P eif2α s51

Manufactured by Cell Signaling Technology

Sourced in United States

P-eIF2α (S51) is an antibody that detects phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α) at serine 51. Phosphorylation of eIF2α is a key regulatory mechanism in the cellular response to various stresses.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

EIF2α (268 citations) P-eIF2α (216 citations)

8 protocols using p eif2α s51

Comprehensive Proteostasis Profiling Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

BTZ (catalog S1013), CFZ (catalog S2853), MEL (catalog S8266), DEX (catalog S1322), and DOX (catalog S1208) were purchased from Selleck Chemicals. Recombinant human PSMB5 protein was custom-made by MedChemExpress. Recombinant human ISG20L2 protein was custom-made by Merry Bio Technology Co., Ltd.
Western blot antibodies against PSMB5 (catalog sc-393931), PSMD8 (catalog sc-514053), PSMD3 (catalog sc-393588), PSMC5 (catalog sc-390631), and eIF2a (catalog sc-133132) were purchased from Santa Cruz Biotechnology. Western blot antibodies against ISG20L2 (catalog 24639-1-AP) and GAPDH (catalog 60004-1-Ig) were purchased from Proteintech. Antibodies against ATF4 (catalog 11815), ATF6 (catalog 65880), cleaved caspase-3 (catalog 9661), p-p38MAPK (T180/Y182) (catalog 9211), p38MAPK (catalog 9212), p-eIF2α (S51) (catalog 3398), and XBP-1s (catalog 27901) were obtained from Cell Signaling Technology. Western blot antibodies against p-PERK (T982) (catalog WL05295), PERK (catalog WL03378), and CHOP (catalog WL00880) were obtained from Wanleibio.

Yang Y., Gao Y., Huang J., Yang Z., Luo H., Wang F., Xu J., Cui Y., Ding H., Lin Z., Zhai X., Qu Y., Zhang L., Liu T., Ye L., Niu T, & Zheng Y. (2022). ISG20L2 suppresses bortezomib antimyeloma activity by attenuating bortezomib binding to PSMB5. JCI Insight, 7(19), e157081.

+ Open protocol

+ Expand

Western Blot Analysis of Cellular Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells were washed and scraped in PBS, centrifuged and the pellet was lysed using RIPA buffer containing protease and phosphatase inhibitor cocktail (Thermo Scientific, Cat. # 78440). 25–30μg of protein was used for SDS PAGE. Tumor tissues were dissociated in RIPA buffer with protease and phosphatase inhibitor cocktail using a pellet pestle followed by sonication for 10 minutes. All primary antibodies were diluted in 5% milk in Tris-buffered Saline/0.1% Tween20 and incubated overnight at 4°C. The following primary antibodies were used: PHGDH (Sigma, Cat. #HPA021241), ATF4 (Cell Signaling Technologies, Cat. #11815S), pERK (T202/204) (Cell Signaling Technologies, Cat. # 9101S), ERK (Cell Signaling Technologies, Cat. # 4695S), p S6 Ribosomal protein (S235/236) (Cell Signaling Technologies, Cat. # 4858S), S6 Ribosomal protein (Cell Signaling Technologies, Cat. # 2217S), p 4EBP1 (S65) (Cell Signaling Technologies, Cat. # 9451S), 4EBP1 (Cell Signaling Technologies, Cat. # 9452S), HA-Tag (Cell Signaling Technologies, Cat. # 3724S), pEIF2α (S51) (Cell Signaling Technologies, Cat. # 3398), EIF2a (Cell Signaling Technologies, Cat. # 5324), GFP (Cell Signaling Technologies, Cat. # 2956S), BRAF (Sigma, Cat. # HPA001328) and β-actin (Invitrogen, Cat. # AM4302)

Jasani N., Xu X., Posorske B., Kim Y., Vera O., Tsai K.Y., DeNicola G.M, & Karreth F.A. (2024). MAPK-mediated PHGDH induction is essential for melanoma formation and represents an actionable vulnerability. bioRxiv.

+ Open protocol

+ Expand

Western Blot Analysis of ER Stress Markers

Check if the same lab product or an alternative is used in the 5 most similar protocols

Whole cell lysates were prepared using the buffers specified by the antibody vendors. Protein concentrations of cleared lysates were measured using a BioRad DC protein assay (BioRad Laboratories, Hercules, CA). Western blot analyses were performed as described (Gao et al. 2008) using commercial antibodies to p‐PERK (T980), PERK, p‐eIF2α(S51), eIF2α, CHOP, cleaved caspase‐3 (Cell Signaling Technology, Beverly, MA), and Nrf2 (Enzo Life Sciences, Farmingdale, NY). Equal protein loading and transfer to blot membranes was verified by Ponceau S Red staining and quantification (Zheng et al. 2010). This method avoids the confounding issues associated with identifying “loading controls” that do not change when the cell treatments being studied alter the synthesis and/or degradation of most cellular proteins.

Woodworth‐Hobbs M.E., Perry B.D., Rahnert J.A., Hudson M.B., Zheng B, & Russ Price S. (2017). Docosahexaenoic acid counteracts palmitate‐induced endoplasmic reticulum stress in C2C12 myotubes: Impact on muscle atrophy. Physiological Reports, 5(23), e13530.

+ Open protocol

+ Expand

Immunoblot Analysis of Cellular Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

Lysates were prepared and electrophoresed on gels as described previously (32 ). Membranes were probed with primary antibodies following each of the supplier’s recommendations: AKT (#4685), pAKT (S473) (#4060), ATG5 (#2630), caspase-3 (#9662), Cleaved PARP (D214) (#9541), p-eIF2α (S51) (#3398), ERK1/2 (#9102), pERK1/2 (T202/Y204) (#9101), GCN2 (#3302), LC3B (#2775), PRAS40 (#2610), pPRAS40 (T246) (#2997), and p62 (#5114) from Cell Signaling Technology (Danvers, MA), and p21 (sc-756), p27 (sc-528), Cyclin D1 (sc-718), α enolase (sc-7455), VDAC1 (sc-8828), and ERK2 (sc-1647) from Santa Cruz Biotechnology (Dallas, TX, USA). The P5CS primary antibody, ALDH18A1 (NBP1-83324), was purchased from Novus Biologicals (Littleton, CO, USA) and pGCN2 (T899) (ab75836) from Abcam. eIF2α and eIF2Bε antibodies were kind gifts from Dr. Scot Kimball. Secondary antibodies goat anti-rabbit IgG-HRP (sc-2004), goat anti-mouse IgG-HRP (sc-2005), and donkey anti-goat IgG-HRP (sc-2020) were purchased from Santa Cruz Biotechnology. The immunoblots were developed using ECL Western Blotting Substrate (Thermo Scientific) or Supersignal West Femto Chemiluminescent Substrate (Thermo Scientific).

Kardos G.R., Wastyk H.C, & Robertson G.P. (2015). Disruption of Proline Synthesis in Melanoma Inhibits Protein Production Mediated by the GCN2 Pathway. Molecular cancer research : MCR, 13(10), 1408-1420.

+ Open protocol

+ Expand

Melanoma Protein Signaling Profiling

Check if the same lab product or an alternative is used in the 5 most similar protocols

The melanoma tumor tissues and cultured cells were harvested in Tween 20 buffer containing 50mM HEPES (pH 8.0), 150mM NaCl, 2.5mM EGTA, 1mM EDTA, 0.1% Tween 20, and protease/phosphatase inhibitors (1mM phenylmethylsulphonyl fluoride, 20 U of aprotinin/ml, 5mg of leupeptin/ml, 1mM DTT, 0.4mM NaF, and 10mM β-glycerophosphate). Lysates were sonicated prior to clearing by centrifugation at 4°C for 30 min. Proteins were resolved by SDS-PAGE, transferred to membrane, and subjected to immunoblot. Antibodies utilized include PERK (Rockland), p-eIF2α S51 (Cell Signaling), BiP (Cell Signaling), total eIF2α (Cell Signaling), Cyclin D1 (mouse monoclonal D1-72-13G), Cul4a (Bethyl, A300-739A), p-AktS473 (Cell Signaling), total Akt (Cell Signaling), GAPDH (Cell Signaling) and β-actin (Sigma Aldrich).

Pytel D., Gao Y., Mackiewicz K., Katlinskaya Y.V., Staschke K.A., Paredes M.C., Yoshida A., Qie S., Zhang G., Chajewski O.S., Wu L., Majsterek I., Herlyn M., Fuchs S.Y, & Diehl J.A. (2016). PERK Is a Haploinsufficient Tumor Suppressor: Gene Dose Determines Tumor-Suppressive Versus Tumor Promoting Properties of PERK in Melanoma. PLoS Genetics, 12(12), e1006518.

+ Open protocol

+ Expand

Investigating Translational Regulation Mechanisms

Check if the same lab product or an alternative is used in the 5 most similar protocols

eFT508 was acquired from eFFECTOR Therapeutics. BBN and BCPN were purchased from TCI Chemicals. Antibodies used were against phospho-eIF4E S209 (Abcam, ab76256), eIF4E (Santa Cruz Biotechnology Inc., sc-271480), rpS6 (Cell Signaling Technology, 2217), p-rpS6 (S240/244) (Cell Signaling Technology, 5364), p-rpS6 (S235/236) (Cell Signaling Technology, 4858), 4EBP1 (Cell Signaling Technology, 9644), p-4EBP1 (T37/46) (Cell Signaling Technology, 2855), eIF2α (Cell Signaling Technology, 2103), p-eIF2α (S51) (Cell Signaling Technology, 3597), eEF2 (Cell Signaling Technology, 2332), p-eEF2 (T56) (Cell Signaling Technology, 2331), puromycin (EMD Millipore, MABE343), and tubulin (MilliporeSigma, T8203).

Jana S., Deo R., Hough R.P., Liu Y., Horn J.L., Wright J.L., Lam H.M., Webster K.R., Chiang G.G., Sonenberg N, & Hsieh A.C. (2021). mRNA translation is a therapeutic vulnerability necessary for bladder epithelial transformation. JCI Insight, 6(11), e144920.

+ Open protocol

+ Expand

Protein Expression Analysis in Cultured Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total cell lysates and nuclear protein-enriched lysates from cultured cells or ectopic bone implants were obtained using the appropriate cell lysis buffers, and Western blot analysis was performed as described previously.^{10 ,12 (link)} Briefly, proteins were separated by SDS-PAGE, and protein-containing nitrocellulose membranes were incubated overnight with primary antibodies specific for the following proteins: AMPK (#2532; Cell Signaling Technology), p-AMPK^T172 (#2535, Cell Signaling Technology), ATF4 (#11815, Cell Signaling Technology), β-actin (A5441, Sigma-Aldrich), eIF2α (#9722, Cell Signaling Technology), p-eIF2α^S51 (#9721, Cell Signaling Technology), Lamin A/C (sc-376248, Santa Cruz Biotechnologies), PHGDH (#66350, Cell Signaling Technology) and PSAT1 (NBP1-55368, Bio-Techne). Appropriate HRP-conjugated secondary antibodies were used for chemiluminescent detection of proteins (Western Lightning Plus, PerkinElmer).

Stegen S., Loopmans S., Stockmans I., Moermans K., Carmeliet P, & Carmeliet G. (2022). De novo serine synthesis regulates chondrocyte proliferation during bone development and repair. Bone Research, 10, 14.

+ Open protocol

+ Expand

Inhibitors of Antiviral Signaling Pathways

Check if the same lab product or an alternative is used in the 5 most similar protocols

PKR inhibitor C16 (Tocris), IKKα:β inhibitor IKK16, TBK1 inhibitor Bx795 and JAK/STAT inhibitor Ruxolitinib (all Selleck Chemicals), and ISR inhibitor ISRIB (Sigma) were dissolved in DMSO and used at the indicated concentrations. Biotinylated HMW poly(I·C) was purchased from InvivoGen and recombinant IFNα2 was from PBL Assay Science. Primary antibodies used in this study were against eIF4G1, eIF4A, GAPDH, IRF3, p-IRF3(S396), STAT1, p-STAT1(Y701), IFNβ, IFIT1, PCBP2, PKR, ADAR, Dicer, IFI16, PACT, MDA5, TBK1, p-TBK1(S172), eIF2a, p-eIF2α(S51) (all Cell Signaling Technology), DHX9, LGP2, IFIT5 (all Proteintech), HelZ2, MCCC1 (ThermoFisher), DHX30 (Novus), NF90 (BD Biosciences), α-tubulin (Sigma-Aldrich), and p-PKR(T446) (LSBio). Anti-2C antibody was a gift from E. Wimmer (Stony Brook Univ., NY). Immunoblots were performed as described previously (59 (link), 60 (link)).

Dobrikov M.I., Dobrikova E.Y., McKay Z.P., Kastan J.P., Brown M.C, & Gromeier M. (2022). PKR Binds Enterovirus IRESs, Displaces Host Translation Factors, and Impairs Viral Translation to Enable Innate Antiviral Signaling. mBio, 13(3), e00854-22.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!