Caveolins (Cavs) are ~20 kDa scaffolding protein that assemble while oligomeric complexes in lipid raft domains to create caveolae, flask-shaped plasma membrane (PM) invaginations. in major skeletal or cardiac muscle tissue isolates or in the immortalized cardiac myoblastic cell lines H9C2 (rat) or HL-1 (mouse). EPZ-6438 manufacturer A few of these cells offer null (or near-null) backgrounds for Cav manifestation; for instance, MCF-7 cells possess suprisingly low Cav1 manifestation, do not communicate cavins, and don’t type caveolae and Personal computer3 cells communicate Cav1 at high amounts but usually do not communicate cavins (74, 93, 210). MDCK, A431, and MEF cells all communicate Cav1, Cavins 1C3, and type caveolae in the PM (13, 74, 193, 210). The variety of mobile backgrounds in study on caveolae can donate to variations in results; for instance, in research of polarized epithelial cells, basolateral and apical membranes perform specific EPZ-6438 manufacturer tasks in endo/exocytosis, signal transduction, mechanosensation, adherence, junction formation, and/or migration compared with studies of caveolae in a sessile adipocyte, contractile primary skeletal muscle cell, or HEK293 cell (70). Cav1 and Cav3 may differ in their biogenesis of caveolae. However, most data on caveolae formation are based on studies of Cav1-expressing systems, in which the basic components and pathway of caveolae formation appear to be similar among different cell types and thus are the focus of this review. Early steps in caveolae assembly: membrane insertion, 8S-Cav oligomerization, and endoplasmic reticulum to Golgi transition. Caveolae are constructed in a complex, stepwise assembly process that EPZ-6438 manufacturer involves endoplasmic reticulum (ER) membrane insertion, oligomerization, and export of Cav; Golgi Cav-cholesterol association, oligomerization, and export to the PM; palmitoylation near the PM; and addition of cavins at the PM to form invaginated structures (reviewed in refs 144, 173) (Fig. 3). The timeline from Cav translation to cavin addition at the PM in CV1 cells includes ER exit site (ERES) localization and initial oligomer (8S-Cav) formation within 5 min; Golgi localization within 15 min; secondary oligomer (70S-Cav) formation, cholesterol addition, and PM translocation by ~60 min; and accumulation of cavin Rabbit Polyclonal to CAMKK2 at PM 70S-Cav oligomers that occurs over 25 min (68). Cavs are the primary protein components of caveolae, influencing membrane composition and protein content EPZ-6438 manufacturer from translation to degradation. Thus, the translation and early oligomerization of Cavs begin the caveolae biogenesis pathway. Open in a separate window Fig. 3. Maturation of caveolae. Cav1 and Cav2 monomers are cotranslationally inserted into the endoplasmic reticulum (ER) membrane and swiftly oligomerized into 8S-Cav oligomers containing 7C14 Cavs. These oligomers are transported to ER exit sites (ERES) within 5 min of synthesis for COPII-dependent transport to the Golgi apparatus 15 min postsynthesis. In the Golgi, cholesterol crystalizes in the membrane and assists in the formation of 70S-Cav complexes composed of 18C25 8S-Cav subunits by ~60 min after synthesis, whereupon 70S-Cav is transported to the PM by four phosphate-adapter protein (FAPP-1, -2)-dependent secretory vesicles. Near or on the PM, palmitoyl acyltransferases palmitoylate 70S-Cav oligomers. Also on the PM, cavin proteins that trimerize in the EPZ-6438 manufacturer cytosol gradually aggregate on the 70S-Cav membrane over the course of more than 25 min and assist in membrane curvature. Mature caveolae consist of three layers: a cholesterol and anionic lipid-rich membrane embedded with a palmitoylated 70S-Cav coat, which is surrounded by a striated oligomerized 60S-Cavin coat. Formation of caveolae is initiated by the cotranslational ER membrane insertion of monomeric Cav1 in a signal recognition particle-dependent manner (68, 123). As quickly as 5 min after synthesis in the ER, Cav1 forms 8S20,w oligomers (8S-Cav, 150C200 kDa with a minor species around 443C669 kDa) that is estimated to contain 7C14 Cavs in a Cav1:Cav2 ratio of 2C4:1 (41, 68, 111, 123, 181, 184). The formation of 8S-Cav oligomers depends on aa residues Cav166C70, Cav181C100 (i.e., the CSD), and Cav1134C178 (i.e., the COOH-terminus) (41, 113, 188). No molecular chaperone has been identified for the formation of 8S-Cav. 8S-Cav complexes translocate to the ERES, where reputation of the NH2-terminal diacidic series theme, Asp-X-Glu (Cav167DFE69), leads to COPII vesicle-dependent export towards the Golgi by ~15 min after synthesis (68, 124, 137). Disruption of.