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P cjuns73

Manufactured by Cell Signaling Technology
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P-cJunS73 is an antibody product developed by Cell Signaling Technology. It is designed to detect the phosphorylation of the c-Jun protein at the serine 73 residue. This phosphorylation event is a known regulatory mechanism for the activity of the c-Jun transcription factor.

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Lab products found in correlation

C jun, by Cell Signaling Technology (4 mentions) β actin, by Merck Group (2 mentions) γh2ax, by Merck Group (2 mentions) Goat anti rabbit igg alexa 488 conjugate, by Thermo Fisher Scientific (2 mentions) Fluorophore conjugated antibodies, by Cell Signaling Technology (2 mentions) P rbs807 s811, by Cell Signaling Technology (2 mentions) Erk1 2, by Cell Signaling Technology (1 mentions) Anti e cadherin, by Proteintech (1 mentions) P erk1 2 t202 y204, by Cell Signaling Technology (1 mentions) Anti n cadherin antibody, by Cell Signaling Technology (1 mentions) P jnk t183 y185, by Cell Signaling Technology (1 mentions) P p38 t180 y182, by Cell Signaling Technology (1 mentions) Twist1, by Cell Signaling Technology (1 mentions) Snail, by Cell Signaling Technology (1 mentions) Anti vimentin, by Cell Signaling Technology (1 mentions) Dc protein assay kit, by Bio-Rad (1 mentions) Pvdf membrane, by Bio-Rad (1 mentions) Pnfκb p65 s536, by Cell Signaling Technology (1 mentions) Cox 2, by Cayman Chemical (1 mentions) Ecf western blotting system, by Cytiva (1 mentions)

8 protocols using p cjuns73

1

Western Blot Analysis of EMT Markers

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The anti-NUAK1 antibody (22723-1-AP) and anti-β-actin antibody (23660-1-AP) were purchased from ProteinTech (Chicago, IL, USA); the anti-E-cadherin (#14472), anti-Vimentin (#46173), anti-N-cadherin antibody (#13116), TWIST1 (#90445), ZEB1 (#83243), Snail (#3879), Slug (#9585), p-c-Jun (S73) (#3270), c-Jun (#9165), p-JNK (T183/Y185) (#4668), JNK (#9252), p-Erk1/2 (T202/Y204) (#4370), Erk1/2 (#4695), p-p38 (T180/Y182) (#54470) and p38 (#9212) were purchased from Cell Signaling Technology (CST, Danvers, MA, USA). SP600125 (HY-12,041), JNK-IN-8 (HY-13,319) and puromycin dihydrochloride (HY-B1743A) were purchased from MCE (NJ, USA).
Yang H., Wei Z., Song Y., Du K., Yin N., Lu H., Li B., Hou L., Xing P., Chen L., Wang C, & Xie S. (2023). NUAK1 promotes tumor metastasis through upregulating slug transcription in esophageal squamous cell carcinoma. Cancer Cell International, 23, 258.
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2

Western Blot Analysis of Signaling Proteins

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The cells were washed twice with ice-cold PBS and collected with the cell lysis buffer (10 mM Tris-HCl, pH 7.4, 1% SDS, and 1 mM Na3VO4). The cell extracts were sonicated, denatured by heating at 100 °C for 5 min, and quantified with a Dc protein assay kit (Bio-Rad Hercules, CA). Equal aliquots of cell extracts were separated on SDS-polyacrylamide gels. The proteins were then transferred to PVDF membranes (Bio-Rad Hercules, CA), blocked, and probed with one of the antibodies against COX-2 (Cayman Chemical Co.), p-c-Jun S73, p-IKKα/β S176/180, p-IκBα S32/36, p-NFκB p65 S536, total c-Jun IκBα and IKKα/β (Cell Signaling Technology, Beverly, MA), NFAT3, p65, and p50 (Santa Cruz Biotechnology, CA, USA), or β-Actin (Sigma). Primary antibody-bound proteins were detected by using an alkaline phosphatase-linked secondary antibody and an ECF Western blotting system (Amersham, Piscataway, NJ).
Wei J., Du K., Cai Q., Ma L., Jiao Z., Tan J., Xu Z., Li J., Luo W., Chen J., Gao J., Zhang D, & Huang C. (2014). Lead induces COX-2 expression in glial cells in a NFAT-dependent, AP-1/NFκB-independent manner. Toxicology, 325, 67-73.
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3

Comprehensive Antibody Characterization for CycIF

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The specific animal sources, catalogue numbers and dilutions for fluorophore-conjugated antibodies from Cell Signaling Technology are listed in Supplementary Table 1. The source and dilution of un-conjugated antibodies used in this study are as follows: p-RbS807/S811 (rabbit mAb; Cell Signaling:#8516 1:1000); p-cJunS73 (rabbit mAb; Cell Signaling:#3270; 1:500); c-Jun (rabbit mAb, Cell Signaling:#9165; 1:500); p53 (mouse mAb, Santa Cruz Biotech. sc-126, 1:200); γH2A.X (mouse mAb, Millipore: 05-636, 1:1000); Goat-anti-Rabbit IgG, Alexa-488 conjugate (Invitrogen:A-11034, 1:2000); Goat-anti-Mouse IgG, Alexa 647 conjugate (Invitrogen:A-21235, 1:2000). In Supplementary table 3, we listed all the antibodies we have already tested in CycIF protocol, including antibodies from several different vendors (Cell Signaling Technology, Abcam, Santa Cruz Technology & Biolegend). More updates will be available in the HMS-LINCS website (http://lincs.hms.harvard.edu/lin-NatCommun-2015/).
Lin J.R., Fallahi-Sichani M, & Sorger P.K. (2015). Highly multiplexed imaging of single cells using a high-throughput cyclic immunofluorescence method. Nature Communications, 6, 8390.
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4

Protein Expression Analysis in Cells

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The protein levels of FAK, p63 (4A4), MMP1, MMP2, BrdU (Santacruz Biotech), Src, pFAK-Y395, MMP14 (Abcam), paxillin (BD biosciences), ppaxillin-Y118, pFAK-Y576/577, pSrc-Y496, pAKT-S493, total AKT, p-c-Jun-S73, c-Jun (Cell Signaling) and β-actin (Sigma-aldrich) were detected in whole cell lysates by immunoblotting as previously described [50 (link)].
Srivastava K., Pickard A., McDade S, & McCance D.J. (2015). p63 drives invasion in keratinocytes expressing HPV16 E6/E7 genes through regulation of Src-FAK signalling. Oncotarget, 8(10), 16202-16219.
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5

Multicolor Immunofluorescence Staining Protocol

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The specific animal sources, catalogue numbers and dilutions for fluorophore-conjugated antibodies from Cell Signaling Technology are listed in Supplementary Table 1. The source and dilution of unconjugated antibodies used in this study are as follows: p-RbS807/S811 (rabbit mAb; Cell Signaling:#8516 1:1,000); p-cJunS73 (rabbit mAb; Cell Signaling:#3270; 1:500 ); c-Jun (rabbit mAb, Cell Signaling:#9165; 1:500); p53 (mouse mAb, Santa Cruz Biotech. sc-126, 1:200); γH2A.X (mouse mAb, Millipore: 05-636, 1:1,000); Goat-anti-Rabbit IgG, Alexa-488 conjugate (Invitrogen:A-11034, 1:2,000); Goat-anti-Mouse IgG, Alexa 647 conjugate (Invitrogen:A-21235, 1:2,000). In Supplementary Table 3, we listed all the antibodies we have already tested in CycIF protocol, including antibodies from several different vendors (Cell Signaling Technology, Abcam, Santa Cruz Technology and Biolegend). More updates will be available in the HMS-LINCS website (http://lincs.hms.harvard.edu/lin-NatCommun-2015/).
Lin J.R., Fallahi-Sichani M, & Sorger P.K. (2015). Highly multiplexed imaging of single cells using a high-throughput cyclic immunofluorescence method. Nature Communications, 6, 8390.
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6

Comprehensive Antibody Characterization and Validation

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All antibodies were purchased from commercial sources as indicated below: R&D Systems (Minneapolis, MN, USA): AXL (for IB) and pAXL-Y779. Cell Signaling Technology (Danvers, MA, USA): pAXL-Y702, pEGFR-Y1068, pMAPK (T202/Y204), MAPK, p-cRAF (S289/296/301), cRAF, p-AKT (S473), AKT, p-rpS6 (S240/244), rpS6, p-c-Jun (S73), c-Jun and GAPDH. Santa Cruz Biotechnology Inc. (Dallas, TX, USA): pEGFR-Y1173, AXL (for IP), and HRP-conjugated goat-anti-rabbit IgG, goat-anti-mouse IgG, donkey-anti-goat IgG. Life Technologies (Carlsbad, CA, USA): AXL (For IF). Abcam (Cambridge, MA, USA): EGFR. Calbiochem (Billerica, MA, USA): α-Tubulin.
Brand T.M., Iida M., Stein A.P., Corrigan K.L., Braverman C., Luthar N., Toulany M., Gill P.S., Salgia R., Kimple R.J, & Wheeler D.L. (2014). AXL mediates resistance to cetuximab therapy. Cancer research, 74(18), 5152-5164.
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7

Immunoblotting Analysis of Melanoma Signaling

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Whole-cell protein lysates (20 μg each) of primary melanocytes and various melanoma cell lines were separated by electrophoresis on 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis gel, transferred to nitrocellulose membrane and analyzed by immunoblotting with antibodies against MALT1 (#2494), pc-Jun(S73) (#9164), pIκBα(Ser32) (#28590), β1-Integrin (#9699) (Cell signaling Technology, Danvers, MA, USA); c-Jun (Ab-243) (GenScript); RelA/p65 (SC109), IκBα (SC847), MKK7 (SC13071) and Actin (SC16160) (Santa Cruz Biotechnology, Santa CruZ, CA, USA) followed by detection with Alexa IRDye-conjugated secondary antibodies (Invitrogen). Blots were scanned using the Odyssey CLX imaging system (LI-COR, Lincoln, NE, USA).
Wang Y., Zhang G., Jin J., Degan S., Tameze Y, & Zhang J.Y. (2017). MALT1 promotes melanoma progression through JNK/c-Jun signaling. Oncogenesis, 6(7), e365-.
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8

Protein Kinase Signaling Pathway Analysis

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Cells were lysed on ice after 24 hours using Triton X-100 Cell Lysis Buffer (Cell Signaling) supplemented with a protease inhibitor tablet (Roche). Lysates were either resolved on SDS–polyacrylamide gel electrophoresis (PAGE) gels followed by Western blotting or used in an in vitro kinase assay (details below). Primary antibodies used were as follows: Flag M2 and α-tubulin (Sigma-Aldrich); MAP2K4, MAP2K7, pJNK (T183/Y185), pMARCKS (S152/S156), pPKC (S676), pThr, and p-cJun (S73) (Cell Signaling Technology). Mouse or rabbit horseradish peroxidase–conjugated secondary antibodies were used (Cell Signaling Technology). All Western blots are representative of at least three independent experiments.
Hudson A.M., Stephenson N.L., Li C., Trotter E., Fletcher A.J., Katona G., Bieniasz-Krzywiec P., Howell M., Wirth C., Furney S., Miller C.J, & Brognard J. (2018). Truncation- and motif-based pan-cancer analysis reveals tumor-suppressing kinases. Science signaling, 11(526), eaan6776.
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