phospho src y416 py416 src antibody  (Cell Signaling Technology Inc)

Journal: Life Science Alliance

Article Title: Tyrosine phosphorylation of lamin A by Src promotes disassembly of nuclear lamina in interphase

doi: 10.26508/lsa.202101120

Figure Lengend Snippet: (A) MCF7 and MDA-MB-231 cells were treated with (+) or without (−) 50 nM dasatinib for 1 h and then lysed. Equal amounts of the whole cell lysates (WCLs) were incubated with anti-lamin A/C or pre-immune serum (IgG) as the control. The immunonocomplexes were analyzed by immunoblotting (IB) with anti-PY or anti-lamin A/C antibodies. An equal amount of WCLs was analyzed by immunoblotting with anti-Src, anti-Src pY416, or anti-actin. The tyrosine phosphorylation of lamin A was quantified and expressed as -fold relative to the level of MCF7 without dasatinib. (B) MCF7 and MDA-MB-231 cells were treated with (+) or without (−) 50 nM dasatinib for 1 h. The cells were fixed and stained for lamin A, lamin B1, and DNA. Representative images are shown. Scale bars, 10 μm. The nuclear circularity (4π × area/perimeter 2 ) was determined. The P -values were calculated from at least 150 cells pooled from three independent experiments. The percentage of the cells with nuclear lobulation was measured (n ≥ 400). The values (mean ± SD) are from three experiments. *** P < 0.001. (C) MDA-MB-231 cells were infected with lentivirues capable of expressing FLAG-lamin A or the mutants (Y45F and Y45D) and selected in the medium with neomycin (neo). An equal amount of WCLs was analyzed by immunoblotting (IB) with antibodies as indicated. (D) The cells as described in panel C were fixed and stained for FLAG-lamin A, lamin B1, and DNA. Representative images are shown. Scale bars, 10 μm. The percentage of the cells with nuclear lobulation was measured (n ≥ 900). Values (means ± SD) are from three experiments. * P < 0.05, ** P < 0.01, *** P < 0.001. Source data are available for this figure.

Article Snippet: The rabbit polyclonal anti-Src pY416 (MAB2685) antibody was purchased from R&D Systems.

Techniques: Incubation, Control, Western Blot, Phospho-proteomics, Staining, Infection, Expressing

Journal: American Journal of Physiology - Cell Physiology

Article Title: Serum starvation regulates E-cadherin upregulation via activation of c-Src in non-small-cell lung cancer A549 cells

doi: 10.1152/ajpcell.00132.2014

Figure Lengend Snippet: Serum starvation induces c-Src activity. A: c-Src activation was analyzed in A549 cells in serum-free media (0, 6 h, 24 h, 48 h) by immunoblotting for tyrosine 416 phosphorylation of c-Src (PY416-c-Src). Bottom panel: PY416-c-Src intensities were analyzed and normalized to β-actin intensities. *P < 0.05 compared with 0-h starvation data. B: A549 Phospho-c-Src Y416 analysis 24 h after exposure to a range of serum concentrations (0%, 0.5%, 1%, 5%, and 10 %) with PY416-c-Src intensities relative to β-actin intensities. *P < 0.05 compared with 0% FBS. C: immunocytochemistry of A549 cells cultured in media with or without serum for 24 h; IgG (green, top panel), PY416-c-Src (green, bottom panels), and DAPI (red) were visualized by fluorescence under equal exposure conditions. Representative immunoblots and images from 3 independent experiments are shown.

Article Snippet: Phospho-Src Y416 (pY416-Src) antibody was obtained from Cell Signaling Technology (Danvers, MA).

Techniques: Activity Assay, Activation Assay, Western Blot, Phospho-proteomics, Immunocytochemistry, Cell Culture, Fluorescence