Cargo Receptor Traffics Many Matrix Proteins Out of the Endoplasmic Reticulum (ER) by Binding to N‐Terminal IleProVal (IPV)‐Like Motif

Abstract

IntroductionMatrix proteins assemble large complexes in the extracellular environment and may be at risk of forming undesired aggregates during intracellular trafficking. Patients with nonsyndromic Dentinogenesis Imperfecta have mutations in the dentin sialophosphoprotein (DSPP) gene. In half of the cases, the N‐terminal IPV tripeptide is changed (e.g., IPD, ITV). We have shown that IPV‐mutant DSPP inefficiently exits the ER and forms aggregates. Many secreted proteins of dentin (e.g., DSPP, DMP1), enamel (AMELX, AMBN, ENAM), bone (SPARC, OCN, OPN) and cartilage (BGN, DCN, ASPN), as well as enzymes (trypsin) and hormones (GH, LEP, PRL, FGF23) also start with conserved IPV‐like tripeptides. We propose that the same cargo receptor binds to these proteins through their IPV‐like motifs thereby trafficking them out of the ER before self‐ or inter‐molecular aggregates form.MethodsThe ER‐trafficking efficiency of DSPP (and other secreted proteins) expressing a series of single amino acid changes in their IPV motifs was studied in HEK293 and HeLa cells. To identify the cargo receptor, we expressed wildtype and mutant constructs in Saccharomyces cell lines each lacking a single ER‐associated protein.ResultsWe have defined the position/property of amino acids in IPV‐like motif that promote ER exit in yeast and higher eukaryotes. Furthermore, gene deletion of a conserved ER‐associated yeast protein specifically delays ER exit of wildtype yeast and transfected matrix proteins.ConclusionWe have defined the amino acids required to bind the proposed cargo receptor and have identified the yeast's receptor as a highly conserved, ER‐associated protein.Supported by Division of Intramural Research of the NIDCR, NIH.