RAF-1 kinase inhibitor protein-mediated cholecystokinin-2 receptor desensitization and extracellular signal-regulated kinase activation

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Title: RAF-1 kinase inhibitor protein-mediated cholecystokinin-2 receptor desensitization and extracellular signal-regulated kinase activation
Author: Jeseong Park
Abstract: Raf -1 kinase inhibitor protein (RKIP ) is initially known as a suppressor for Raf -1 -mediated ERK activation . Moreover , recent findings indicate that RKIP also has a role in G -protein -coupled receptor (GPCR ) desensitization . Protein kinase C (PKC ) -mediated phosphorylation at Serine 153 (S153 ) on RKIP switches RKIP association from Raf -1 to GPCR kinase -2 (GRK2 ) for inhibiting GRK2 -mediated G -protein -coupled receptor (GPCR ) desensitization . As a member of the GPCR superfamily , Cholecystokinin -2 receptor (CCK2R ) is a physiological receptor for gastrin (G17 ) and activates extracellular signal -regulated kinase (ERK ) via the PKC activity . The inhibition of classical PKCs (cPKC , PKC - & #945 ; , - & #946 ; , and - & #947 ; ) by Gö6976 indicated the augment in ERK activation compared to vehicle control , suggesting cPKC’s involvement in CCK2R desensitization . CCK2R -mediated ERK activation was significantly decreased when PKC - & #948 ; was selectively silenced by siRNAs , indicating that PKC - & #948 ; , a member of the novel PKC family , mediates CCK2R -induced ERK activation . Furthermore , the data for CCK2R desensitization showed that inhibited cPKC activity by Gö6976 facilitated CCK2R desensitization . However , the silencing for PKC - & #945 ; , - & #946 ; , or – & #948 ; by siRNAs indicated that each knockdown of PKC isozymes attenuated CCK2R desensitization . The PKC involvement in CCK2R desensitization and ERK activation also suggested a potential role of RKIP in regulation of CCK2R activity . By either silencing or overexpressing RKIPs , I proved that RKIP acts as a suppressor for CCK2R desensitization , and the phosphorylation at S153 on RKIP plays a crucial role for inhibiting desensitization . The RKIP -mediated inhibition of CCK2R desensitization also resulted in augmentation of receptor -induced ERK activation , and this finding indicates that RKIP acts as a modulator for CCK2R -mediated signaling . The mechanism for RKIP -mediated receptor desensitization was investigated by co -immunoprecipitation of GRK2 with RKIP . The data indicated that RKIP strongly associated onto GRK2 when PKC was activated by phorbol 12 -myristate 13 -acetate (PMA ) treatment , but either G17 stimulation or Gö6976 did not affect on the association . It suggests that PMA -sensitive PKC isozymes are responsible for RKIP phosphorylation ; however , CCK2R -mediated PKC isozymes are not involved in RKIP phosphorylation directly , rather PKC activation by other cellular mechanisms mediate RKIP phosphorylation resulting in GRK2 association . Therefore , I conclude that RKIP mediates CCK2R desensitization and ERK activation through PKC activation .
URI: http : / /hdl .handle .net /2152 .3 /160
Date: 2009-06-29

Citation

RAF-1 kinase inhibitor protein-mediated cholecystokinin-2 receptor desensitization and extracellular signal-regulated kinase activation. Doctoral dissertation, The University of Texas Medical Branch. Available electronically from http : / /hdl .handle .net /2152 .3 /160 .

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