Ptpn22

A high-affinity HSP90-inhibitor, AUY922 [30 (link)] was purchased from Selleckchem. Antibodies to HSP90_αβ, PLCγ2, BCAP, CD19, AXL, BCL2, GAPDH and actin were purchased from Santa Cruz Biotechnologies. Antibodies to CD79a, CD79b, LYN, SYK, BTK, AKT, P-ERK1/2, ERK1/2, STAT3, PTPN22, FGFR3, and MCL-1 were purchased from Cell Signaling Technologies. XIAP antibody, chromogen-conjugated antibodies to human CD5 and CD19 or fluorescein isothiocyanate (FITC)-conjugated annexin V were obtained from BD Biosciences or Invitrogen, respectively. Propidium iodide (PI) and other chemicals were purchased from Sigma or Bio-Rad. Replication-deficient lentiviral constructs expressing HSP90-specific shRNA or GFP tagged control scrambled shRNA were purchased from Santa Cruz Biotechnologies.

The antibodies used in this study are as follows: HDAC10 (Sigma, H3413), ERK1/2 (Cell Signaling Technology, #4695), p-ERK1/2 (Thr202/Tyr204) (Cell Signaling Technology, #4370), PTPN22 (Cell Signaling Technology, #14693), and GAPDH (Santa Cruz Biotechnology, sc-47724). For Western blot, HUVECs were lysed with radio-immunoprecipitation assay buffer (50 mM Tris, 0.1% sodium dodecyl sulfate, 1.0% IGEPAL CA-630, 150 mM NaCl, and 0.5% sodium deoxycholate, pH 8.0, The Shanghai Sheng Gong Company) on ice for 25 min. The lysates were centrifuged at 12,000 rpm for 15 minutes at 4°C, and the protein concentrations in the samples were determined using the bicinchoninic (BCA) method. Equal amounts of protein were loaded onto SDS-PAGE gels, separated by electrophoresis, transferred to membranes and incubated with the appropriate antibodies. Then, the signals were developed using the Clarity Western enhanced chemiluminescence (ECL) substrate (1705061, Bio-Rad).