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mCLCA4 ER processing and secretion requires luminal sorting motifs.

Chunlei Huan1, Kai Su Greene, Bo Shui

  • 1Biomedical Sciences Department, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853-6401, USA.

American Journal of Physiology. Cell Physiology
|May 23, 2008
PubMed
Summary

Calcium-activated chloride channel (CLCA) proteins, crucial for chloride transport, are regulated by specific sequences. Murine CLCA4 protein processing and secretion are controlled by luminal diarginine and dileucine trafficking signals.

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Calcium-activated chloride channel (CLCA) proteins are implicated in chloride transport and are upregulated during inflammation.
  • The CLCA gene family exhibits high relatedness and clustering in mammals.

Purpose of the Study:

  • To elucidate the cellular processing and regulatory sequences of murine (m) CLCA4 proteins.
  • To understand the mechanisms governing mCLCA4 secretion and proteolytic cleavage.

Main Methods:

  • Analysis of mCLCA4 protein processing and localization.
  • Site-directed mutagenesis of putative trafficking signals (diarginine and dileucine motifs).
  • Expression of modified enhanced green fluorescent protein (eGFP) to assess signal function.

Main Results:

  • The full-length 125-kDa mCLCA4 protein is retained in the endoplasmic reticulum (ER).
  • Proteolytic cleavage occurs, yielding 90- and 40-kDa secreted fragments found in cell media and associated with the plasma membrane.
  • Luminal diarginine retention and dileucine forward trafficking signals regulate ER export and processing; mutations trap the protein in the ER.

Conclusions:

  • Specific luminal sequences (diarginine and dileucine) dictate mCLCA4 ER export and proteolytic processing.
  • These trafficking motifs are critical for the proper maturation and secretion of mCLCA4.
  • The identified signals can direct the processing of other secreted proteins, suggesting a conserved mechanism.