hybridization
Vertebrates
Actins
Protein Isoforms
Proteins
Complementary DNA
Polymerase Chain Reaction

Keywords

  • Heart muscle
  • Myofibril
  • Nebulin
  • Sarcomere
  • Thin filament

ASJC Scopus subject areas

  • Virology

Cite this

The complete mouse nebulin gene sequence and the identification of cardiac nebulin. / Kazmierski, Steven T.; Antin, Parker B; Witt, Christian C.; Huebner, Norbert; McElhinny, Abigail S.; Labeit, Siegfried; Gregorio, Carol.

In: Journal of Molecular Biology, Vol. 328, No. 4, 09.05.2003, p. 835-846.

Research output: Contribution to journalArticle

Kazmierski, Steven T. ; Antin, Parker B ; Witt, Christian C. ; Huebner, Norbert ; McElhinny, Abigail S. ; Labeit, Siegfried ; Gregorio, Carol. / The complete mouse nebulin gene sequence and the identification of cardiac nebulin. In: Journal of Molecular Biology. 2003 ; Vol. 328, No. 4. pp. 835-846.
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