Arf6 guanine nucleotide exchange factor cytohesin-2 binds to CCDC120 and is transported along neurites to mediate neurite growth

J Biol Chem. 2014 Dec 5;289(49):33887-903. doi: 10.1074/jbc.M114.575787. Epub 2014 Oct 17.

Abstract

The mechanism of neurite growth is complicated, involving continuous cytoskeletal rearrangement and vesicular trafficking. Cytohesin-2 is a guanine nucleotide exchange factor for Arf6, an Arf family molecular switch protein, controlling cell morphological changes such as neuritogenesis. Here, we show that cytohesin-2 binds to a protein with a previously unknown function, CCDC120, which contains three coiled-coil domains, and is transported along neurites in differentiating N1E-115 cells. Transfection of the small interfering RNA (siRNA) specific for CCDC120 into cells inhibits neurite growth and Arf6 activation. When neurites start to extend, vesicles containing CCDC120 and cytohesin-2 are transported in an anterograde manner rather than a retrograde one. As neurites continue extension, anterograde vesicle transport decreases. CCDC120 knockdown inhibits cytohesin-2 localization into vesicles containing CCDC120 and diffuses cytohesin-2 in cytoplasmic regions, illustrating that CCDC120 determines cytohesin-2 localization in growing neurites. Reintroduction of the wild type CCDC120 construct into cells transfected with CCDC120 siRNA reverses blunted neurite growth and Arf6 activity, whereas the cytohesin-2-binding CC1 region-deficient CCDC120 construct does not. Thus, cytohesin-2 is transported along neurites by vesicles containing CCDC120, and it mediates neurite growth. These results suggest a mechanism by which guanine nucleotide exchange factor for Arf6 is transported to mediate neurite growth.

Keywords: ADP Ribosylation Factor (ARF); GTPase; Guanine Nucleotide Exchange Factor (GEF); Neurite Outgrowth; Vesicles.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Cell Line
  • Cell Nucleus / metabolism
  • Cytoplasm / metabolism
  • GTPase-Activating Proteins / genetics*
  • GTPase-Activating Proteins / metabolism
  • Gene Expression Regulation
  • Humans
  • Molecular Sequence Data
  • Nerve Tissue Proteins / genetics*
  • Nerve Tissue Proteins / metabolism
  • Neurogenesis / genetics*
  • Neurons / cytology
  • Neurons / metabolism*
  • Protein Binding
  • Protein Structure, Tertiary
  • Protein Transport
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Signal Transduction
  • Transport Vesicles / metabolism*

Substances

  • CCDC120 protein, human
  • GTPase-Activating Proteins
  • Nerve Tissue Proteins
  • RNA, Small Interfering
  • cytohesin-2