Submitted on February 5, 2008
Revised on May 2, 2008
Accepted on May 18, 2008
mCLCA4 Processing and Secretion Requires Luminal Sorting Motifs
Chunlei Huan1, Kai Su Greene1, Bo Shui, Gwendolyn Spizz1, Haitao Sun1, Robert Doran, Patricia J Fisher, Mark Roberson, Randolph C Elble2, and Michael I Kotlikoff1*
1 Cornell University
2 Southern Illinois University
* To whom correspondence should be addressed. E-mail: mik7{at}cornell.edu.
CLCA proteins are encoded by a family of highly related and clustered genes in mammals that are markedly upregulated in inflammation and have been shown to affect chloride transport. Here we describe the cellular processing and regulatory sequences underlying mCLCA4 proteins. The 125 kD mCLCA4 gene product is cleaved to 90 kD and 40 kD fragments and the N and C -terminal fragments are secreted, where they are found in cell media and associated with the plasma membrane. The 125 kD full-length protein is only found in the ER, and specific luminal diarginine retention and dileucine forward trafficking signals contained within the CLCA4 sequence regulate export from the ER and proteolytic processing. Mutation of the dileucine luminal sequences resulted in ER trapping of the immaturely glycosylated 125 kD peptide, indicating that proteolytic cleavage occurs following recognition of the trafficking motifs. Moreover, the mutated dileucine and diarginine signal sequences directed processing of a secreted form of enhanced green fluorescent protein in a manner consistent with the effects on mCLCA4.
