A novel di-acidic motif facilitates ER export of the syntaxin SYP31.
It is generally accepted that ER protein export is largely influenced by the transmembrane domain (TMD). The situation is unclear for membrane-anchored proteins such as SNAREs, which are anchored to the membrane by their TMD at the C-terminus. For example, in plants, Sec22 and SYP31 (a yeast Sed5 homologue) have a 17 aa TMD but different locations (ER/Golgi and Golgi), indicating that TMD length alone is not sufficient to explain their targeting. To establish the identity of factors that influence SNARE targeting, mutagenesis and live cell imaging experiments were performed on SYP31. It was found that deletion of the entire N-terminus domain of SYP31 blocked the protein in the ER. Several deletion mutants of different parts of this N-terminus domain indicated that a region between the SNARE helices Hb and Hc is required for Golgi targeting. In this region, replacement of the aa sequence MELAD by GAGAG or MALAG retained the protein in the ER, suggesting that MELAD may function as a di-acidic ER export motif EXXD. This suggestion was further verified by replacing the established di-acidic ER export motif DLE of a type II Golgi protein AtCASP and a membrane-anchored type I chimaera, TMcCCASP, by MELAD or GAGAG. The MELAD motif allowed the proteins to reach the Golgi, whereas the motif GAGAG was found to be insufficient to facilitate ER protein export. Our analyses indicate that we have identified a novel and transplantable di-acidic motif that facilitates ER export of SYP31 and may function for type I and type II proteins in plants.
Chatre
Laurent
L
University of Bordeaux 2, Membrane Biogenesis Laboratory, CNRS UMR 5200, 146, rue Léo Saignat, 33076 Bordeaux Cedex, France.
Wattelet-Boyer
Valérie
V
Melser
Su
S
Maneta-Peyret
Lilly
L
Brandizzi
Federica
F
Moreau
Patrick
P
D020816:Amino Acid Motifs; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006056:Golgi Apparatus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D050765:Qa-SNARE Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D050600:SNARE Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
D029681:Arabidopsis Proteins; C504617:CASP protein, Arabidopsis; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; D050600:SNARE Proteins; C544320:SYP31 protein, Arabidopsis