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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

Insights

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.

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.

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