Western Blot Analysis of ATXN2L, pAKT, and GAPDH

For Western blot, antibodies of ATXN2L (Proteintech, Rosemont, IL), pAKT (Cell Signaling Tech., Danvers, MA, US), and GAPDH (Proteintech) were used. Procedures was as previously described^{21 (link)}. Generally, cells or tissues were solubilized in lysis buffer (KeyGEN, Nanjing, China), and total protein concentration was determined by BioRad BCA method (Pierce, Rockford, IL, USA). After electrophoresed on 10% sodium dodecyl sulfate-polyacrylamide gels, protein was electroblotted onto polyvinylidene fluoride membranes. Then, membranes were blocked with 5% bovine serum albumin and incubated overnight at 4 °C with antibodies against ATXN2L, pAKT, or GAPDH, respectively. After incubated by secondary antibodies, blots were visualized using enhanced chemiluminescence.