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. Author manuscript; available in PMC: 2014 Sep 1.
Published in final edited form as: Mol Cell Neurosci. 2013 Jul 10;0:283–297. doi: 10.1016/j.mcn.2013.07.002

Figure 3. Cav-1 and Cav-3 are associated with cytoskeletal fractions in BV2 and PMG.

Figure 3

A. Detergent resistant fractions (DRM) from HyC and D10% cells – top panel shows Cav-1 is found in DRM f4–6. Bottom panel for Cav-3 shows a minimal amount of Cav-3 protein in f5 with the majority of detectable Cav-3 in the heaviest non-lipid raft fractions (f11–12). B. HyC cells have less flotillin-1 and increased flotillin-2 compared to D10% cells. C. In a semi-quantitive assay based on total protein, HyC cells show decreased tubulin and slightly increased actin whereas D10% cells show the converse pattern. Blots are normalized to HSP90, since GAPDH was too abundant for accurate normalization on the stripped blots. D. Qproteome cell compartmentation assay was used to determine Cav-1 and Cav-3 cellular distribution between cytosol, membrane, nuclear and cytoskeletal compartments in BV2 cells – HyC (right) D10% (left). In either condition, Cav-1 distributes to the membrane component (lane 2). Cav-3 is predominantly distributed to the cytoskeleton component (lane 4). Tubulin is confined to cytosolic and membrane fractions, while actin is distributed across the fractions. E. Qproteome cell compartmentation assay was used to determine Cav-1 and Cav-3 cellular distribution between cytosol, membrane, nuclear and cytoskeletal compartments in PMG – HyC (right) D10% (left). Cav-1 is found in membrane fractions, cytosolic fractions and also distributes to the heavy cytoskeletal fractions, along with tubulin, unlike the exclusively membrane localized Cav-1 in BV2. Cav-3 is found exclusively in the cytosol and cytoskeletal fractions. As with BV2 cells, actin is found in all fractions, but a shift to cytosolic fractions is seen in the D10% cells.