Phorbol-induced surface expression of NR2A subunit homologues in HEK293 cells  

作　　者：Chan-ying ZHENG Xiu-juan YANG Zhan-yan FU Jian-hong LUO 

机构地区：[1]Department of Neurobiology, Zhejiang University School of Medicine, Hangzhou 310031, China [2]Institute of Neuroscience, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai 200031, China [3]Department of Physiology and Biophysics, Georgetown University School of Medicine, Washington 20057, USA

出　　处：《Acta Pharmacologica Sinica》2006年第12期1580-1585,共6页中国药理学报（英文版）

基　　金：Project supported by the National Basic Research Program of China(G2002CB713808);the National Natural Science Foundation of China(№30270436 and 30470547);by the Outstanding Cross-century Faculty Grant from the Ministry of Education of China.

摘　　要：Aim： N-methyl-D-aspartate receptors （NMDAR） are heteromeric complexes primarily assembled from NR1 and NR2 subunits. In normal conditions, NR2 subunits assemble into homodimers in the endoplasmic reticulum （ER）. These homodimers remain in the ER until they coassemble with NR1 dimers and are trafficked to the cell surface. However, it still remains unclear whether functional homomeric NMDAR exist in physiological or pathological conditions. Methods： We transfected GFP-NR2A alone into HEK293 cells, treated the cells with PKC activator 12-myristate-13 acetate （PMA）, and then detected surface NR2A subunits with a live cell immunostaining method. We also used a series of NR2A mutants with a partial deletion of its C-terminus to identify the regions that are involved in the PMA-mediated surface expression of NR2A subunits. Results： NR2A subunits were expressed on the cell membrane after incubation with PMA （200 nmol/L, 30 min）, although no functional NMDA channels were detected after PMA-induced membrane trafficking. Immunostaining with an ER marker also revealed that NR2A subunits were exported from the ER after PMA treatment. Furthermore, the deletion of amino acids between 1149-1347 or 1354-1464 of NR2A inhibited PMA-induced surface expression of NR2A subunits. Conclusion： First, our data suggests that PMA treatment can induce the surface expression of homomeric NR2A subunits. Furthermore, this process is probably mediated by the NR2A C-terminal region between positions 1149 and 1464.Aim： N-methyl-D-aspartate receptors （NMDAR） are heteromeric complexes primarily assembled from NR1 and NR2 subunits. In normal conditions, NR2 subunits assemble into homodimers in the endoplasmic reticulum （ER）. These homodimers remain in the ER until they coassemble with NR1 dimers and are trafficked to the cell surface. However, it still remains unclear whether functional homomeric NMDAR exist in physiological or pathological conditions. Methods： We transfected GFP-NR2A alone into HEK293 cells, treated the cells with PKC activator 12-myristate-13 acetate （PMA）, and then detected surface NR2A subunits with a live cell immunostaining method. We also used a series of NR2A mutants with a partial deletion of its C-terminus to identify the regions that are involved in the PMA-mediated surface expression of NR2A subunits. Results： NR2A subunits were expressed on the cell membrane after incubation with PMA （200 nmol/L, 30 min）, although no functional NMDA channels were detected after PMA-induced membrane trafficking. Immunostaining with an ER marker also revealed that NR2A subunits were exported from the ER after PMA treatment. Furthermore, the deletion of amino acids between 1149-1347 or 1354-1464 of NR2A inhibited PMA-induced surface expression of NR2A subunits. Conclusion： First, our data suggests that PMA treatment can induce the surface expression of homomeric NR2A subunits. Furthermore, this process is probably mediated by the NR2A C-terminal region between positions 1149 and 1464.

关 键 词：NMDA receptor NR2A subunit PMA surface expression HEK293 cells 

分 类 号：Q2[生物学—细胞生物学]