Functional analysis of CLIP-115 and its binding to microtubules

J Cell Sci. 2000 Jun:113 ( Pt 12):2285-97. doi: 10.1242/jcs.113.12.2285.

Abstract

Cytoplasmic linker proteins (CLIPs) bind to microtubules and are proposed to link this cytoskeletal network to other intracellular structures. We are interested in CLIP-115, since this protein is enriched in neuronal dendrites and may operate in the control of brain-specific organelle translocations. Each CLIP monomer is characterized by two microtubule-binding (MTB) motifs, surrounded by basic, serine-rich regions. This head domain is connected to the C-terminal tail through a long coiled-coil structure. The MTB domains are conserved as a single domain in other proteins involved in microtubule based transport and dynamics, such as p150(Glued). Here we provide evidence that efficient binding of CLIP-115 to microtubules is sensitive to phosphorylation and is not mediated by the conserved MTB domains alone, but requires the presence of the basic, serine rich regions in addition to the MTB motifs. In transfected COS-1 cells, CLIP-115 initially accumulates at the distal ends of microtubules and coincides with CLIP-170, indicating that both proteins mark growing microtubule ends. However, when expressed at higher levels, CLIP-115 and -170 affect the microtubule network differently. This might be partly due to the divergent C-termini of the two proteins. We demonstrate that, similar to CLIP-170, CLIP-115 forms homodimers, which, at least in vitro, are linked by disulfide bridges. Cysteine(391) of CLIP-115, however, is specific in that it controls the microtubule bundling capacity of certain mutant CLIP-115 molecules. Therefore, both similar and specific mechanisms appear to regulate the conformation of CLIPs as well as their binding to microtubules.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Brain / cytology
  • Brain / metabolism*
  • Dimerization
  • Escherichia coli
  • Mice
  • Microtubule-Associated Proteins / metabolism*
  • Microtubules / metabolism*
  • Molecular Sequence Data
  • Nerve Tissue Proteins / metabolism*
  • Neurons / metabolism*
  • Neurons / ultrastructure*
  • Protein Binding

Substances

  • Microtubule-Associated Proteins
  • Nerve Tissue Proteins
  • cytoplasmic linker protein 115