perbb3 antibody  (Cell Signaling Technology Inc)

Journal: Glia

Article Title: Schwann cell-specific deletion of the endosomal PI 3-kinase Vps34 leads to delayed radial sorting of axons, arrested myelination, and abnormal ErbB2-ErbB3 tyrosine kinase signaling

doi: 10.1002/glia.23173

Figure Lengend Snippet: Proposed model for the role of Vps34 in endo-lysosomal trafficking of ErbB2/3 in Schwann cells. Binding of axonal Nrg1-III to ErbB3 induces receptor heterodimerization and stimulates ErbB2 kinase activity. ErbB2 phosphorylates both itself and ErbB3, creating phosphotyrosine-based docking sites for downstream signaling proteins (lightning bolt). Activation of ErbB2/3 triggers internalization by clathrin-mediated endocytosis. Fusion of ErbB2/3 containing vesicles with early endosomes requires PI3P, which is produced by Vps34, and requires EEA1 and Rab5 (Step 1) (Simonsen et al., 1998). Internalized ErbB2/3 heterodimers likely continue to signal from endosomal compartments, and can be recycled from endosomes to the cell membrane. On endosomes, ubiquitination (Ub) of ErbB2/3 targets the heterodimer for sorting into multivesicular bodies (MVBs). In a second Vps34-dependent step, PI3P recruits Hrs, which binds concomitantly to ubiquitin on ErbB2/3. Hrs recruits the ESCRT complex, which drives the budding event that generates the receptor-containing internal vesicles of MVBs (Step 2) (Fernandez-Borja et al., 1999). Once in MVBs, ErbB2/3 signaling is terminated (Lee et al., 2017). PI3P also mediates early-to-late endosome maturation through the recruitment of Rab7 (Poteryaev et al., 2010) (Step 3). ErbB2/3 receptors are degraded when late endosomes fuse with LAMP1-positive lysosomes (Lee et al., 2017). We propose that, in Vps34−/− Schwann cells the depletion of PI3P results in a slowing or blockade of trafficking Steps 1, 2, and 3 (red dashed lines). Such trafficking defects are predicted to reduce the fusion of ErbB2/3-bearing endocytic vesicles with early endosomes (Step 1), and prevent the sorting of phosphorylated ErbB2/3 into MVBs (Step 2). This proposed abnormal trafficking of ErbB2/3 is consistent with our observations of both enhanced phosphorylation of ErbB3 on Y1289 and altered posttranslational modification of ErbB2 in Vps34SCKO peripheral nerves. Vps34−/− Schwann cells are also likely deficient in Rab7-dependent late endosome maturation (Step 3). Finally, abnormal MVBs/late endosomes may fuse inefficiently with lysosomes (Step 4), resulting in lysosome accumulation.

Article Snippet: Rabbit monoclonal antibodies (mAb) for Vps34, EEA1, Rab5, ErbB3, pErbB3 (Tyr1289), ErbB2, S6, pS6 (Ser235/236), pAkt (Ser473), pAkt (Thr308), and pJNK (Thr183/Tyr185) were from Cell Signaling Technology.

Techniques: Binding Assay, Activity Assay, Activation Assay, Produced, Membrane, Ubiquitin Proteomics, Phospho-proteomics, Modification