Anti rpl4 11302 1 ap

HeLa cells were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM) with 10% fetal bovine serum (FBS). HSF1^+/+ and HSF1^−/− MEFs were kindly provided by I. J. Benjamin (University of Utah). Cycloheximide (CHX), poly-U and puromycin were purchased from Sigma. MitoTracker, Mitochondria-GFP Alexa Fluor 488 or 546 labeled secondary antibodies [donkey anti-mouse IgG (H+L)] and Hoechst were purchased from Invitrogen. Anti-MRPL18 (HPA028774), anti-MRPL38 (HPA023054), anti-RPS20 (HPA003570), anti-RPL5 (HPA043717), anti-myc (C3956) and β-actin (A5441) monoclonal antibody was purchased from Sigma; anti-MRPS18B (16139-1-AP) and anti-RPL4 (11302-1-AP) was from Proteintech; anti-Hsp70 (SPA810) was from Enzo Life Sciences; anti-RPS6 (#2217), anti-HSP60 (#4870), anti-Caspase 3 (#9662)and cleaved Caspase 3 (#9661) antibody were from Cell Signaling. Dual luciferase kit was purchased from Promega. siRNA SMARTpools targeting human MRPL18 and control were purchased from Dharmacon. siRNA targeting human Lyn, MRPL38 and MRPS18B were purchase from Santa Cruz Biotechnology. Plasmids and siRNA transfections were performed using Lipofectamine 2000 (Invitrogen) following the manufacturer’s instruction.

Ogt^F/Y(mER-Cre) and Ogt^F/Y(GFP) MEFs were provided by Natasha E. Zachara (The Johns Hopkins University School of Medicine) and were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM) with 10% FBS (Sigma Aldrich). For amino acid starvation treatment, MEF cells were incubated in HBSS buffer (Lonza) with 10% dialyzed FBS. Samples were collected at indicated time points. 4-Hydroxytamoxifen (H7904) and Thiamet G (SML0244) were purchased from Sigma. Anti-eIF3A (3411), anti-RPS6 (2217), anti-p-eIF2α Ser51 (3398), anti-eIF2α (9722), anti-MMK6/MAP2K6 (9264), anti-eIF3C (2068), and anti-ATF4 (11815) were purchased from Cell Signaling. Anti-O-GlcNAc (RL2, MA1–072) was purchased from Fisher Scientific; anti-Puromycin (PMY-2A4) was purchased from Developmental Studies Hybridoma Bank; β-actin monoclonal antibody (F1804) from Sigma; anti-OGT (11576–2-AP), anti-eIF3g (11165–1-AP) and anti-RPL4 (11302–1-AP) from Proteintech; anti-GFP (NB600–308) from Novus; siRNA targeting human OGT was purchased from Santa Cruz. WGA Agarose beads were purchased from Vector Laboratories (AL-1023–2).