Analysis of Nesprin-2 Interaction with Its Binding Partners and Actin

Methods Mol Biol. 2018:1840:35-43. doi: 10.1007/978-1-4939-8691-0_4.

Abstract

Nuclei are connected to the actin cytoskeleton for controlling its position in the cell and for mechanochemical signaling. Nesprin-2G is one of the major outer nuclear membrane proteins that links the nucleus to the actin cytoskeleton. In addition to its paired calponin homology (CH) domains, nesprin-2G interacts with actin filaments by binding the actin-bundling proteins FHOD1 and fascin. We describe methods to measure the interaction of nesprin-2G with actin filaments using an actin co-sedimentation assay and with its binding partner FHOD1 using a GST pull-down method.

Keywords: Actin filaments; Calponin homology domains; FHOD1; Fascin; LINC complex; Spectrin repeats.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Actins / metabolism*
  • Animals
  • Carrier Proteins / chemistry
  • Carrier Proteins / metabolism*
  • Fetal Proteins / chemistry
  • Fetal Proteins / metabolism
  • Formins
  • Humans
  • Mice
  • Nerve Tissue Proteins / chemistry
  • Nerve Tissue Proteins / metabolism*
  • Nuclear Proteins / chemistry
  • Nuclear Proteins / metabolism*
  • Protein Binding
  • Protein Interaction Domains and Motifs
  • Recombinant Fusion Proteins

Substances

  • Actins
  • Carrier Proteins
  • FHOD1 protein, mouse
  • Fetal Proteins
  • Formins
  • Nerve Tissue Proteins
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • Syne2 protein, mouse