Compared to animals, evolution of plant calcium (Ca) physiology has led to a loss of proteins for influx and small ligand-operated control of cytosolic Ca, leaving many Ca mechanisms unaccounted for. Here, we show a mechanism for sorting and activation of glutamate receptor-like channels (GLRs) by CORNICHON HOMOLOG (CNIH) proteins. Single mutants of pollen-expressed GLRs (GLRs) showed growth and Ca flux phenotypes expected for plasma membrane Ca channels. However, higher-order mutants of GLR3.3 revealed phenotypes contradicting this assumption. These discrepancies could be explained by subcellularGLR localization, and we explored the implication of CNIHs in this sorting. We found thatGLRs interact with CNIH pairs, yielding specific intracellular localizations.CNIHs further trigger GLR activity in mammalian cells without any ligand. These results reveal a regulatory mechanism underlying Ca homeostasis by sorting and activation of GLRs byCNIHs.

HighlightThe cargo receptor cornichon, located in the endoplasmic reticulum, interacts with the low-affinity Na+ transporter OsHKT1;3 for its delivery to the Golgi apparatus.

The export of membrane proteins along the secretory pathway is initiated at the endoplasmic reticulum after proteins are folded and packaged inside this organelle by their recruiting into the coat complex COPII vesicles. It is proposed that cargo receptors are required for the correct transport of proteins to its target membrane, however, little is known about ER export signals for cargo receptors. Erv14/Cornichon belong to a well conserved protein family in Eukaryotes, and have been proposed to function as cargo receptors for many transmembrane proteins. Amino acid sequence alignment showed the presence of a conserved acidic motif in the C-terminal in homologues from plants and yeast. Here, we demonstrate that mutation of the C-terminal acidic motif from ScErv14 or OsCNIH1, did not alter the localization of these cargo receptors, however it modified the proper targeting of the plasma membrane transporters Nha1p, Pdr12p and Qdr2p. Our results suggest that mistargeting of these plasma membrane proteins is a consequence of a weaker interaction between the cargo receptor and cargo proteins caused by the mutation of the C-terminal acidic motif.