The distribution of synaptotagmin II in RBL-2H3 and its regulation on exocytosis of lysosomes in RBL-2H3.
Synaptotagmin (Syt) constitutes a family of membrane-trafficking proteins, so far nearly 20 Syts have been discovered. Extensive work showed that synatotagmins were a potential Ca2+ sensor for regulated exocytosis. This study was to investigate the expression and location of synaptotagmin II (Syt2) in RBL-2H3 (RBL) and its role in regulating exocytosis of RBL. The expression of Syt2 in RBL was confirmed by Western blot. The recombinant expression vector pEGFP-N1-Syt2 was constructed and transfected into RBL by electroporation, the stable transfectant RBL-Syt2-S expressing fusion protein Syt2-EGFP were obtained and Syt2 was highly concentrated at plasma membrane with little detected in cytoplasm. To analyze the role of Syt2 during exocytosis of RBL, the release of cathepsin D was assayed by immunoblotting. Compared with control, the release of cathepsin D by RBL-Syt2-S was markedly decreased. The results indicated that Syt2 played a negative regulation in exocytosis of lysosomes in RBL.
Zhang
Jicheng
J
Department of Medical Laboratory, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Wu
Jianmin
J
Pan
Shixiu
S
Lv
Wenli
W
D000818:Animals; D015153:Blotting, Western; D002402:Cathepsin D / Q000032:analysis / Q000378:metabolism; D045744:Cell Line, Tumor; D005089:Exocytosis; D015870:Gene Expression; D008247:Lysosomes / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D051381:Rats; D011993:Recombinant Fusion Proteins; D050861:Synaptotagmin II / Q000378:metabolism*; D014162:Transfection
D011993:Recombinant Fusion Proteins; D050861:Synaptotagmin II; D002402:Cathepsin D