Mutational analysis of betaCOP (Sec26p) identifies an appendage domain critical for function

BMC Cell Biol. 2008 Jan 22;9:3. doi: 10.1186/1471-2121-9-3.

Abstract

Background: The appendage domain of the gammaCOP subunit of the COPI vesicle coat bears a striking structural resemblance to adaptin-family appendages despite limited primary sequence homology. Both the gammaCOP appendage domain and an equivalent region on betaCOP contain the FxxxW motif; the conservation of this motif suggested the existence of a functional appendage domain in betaCOP.

Results: Sequence comparisons in combination with structural prediction tools show that the fold of the COOH-terminus of Sec26p is strongly predicted to closely mimic that of adaptin-family appendages. Deletion of the appendage domain of Sec26p results in inviability in yeast, over-expression of the deletion construct is dominant negative and mutagenesis of this region identifies residues critical for function. The ArfGAP Glo3p was identified via suppression screening as a potential downstream modulator of Sec26p in a manner that is independent of the GAP activity of Glo3p but requires the presence of the COOH-terminal ISS motifs.

Conclusion: Together, these results indicate an essential function for the predicted betaCOP appendage and suggest that both COPI appendages perform a biologically active regulatory role with a structure related to adaptin-family appendage domains.

Publication types

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

MeSH terms

  • Adaptor Protein Complex alpha Subunits / genetics
  • Adaptor Protein Complex alpha Subunits / metabolism
  • Amino Acid Motifs / genetics
  • Base Sequence / genetics
  • COP-Coated Vesicles / genetics*
  • COP-Coated Vesicles / metabolism*
  • COP-Coated Vesicles / ultrastructure
  • Coatomer Protein / chemistry*
  • Coatomer Protein / genetics*
  • Endoplasmic Reticulum, Rough / metabolism
  • GTPase-Activating Proteins / genetics
  • GTPase-Activating Proteins / metabolism
  • Gene Expression Regulation, Fungal / genetics
  • Golgi Apparatus / metabolism
  • Mutagenesis, Site-Directed
  • Mutation / genetics
  • Protein Structure, Tertiary / genetics
  • Protein Transport / physiology
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / chemistry*
  • Saccharomyces cerevisiae Proteins / genetics*
  • Saccharomyces cerevisiae Proteins / metabolism

Substances

  • Adaptor Protein Complex alpha Subunits
  • Coatomer Protein
  • GTPase-Activating Proteins
  • Glo3 protein, S cerevisiae
  • SEC26 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins