The 160 N-terminal residues of calnexin define a novel region supporting viability in Schizosaccharomyces pombe

Yeast. 2007 Feb;24(2):89-103. doi: 10.1002/yea.1440.

Abstract

Protein secretion is a complex process that can be modulated by folding factors in the endoplasmic reticulum (ER), such as calnexin, a highly-conserved molecular chaperone involved in quality control. In Schizosaccharomyces pombe, calnexin (Cnx1p) is essential for cell viability. The calnexin/Cnx1p determinants required for viability have been mapped within the last 123 residues of its C-terminus. To better understand the role(s) of calnexin/Cnx1p in secretion, we screened for cnx1 mutants 'super-secreting' cellulase. We identified ss14_cnx1, a mutant secreting 10-fold higher levels of the glycoprotein cellulase than the wild-type strain. While cellulase did not interact with ss14_Cnx1p, the ratio of secreted activity/quantity for this enzyme was not affected, suggesting that the quality control of folding in the ER was adequate in the mutant strain. Surprisingly, the ss14_Cnx1p mutant is composed of the 160 N-terminal amino acids of the mature molecule, thus this mutant defines a novel calnexin/Cnx1p region supporting Sz. pombe viability. Interestingly, like viable mutants spanning the last 52 aa of calnexin/Cnx1p, the 160 N-terminal residues encoded by ss14_cnx1 also forms a complex with the essential BiP chaperone. These results reveal the so far unidentified importance of the N-terminal region of calnexin/Cnx1p.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Aspergillus / enzymology
  • Aspergillus / genetics
  • Blotting, Southern
  • Calnexin / genetics
  • Calnexin / physiology*
  • Cellulase / genetics
  • Cellulase / metabolism
  • Cellulase / physiology
  • DNA, Fungal / genetics
  • Fungal Proteins / genetics
  • Fungal Proteins / physiology*
  • Immunoblotting
  • Microscopy, Interference
  • Mutagenesis, Insertional
  • Plasmids / genetics
  • Polymerase Chain Reaction
  • Protein Folding
  • Schizosaccharomyces / enzymology
  • Schizosaccharomyces / genetics
  • Schizosaccharomyces / physiology*

Substances

  • Cnx1 protein, S pombe
  • DNA, Fungal
  • Fungal Proteins
  • Calnexin
  • Cellulase