rabbit polyclonal antibodies against arp2  (Cell Signaling Technology Inc)

Journal: PLoS ONE

Article Title: The Function of Cortactin in the Clustering of Acetylcholine Receptors at the Vertebrate Neuromuscular Junction

doi: 10.1371/journal.pone.0008478

Figure Lengend Snippet: Cultured embryonic Xenopus muscle cells labeled with rhodamine-α-bungarotoxin (R-BTX) were stimulated overnight with polystyrene beads coated with heparan-binding growth-associated molecule (HB-GAM) (A, D; asterisks) to induce AChR clusters (C, F). Cells were then fixed and labeled with affinity-purified polyclonal antibodies against the Arp2/3 complex proteins Arp2 (B) and p34arc (E) followed by FITC-linked anti-rabbit secondary antibodies. Separately, bead-stimulated muscle cells (G) were labeled with anti-p34arc polyclonal (H) and anti-cortactin monoclonal (I; mAb4F11) antibodies and then FITC-conjugated anti-rabbit and rhodamine-conjugated anti-mouse secondary antibodies. AChRs, Arp2 and p34arc were clustered at bead-muscle contacts (A-F; arrows) where cortactin localized and overlapped in distribution with p34arc (H and I; arrows). In primary muscle cultures non-muscle cells were occasionally found (J) and in these cells p34arc (K) and cortactin (L) localized along the cell periphery (arrowheads) but were not clustered at bead-cell contacts (“b” in K and L corresponds to bead indicated by asterisk in J).

Article Snippet: These reagents were purchased: rhodamine-conjugated α-bungarotoxin (R-BTX) (Molecular Probes; Eugene, OR, USA); monoclonal antibodies against cortactin (4F11) and phosphotyrosine (4G10) and a rabbit polyclonal antibody against p34arc (Upstate Biotechnology; Lake Placid, NY, USA); rabbit polyclonal antibodies against cortactin phosphorylated on Y421, Y466 or Y482 (Cell Signaling Technology; Danvers, MA, USA); rabbit polyclonal antibodies against Arp2 and Y390-phosphorylated AChR β-subunit (Santa Cruz Biotechnology; Santa Cruz, CA, USA); monoclonal anti-Shp2 and anti-neurexin-1 antibodies (BD Biosciences; San Jose, CA, USA); monoclonal anti-α-tubulin antibody DM1A (Sigma; St Louis, MO, USA); rhodamine- and FITC-conjugated secondary antibodies (Zymed; South San Francisco, CA, USA); horseradish-peroxidase (HRP)-conjugated secondary antibodies (Jackson Immuno Research Laboratories; West Grove, PA, USA); and Triton X-100 (TX-100) and West Pico enhanced chemiluminescence (ECL) reagent (Pierce; Rockford, IL, USA).

Techniques: Cell Culture, Labeling, Binding Assay, Affinity Purification

Journal: PLoS ONE

Article Title: The Function of Cortactin in the Clustering of Acetylcholine Receptors at the Vertebrate Neuromuscular Junction

doi: 10.1371/journal.pone.0008478

Figure Lengend Snippet: Activation of MuSK by agrin induces AChR clustering in an actin polymerization-dependent manner. This model depicts a possible way in which cortactin signaling might promote the AChR clustering process. Initiation of intracellular signaling by the activated MuSK complex could enhance cortactin's tyrosine phosphorylation through src family tyrosine kinases (SFKs) (and possibly other kinases such as abl), and cortactin, in turn, could increase actin polymerization. Alternatively, cortactin might trigger actin polymerization by activating the Arp2/3 complex, either on its own or in concert with WASP-related proteins (N-WASP, WIP, etc.) to which it could be linked by the adapter Nck. In parallel, via other signaling intermediates, MuSK could stimulate Rho-family GTPases and, through them, F-actin assembly. Such enhanced and dynamic actin polymerization at synaptic sites could generate a scaffold which “traps” AChRs through rapsyn.

Article Snippet: These reagents were purchased: rhodamine-conjugated α-bungarotoxin (R-BTX) (Molecular Probes; Eugene, OR, USA); monoclonal antibodies against cortactin (4F11) and phosphotyrosine (4G10) and a rabbit polyclonal antibody against p34arc (Upstate Biotechnology; Lake Placid, NY, USA); rabbit polyclonal antibodies against cortactin phosphorylated on Y421, Y466 or Y482 (Cell Signaling Technology; Danvers, MA, USA); rabbit polyclonal antibodies against Arp2 and Y390-phosphorylated AChR β-subunit (Santa Cruz Biotechnology; Santa Cruz, CA, USA); monoclonal anti-Shp2 and anti-neurexin-1 antibodies (BD Biosciences; San Jose, CA, USA); monoclonal anti-α-tubulin antibody DM1A (Sigma; St Louis, MO, USA); rhodamine- and FITC-conjugated secondary antibodies (Zymed; South San Francisco, CA, USA); horseradish-peroxidase (HRP)-conjugated secondary antibodies (Jackson Immuno Research Laboratories; West Grove, PA, USA); and Triton X-100 (TX-100) and West Pico enhanced chemiluminescence (ECL) reagent (Pierce; Rockford, IL, USA).

Techniques: Activation Assay