Anti aldolase a

The antibodies against SPAK (#2281; Cell Signalling Technology, Danvers, MA), OSR1 (#3729; Cell Signalling Technology), CFTR (M3A7; Upstate Biotechnology, Waltham, MA), Flag epitope (F3165; Sigma-Aldrich, St. Louis, MO), Myc epitope (sc-40; Santa Cruz Biotechnology, Santa Cruz, CA), anti-Aldolase A (ab78339; Abcam, Cambridge, United Kingdom), and β-actin (sc-1616; Santa Cruz Biotechnology) were obtained from commercial companies. For immunoblot analysis, transfected HEK293T cells were washed 3 times with ice-cold phosphate-buffered saline and then harvested with lysis buffer consisting of 50-mM Tris-HCl (pH 7.4), 150-mM NaCl, 10% (v/v) glycerol, 1% (v/v) Triton X-100, 5-mM EDTA, 5-mM EGTA, and complete protease inhibitor cocktail (Roche Applied Science, Mannheim, Germany). The cells were centrifuged, the cell lysates were diluted in 2× sodium dodecyl sulfate (SDS) sample buffer, and the protein samples were separated by SDS-polyacrylamide gel electrophoresis. The separated proteins were transferred to a nitrocellulose membrane and blotted with the appropriate primary and secondary antibodies. Protein bands were detected by enhanced chemiluminescence (Amersham Biosciences, Piscataway, NJ).

Thapsigargin (Sigma-Aldrich, T9033), STF-083010 (TOCRIS, 4509), APY29 (TOCRIS, 4865), MSC 2032964A (TOCRIS, 5641), VX-809 (Selleckchem, Chemical Abstracts Service registry no. 936727-05-8), VX-770 (Selleckchem, CAS registry no. 873054-44-5), and cycloheximide (Sigma-Aldrich, C4859) were purchased commercially. CSTMP was custom-synthesized by Cayman Chemical (Ann Arbor, MI, USA) (CAS registry no. 1000672-89-8).
The following antibodies were acquired commercially: anti-CFTR M3A7 (Millipore, Billerica, MA, USA), anti-IRE1α (Cell Signaling Technology, 3294), anti–phospho-S724 IRE1α (Abcam, ab48187), anti-XBP1 (Abcam, ab198999), anti-ASK1 (Cell Signaling Technology, 3762), anti–phospho-Thr⁸⁴⁵ ASK1 (Cell Signaling Technology, 3765), anti-BiP (Cell Signaling Technology, 3177), anti-CHOP (Cell Signaling Technology, 2895), anti-pro/p17–caspase 3, anti–cleaved PARP1 (Abcam, ab136812), anti–Na- and K-dependent ATPase (Na,K-ATPase) α1 (Cell Signaling Technology, 23565), anti-HA (Cell Signaling Technology, 2367), anti-Myc (Cell Signaling Technology, 2276), anti–aldolase A (Abcam, ab78339) anti–β-actin (Santa Cruz Biotechnology, sc47778), and anti-pendrin (Santa Cruz Biotechnology, sc23779). The anti-R4 polyclonal antibody was raised against peptides corresponding to amino acids 1458 to 1471 of human CFTR, as described previously (12 (link)).