AKAP350 at the Golgi Apparatus

Abstract

AKAP350 can scaffold a number of protein kinases and phosphatases at the centrosome and the Golgi apparatus. We performed a yeast two-hybrid screen of a rabbit parietal cell library with a 3.2-kb segment of AKAP350 (nucleotides 3611-6813). This screen yielded a full-length clone of rabbit chloride intracellular channel 1 (CLIC1). CLIC1 belongs to a family of proteins, all of which contain a high degree of homology in their carboxyl termini. All CLIC family members were able to bind a 133-amino acid domain within AKAP350 through the last 120 amino acids in the conserved CLIC carboxyl termini. Antibodies developed against a bovine CLIC, p64, immunoprecipitated AKAP350 from HCA-7 colonic adenocarcinoma cell extracts. Antibodies against CLIC proteins recognized at least five CLIC species including a novel 46-kDa CLIC protein. We isolated the human homologue of bovine p64, CLIC5B, from HCA-7 cell cDNA. A splice variant of CLIC5, the predicted molecular mass of CLIC5B corresponds to the molecular mass of the 46-kDa CLIC immunoreactive protein in HCA-7 cells. Antibodies against CLIC5B colocalized with AKAP350 at the Golgi apparatus with minor staining of the centrosomes. AKAP350 and CLIC5B association with Golgi elements was lost following brefeldin A treatment. Furthermore, GFP-CLIC5B-(178 -410) targeted to the Golgi apparatus in HCA-7 cells. The results suggest that AKAP350 associates with CLIC proteins and specifically that CLIC5B interacts with AKAP350 at the Golgi apparatus in HCA-7 cells.

c-Src Control of Chloride Channel Support for Osteoclast HCl Transport and Bone Resorption

“…This could result from activation of the chloride channel in pump containing membranes or physical approximation of the active channel and pump. Previously CLIC-4 and -5 proteins and the v-H ϩ -ATPase have been shown to associate with cytoskeleton proteins that are c-Src substrates (42,78,85). In these studies the proton pump is found in cytoskeletal-associated vesicles while the soluble CLIC-proteins directly interact with F-actin-based structures.…”

Identification of chloride intracellular channel proteins in spermatozoa

“…For example, CLIC3 was originally discovered as a binding partner for ERK7, a member of the mitogen-activated protein kinase family [18], and bovine p64 has been shown to associate directly with p59 fyn , a Src family tyrosine kinase [19]. In addition, multiple CLIC family members can associate with AKAP350, a scaffolding protein that sequesters cAMP-dependent protein kinase and several other protein kinases and phosphatases to particular subcellular sites [16,17]. More recently, CLIC6 has been shown to bind scaffolding proteins, such as radixin and multi-PDZ protein, as well as dopamine D 2 -like receptors [24].…”

“…CLICs constitute a new family of putative chloride channel proteins with several distinct members, including bovine p64 and human CLIC1-6 [10,11]. CLIC proteins are localized to a variety of subcellular compartments in various cell types, and although they have been implicated in chloride ion transport [12,13], several reports show that certain family members can associate with the cytoskeleton, scaffolding proteins, and signaling enzymes [14][15][16][17][18][19]. The identification of multiple CLIC proteins in spermatozoa, which may interact with PP1c2, opens a new area of study on the role of CLICs in sperm function.…”

The Organellular Chloride Channel Protein CLIC4/mtCLIC Translocates to the Nucleus in Response to Cellular Stress and Accelerates Apoptosis

“…CLIC4 has several consensus phosphorylation sites, and it interacts with a 14-3-3 isoform (14). Furthermore, recent evidence indicates that CLIC family members interact with the AKAP350 chaperone protein (26). p64 is a substrate for the Fyn kinase (27), and CLIC3 associates with ERK7 (3).…”