TAILIEUCHUNG - Báo cáo khoa học: Unusual transfer of CutA into the secretory pathway, evidenced by fusion proteins with acetylcholinesterase

The mouse CutA protein exists as long and short components of 20 and 15 kDa, produced by the use of different in-frame ATGs initiation codons, and by proteolytic cleavage. We recently showed that, surprisingly, the longer, uncleaved component resides mostly in the secretory pathway and is secreted, whereas the shorter component resides mostly in the cytoplasm. | ỊFEBS Journal Unusual transfer of CutA into the secretory pathway evidenced by fusion proteins with acetylcholinesterase Dong Liang1 2 Stephanie Carvalho1 Suzanne Bon1 and Jean Massoulie1 1 Laboratoire de Neurobiologie CNRS UMR 8544 Paris France 2 Key Lab of Brain FunctionalGenomics MOE STCSM Shanghai Institute of Brain FunctionalGenomics China Keywords acetylcholinesterase CutA folding secretion signal peptide Correspondence J. Massoulie Laboratoire de Neurobiologie CNRS UMR 8544 Ecole Normale Superieure 46 rue d Ulm 75005 Paris France Fax 33 1443 23887 Tel 33 1443 23891 E-mail Received 5 December 2008 revised 6 June 2009 accepted 15 June 2009 doi The mouse CutA protein exists as long and short components of 20 and 15 kDa produced by the use of different in-frame ATGs initiation codons and by proteolytic cleavage. We recently showed that surprisingly the longer uncleaved component resides mostly in the secretory pathway and is secreted whereas the shorter component resides mostly in the cytoplasm. To confirm these subcellular localizations we constructed fusion proteins in which the catalytic domain of rat acetylcholinesterase was placed downstream of the CutA variants. The acquisition of an active conformation and N-glycosylation of the fusion proteins proved their transfer into the secretory pathway. We show that the CutA-AChE fusion proteins produced and secreted active N-glycosylated molecules while an AChE mutant lacking its secretory signal peptide did not produce any significant activity. Thus an N-terminal CutA domain actually drives AChE into the endoplasmic reticulum and allows its secretion. This was observed with full length CutA starting at Met1 and at a much lower level with the shorter mutants starting at Met24 and Met44 although the latter is not predicted to possess any signal peptide. These experiments illustrate the value of using AChE as a reporter and reveals an unusual protein .

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