Functional dissection of nebulette demonstrates actin binding of nebulin-like repeats and Z-line targeting of SH3 and linker domains

Carole L. Moncman, Kuan Wang

Research output: Contribution to journalArticlepeer-review

50 Citations (Scopus)


Nebulette, a 107 kDa protein associated with the I-Z-I complex of cardiac myofibrils, may play an important role in the assembly of the Z-line. Determination of the complete primary structure of 1011 residue human fetal nebulette reveals a four-domain layout similar to skeletal muscle nebulin: a short N-terminal domain, followed by 22 nebulin-like repeats that are linked to a C-terminal Src homology 3 (SH3) domain via a short linker domain. To elucidate the mechanisms of assembly for nebulette in the Z-line, the complete coding sequence or fusions of nebulette domains with green fluorescent protein (GFP) were expressed in cardiomyocytes and fibroblasts. The complete protein localized to Z-lines in cardiac cells and to dense bodies in nonmuscle cells. The GFP-repeat domain forms bundles that are associated with actin filaments in both cell types and disrupts the microfilament network. In contrast, the GFP-repeat plus linker shows limited interaction with dense bodies in nonmuscle cells and the Z-lines of cardiomyocytes. Interestingly, the tagged linker or SH3 is diffusely distributed in nonmuscle cells, but localizes to the Z-lines in cardiomyocytes. Supporting the cellular localization work, recombinant nebulette fragments bind to actin, tropomyosin, and α-actinin in in vitro binding assays. These results suggest the repeat domain contains actin binding functions and that the linker domain may target this interaction to Z-lines and dense bodies. Our data also indicate that the linker and SH3 domains can distinguish between dense bodies and Z-lines, suggesting that the ligands for their interactions are specific to these muscular substructures.

Original languageEnglish
Pages (from-to)1-22
Number of pages22
JournalCell Motility and the Cytoskeleton
Issue number1
Publication statusPublished - 1999
Externally publishedYes


  • Cardiac muscle
  • Cytoskeleton
  • Green fluorescent protein
  • Myofibrillogenesis
  • Nebulin

ASJC Scopus subject areas

  • Structural Biology
  • Cell Biology


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