Journal: The Journal of Biological Chemistry

Article Title: PtpA, a secreted tyrosine phosphatase from Staphylococcus aureus , contributes to virulence and interacts with coronin-1A during infection

doi: 10.1074/jbc.RA118.003555

Figure Lengend Snippet: PtpA interacts with Coro-1A in vitro. A, the interaction between PtpA and D. discoideum CorA was confirmed by GST pulldown assay. The indicated GST fusion proteins were expressed in E. coli, bound to GSH beads, and then incubated with D. discoideum cell lysates expressing myc-tagged-Dd_Coro-1 (Dd_Coro-A_myc). Beads were washed, eluted by boiling, and bound proteins were revealed by Western blot analysis with an anti-myc antibody. B, the interaction between PtpA and human Coro-1A was tested by pulldown analysis. His-tagged versions of PtpA (WT) and the catalytically inactive PtpA_C8S (C8S) were constructed, produced. and purified as described under “Experimental procedures.” BL21 lysates expressing Hs_Coro-1A_GST or GST alone were prepared and incubated with the PtpA_His derivatives immobilized on Ni-NTA-agarose beads. The bound proteins were eluted and resolved by SDS-PAGE followed by immunoblotting using anti-GST and anti-Coro-1A antibodies. Empty Ni-NTA-agarose beads were used as a control. *, nonspecific signals co-precipitated with PtpA_C8S_His. M kDa, molecular markers.

Article Snippet: Immunoprecipitation of Coro-1A from infected macrophages Murine macrophage coronin-1A (Mm_Coro-1A) was obtained from S. aureus- infected RAW 264.7 macrophage extracts by immunoprecipitation with anti-Coro-1A antibody (5 μg, rabbit anti-coronin-1A; SAB4200078, Sigma).

Techniques: In Vitro, GST Pulldown Assay, Incubation, Expressing, Western Blot, Construct, Produced, Purification, SDS Page

Journal: The Journal of Biological Chemistry

Article Title: PtpA, a secreted tyrosine phosphatase from Staphylococcus aureus , contributes to virulence and interacts with coronin-1A during infection

doi: 10.1074/jbc.RA118.003555

Figure Lengend Snippet: Coro-1A is phosphorylated on tyrosines in vivo but is not dephosphorylated by PtpA in vitro. A, immunoblot analysis with anti-Coro-1A (upper panel) or anti-phosphotyrosine antibodies (bottom panel) of immunoprecipitated (IP) endogenous Coro-1A (Mm_Coro-1A) from lysates of RAW 264.7 macrophages infected with cells of S. aureus Newman (WT) or the isogenic ΔptpA mutant. Noninfected macrophages and infected macrophages were treated with gentamicin for 30 min and subsequently incubated with lysostaphin to kill extracellular bacteria that might be released from lysed macrophages during the successive incubation time. Noninfected and infected macrophages were lysed 30 min (NI T30 min and T30 min) and 3 h (NI T3 h and T3 h) pGt treatment. B, immunoprecipitated Mm_Coro-1A from lysates of macrophages infected with strain Newman for 30 min pGt were incubated in the absence (−) or presence (+) of 2 μg of PtpA_His or PtpA-D120A_His at 37 °C for the time points indicated (T0, 30 and 60 min). Proteins were resolved by SDS-PAGE and probed with anti-Coro-1A (upper panel) or anti-phosphotyrosine (bottom panel) antibodies on the same blot. Contents of Mm_Coro-1A and Tyr-phosphorylated Mm_Coro-1A in lysates of S. aureus-infected macrophages prior to concentration by IP are indicated in the input lane (input). M kDa, molecular markers.

Article Snippet: Immunoprecipitation of Coro-1A from infected macrophages Murine macrophage coronin-1A (Mm_Coro-1A) was obtained from S. aureus- infected RAW 264.7 macrophage extracts by immunoprecipitation with anti-Coro-1A antibody (5 μg, rabbit anti-coronin-1A; SAB4200078, Sigma).

Techniques: In Vivo, In Vitro, Western Blot, Immunoprecipitation, Infection, Mutagenesis, Incubation, SDS Page, Concentration Assay

Journal: The Journal of Biological Chemistry

Article Title: PtpA, a secreted tyrosine phosphatase from Staphylococcus aureus , contributes to virulence and interacts with coronin-1A during infection

doi: 10.1074/jbc.RA118.003555

Figure Lengend Snippet: Strains, plasmids, and primers used in this study

Article Snippet: Immunoprecipitation of Coro-1A from infected macrophages Murine macrophage coronin-1A (Mm_Coro-1A) was obtained from S. aureus- infected RAW 264.7 macrophage extracts by immunoprecipitation with anti-Coro-1A antibody (5 μg, rabbit anti-coronin-1A; SAB4200078, Sigma).

Techniques: Plasmid Preparation, Sequencing, FLAG-tag, Modification, Expressing

Journal: The Journal of Physiology

Article Title: Dynamics of neuroeffector coupling at cardiac sympathetic synapses

doi: 10.1113/JP275693

Figure Lengend Snippet: Primary antibodies used in the present study

Article Snippet: Rabbit anti‐synapsin 1a , Millipore , 1:1000 , IF.

Techniques:

Journal: eLife

Article Title: A pre-screening strategy to assess resected tumor margins by imaging cytoplasmic viscosity and hypoxia

doi: 10.7554/eLife.70471

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-HIF-1A antibody (Rabbit polyclonal) , Sangon Biotech , Cat#D162108RRID: AB_1674786 , WB (1:500).

Techniques: Recombinant, Concentration Assay, Bicinchoninic Acid Protein Assay, CCK-8 Assay, Gel Extraction, Software

Journal: Cell reports

Article Title: HIF-1α accumulation in response to transient hypoglycemia may worsen diabetic eye disease

doi: 10.1016/j.celrep.2022.111976

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-HIF-1a , Gene Tex , Cat# GTX127309; RRID:AB_2616089.

Techniques: Virus, Recombinant, Tube Formation Assay, In Vivo, Angiogenesis Assay, Enzyme-linked Immunosorbent Assay, Lactate Assay, Plasmid Preparation, Software