Gapdh sc 47724

Western blot and immunoprecipitation analysis were performed as described previously [13 (link)]. The antibodies for GAPDH (sc-47724; 1 : 1000), p62 (sc-101542; 1 : 1000), anti-LC3 (sc-398822; 1 : 1000), and individual secondary antibodies were purchased from Santa Cruz (Santa Cruz Biotechnology, Santa Cruz, CA). The antibodies for ATG5 (#12994; 1 : 1000), ATG13 (#13468; 1 : 1000), ULK1 (#8054; 1 : 1000), and NEDD4L (#4013; 1 : 1000) were all obtained from Cell Signaling Technology (Cell Signaling Technology Inc., Beverly, USA).

C3H10T1/2 cells were bought from ATCC (Manassas, VA, USA) and cultured with complete Dulbecco's modified Eagle's medium, which containing 10% fetal bovine serum (FBS), 100 μg/ml streptomycin, and 100 U/ml penicillin. Besides this, cells were saturated at 37°C and 5% CO₂. Primary antibodies against PTEN (sc-7974), Runx2 (sc-390715), OPN (sc-21742), and GAPDH (sc-47724) were ordered from Santa Cruz Biotechnology; Wnt10b (ab70816), CREB (ab32515), and p-CREB (ab32096) were ordered from Abcam; Smad1/5/9 (13820S) was ordered from CST; and p-Smad1/5/9 (AF8313) was bought from Affinity Biotech.

Cells were lysed, and total protein was extracted. The proteins were separated with sodium dodecyl sulfate polyacrylamide gel electrophoresis and were then transferred to a PVDF membrane. After blocking, the membranes were incubated with primary antibodies (GAPDH (sc‐47 724) from Santacruz, Delaware Ave, USA; PTEN (9559L), FGF2 (3196S), FGFR (3471S), FRS2 (3864L) from cell signaling, Boston, USA) at 4°C overnight and then incubated with secondary antibody. Protein bands were analyzed using the Chemi Doc XRS System (Bio‐Rad, Hercules, CA, USA).

A protease inhibitor cocktail (P2741), a phosphatase inhibitor cocktail (P5726), pan caspase inhibitor Z-VAD-FMK (V116), Ca²⁺ chelator BAPTA-AM (A1076) and monoclonal antibody against FLAG epitope (SAB420007) were procured from Merck (Sigma-Aldrich, St. Louis, Missouri, USA). Both mouse monoclonal antibodies (ab107359) and rabbit polyclonal antibodies (ab73864) against viperin were purchased from Abcam, USA. Mouse monoclonal antibodies against COX-4 (sc-376731) and GAPDH (sc-47724) and rabbit polyclonal antibodies against calnexin (sc-11397) were purchased from Santa Cruz Biotechnology, USA. Rabbit polyclonal antibodies against cleaved caspase-3 (9661s) and cleaved caspase-9 (9501) were procured from Cell Signaling Technology, USA. Monoclonal antibodies against rotaviral structural protein VP6 (3R103C10) were obtained from HyTest, Finland. Monoclonal antibodies against cytochrome c (CH2.B4) were acquired from BD Biosciences. Antisera against rotaviral non-structural protein NSP4 and VP7 were raised in rabbits, according to standard protocols at the Department of Virology and Parasitology, Fujita Health University School of Medicine, Aichi, Japan.

Immunoblotting was performed as previously described^{55 (link)} with the following antibodies: mouse monoclonal to pSMAD2 3104S (Cell Signaling, MA, USA) and mouse monoclonal GAPDH sc-47724 (Santa Cruz, TX, USA). Protein bands were visualized by chemiluminescence detection by BIO-RAD ChemiDoc MP Imaging System (Thermo Scientific, MA, USA).