The antibodies used were: ASCL1 (Abcam, #ab211327), INSM1 (Abcam, #ab170876), E2F7 (Abcam, ab245655), RB1 (Abcam, ab181616), SYP (Santa Cruz, #sc-17750), RCOR1 (Santa Cruz, #sc-376567), CCN1 (Santa Cruz, #sc-374129), CCN2 (Santa Cruz, #sc-365970), YAP/TAZ (Santa Cruz, #sc-101199), P53 (Santa Cruz, #sc-126), E2F1 (Santa Cruz, #sc-251), GAPDH (Santa Cruz, #sc-47724), LATS1 (Cell signaling, #3477), LATS2 (Cell signaling, #5888), p-LATS (Cell signaling, #8654), H3 (Cell signaling, #4499), H3K27ac (Cell signaling, #8173), H3K4me3 (Cell signaling, #9751), VIN (Cell signaling, #13901), Flag (Sigma, #1804),
Yap taz
Manufactured by Santa Cruz Biotechnology
Sourced in United States
YAP/TAZ is a protein complex that plays a central role in the Hippo signaling pathway, which regulates cell growth, proliferation, and apoptosis. The YAP/TAZ complex acts as a transcriptional co-activator, transducing signals from the Hippo pathway to influence gene expression. This product is a research tool used to study the Hippo signaling pathway and its implications in cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Lats1, by Cell Signaling Technology (3 mentions)
Gapdh, by Santa Cruz Biotechnology (3 mentions)
Lats2, by Cell Signaling Technology (2 mentions)
Alexa fluor 488 phalloidin, by Thermo Fisher Scientific (2 mentions)
Jmjd1a 12835 1 ap, by Proteintech (2 mentions)
P lat, by Cell Signaling Technology (1 mentions)
Flag tag (flag), by Merck Group (1 mentions)
H3k27ac, by Cell Signaling Technology (1 mentions)
Ascl1, by Abcam (1 mentions)
Insm1, by Abcam (1 mentions)
Rcor1, by Santa Cruz Biotechnology (1 mentions)
H3k4me3, by Cell Signaling Technology (1 mentions)
Protease inhibitor, by Roche (1 mentions)
Phosphatase inhibitor, by Thermo Fisher Scientific (1 mentions)
P yap, by Cell Signaling Technology (1 mentions)
Flag tag (flag), by Cell Signaling Technology (1 mentions)
Anti flag hrp, by Merck Group (1 mentions)
General western blot protocol, by Abcam (1 mentions)
Ncor2, by Cell Signaling Technology (1 mentions)
Myc tag, by Cell Signaling Technology (1 mentions)
8 protocols using yap taz
1
Comprehensive Immunohistochemistry Profiling
2
Immunoblot and Immunoprecipitation Protocol
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Immunoblot was performed per a general western-blot protocol (Abcam). Antibodies for Flag-tag (#14793, 1:1000), HA-tag (#2367, 1:1000), Myc-tag (#2278, 1:1000), YAP (#14074, 1:1000), pYAP(S127, 1:2000) (#4911), LATS1 (#3477, 1:1000), LATS2 (#5888, 1:1000), pLATS1/2 (HM) (#8654, 1:1000), NCOR2 (#62370, 1:1000) and ERα (human) (#8644, 1:2000) were from Cell Signaling Technology. The GAPDH (#sc-25778, 1:3000), YAP/TAZ (#sc-101199, 1:2000) and ERα (mouse) (#sc-542, 1:1000) were obtained from Santa Cruz Biotechnology. The anti-Flag (HRP) (#A5892, 1:3000) was from Sigma.
For immunoprecipitations, cells were rinsed with ice-cold DPBS and then lysed in mild lysis buffer (150 mM NaCl, 50 mM Tris-Cl pH7.5, 0.5% Triton X-100) with protease inhibitors (Roche, #11873580001) and phosphatase inhibitors (Thermo Scientific, #88667) on ice for 30 min and centrifuged at 12,000 rpm for 10 min. Primary antibodies and Protein A/G magnetic beads (Pierce, #88803) were added to the supernatants and incubated with rotation for 3 h at 4 °C. Immunoprecipitants were washed four times with lysis buffer and were denatured by SDS-PAGE sample buffer and boiled for 5 min. Sample were followed by immunoblot analysis with antibodies indicated in the figures.
For immunoprecipitations, cells were rinsed with ice-cold DPBS and then lysed in mild lysis buffer (150 mM NaCl, 50 mM Tris-Cl pH7.5, 0.5% Triton X-100) with protease inhibitors (Roche, #11873580001) and phosphatase inhibitors (Thermo Scientific, #88667) on ice for 30 min and centrifuged at 12,000 rpm for 10 min. Primary antibodies and Protein A/G magnetic beads (Pierce, #88803) were added to the supernatants and incubated with rotation for 3 h at 4 °C. Immunoprecipitants were washed four times with lysis buffer and were denatured by SDS-PAGE sample buffer and boiled for 5 min. Sample were followed by immunoblot analysis with antibodies indicated in the figures.
3
Immunohistochemistry of Fibroblast Markers
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Immunohistology was performed as described previously [12 (link)]. Briefly, fibroblasts were fixed in 2% paraformaldehyde for one hour at room temperature and were incubated with 0.5% Nonidet P-40 and then blocked with 2% bovine serum albumin (BSA). The cells were washed with PBS three times and incubated with primary antibodies (YAP/TAZ and Smad7, Santa Cruz Biotechnology, Santa Cruz, CA; Phospho Smad3, Abcam, Cambridge, MA) for 1 h at room temperature, followed by incubation with Super Sensitive MultiLink (BioGenex, Fremont CA) for 10 min and streptavidin-conjugated AlexaFluor 594 or 488 (Invitrogen-Molecular Probes, San Diego, CA) for 10 min. Mounting medium with DAPI was added to stain cell nuclei. Corresponding IgG isotype (negative control) show no specific staining (data not shown). For phalloidin (Sigma, St. Louis, MO, USA) staining, fibroblasts were washed with PBS and were fixed in 2% paraformaldehyde for 30 min followed by phalloidin staining for one hour.
4
Immunoblotting and Microscopy Protocols
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The following antibodies were purchased from Cell Signaling and used at the indicated dilution for Western blot analysis: Merlin (NF2) (1:1000; 6995), phospho-Merlin Ser518 (1:1000; 9163), YAP (1:1000; 14074), pYAP-Ser127 (1:1000; 4911), LATS1 (1:1000; 3477), phospho-LATS HM (1:1000; 8654), phospho-MK2 Thr334 (1:1000; 3007), phospho-p38 Thr180/Tyr182 (1:1000; 4511), and HA HRP-conjugated (1:10,000; 2999). GAPDH (1:2000) and YAP/TAZ (1:500) were obtained from Santa Cruz Biotechnology (sc-25778 and sc-101199, respectively). Vinculin (1:5000) and Flag HRP conjugated (1:10,000) were from Sigma (V9131 and A8592, respectively. N-Cadherin (1:1000) was from BD Biosciences (610920).
The following antibodies were used for immunofluorescent microscopy experiments at the indicated dilutions: GFP (1:200) was from Abcam (ab6673), PI(4,5)P2 (1:200) was from Echelon Biosciences (Z-G045), Flag (1:500) and HA (1:500) were from Cell Signaling (14793 and 3724, respectively), and YAP (1:200) was from Santa Cruz Biotechnology (sc-101199).
Secondary antibodies Alexa fluor 488 and 555 and phalloidin were from Invitrogen and used in 1:1000 dilution.
Chemicals sorbitol was from Fisher Scientific (S459-500), latrunculin B (Lat B) was from Abcam (ab144291), 2-deoxy-D-glucose (2-DG) was from Sigma (D8375), and p38 inhibitor SB203580 (1202) and JNK inhibitor SP600125 (1496) were from Tocris.
The following antibodies were used for immunofluorescent microscopy experiments at the indicated dilutions: GFP (1:200) was from Abcam (ab6673), PI(4,5)P2 (1:200) was from Echelon Biosciences (Z-G045), Flag (1:500) and HA (1:500) were from Cell Signaling (14793 and 3724, respectively), and YAP (1:200) was from Santa Cruz Biotechnology (sc-101199).
Secondary antibodies Alexa fluor 488 and 555 and phalloidin were from Invitrogen and used in 1:1000 dilution.
Chemicals sorbitol was from Fisher Scientific (S459-500), latrunculin B (Lat B) was from Abcam (ab144291), 2-deoxy-D-glucose (2-DG) was from Sigma (D8375), and p38 inhibitor SB203580 (1202) and JNK inhibitor SP600125 (1496) were from Tocris.
5
Quantitative Nuclear-Cytosolic YAP/TAZ Ratio
6
Phosphotyrosine-mediated JMJD1a regulation
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Standard western blotting techniques and Amersham ECL Plus Western blotting reagent were used. Following antibodies were used: JMJD1a (12835-1-AP, Proteintech, USA, 1:1,000), YAP/TAZ (sc-101199, Santa Cruz Biotechnology, USA, 1:500), GAPDH (5G4, HyB test 1:5,000), Tubulin (12G10, Hybridoma bank, 1:5,000), Lamin A/C (sc-7292, Santa Cruz Biotechnology, 1:1,000), H3K9me2 (#7658, Cell Signaling, 1:1,000), Histone 3 (#4499, Cell Signaling, 1:1,000), actin (clone AC-74, Sigma, 1:1,000), alpha-SMA (A2547, Sigma, 1:1,000) and antiphosphotyrosine antibody (APY03, Cytoskeleton, 1:1,000). For the phosphotyrosine pull-downs, MDA-MB-231 cells co-transfected with GFP-JMJD1a and CA-Src or empty vector were lysed and subjected to pulldown with beads of APY03 covalently coupled to sepharose (Anti-Phosphotyrosine Affinity Beads # APY03-Beads (Cytoskeleton) according to the manufacturer's instructions. The pulldowns and cell lysate were resolved on SDS–PAGE and subjected to western blot analysis with anti-GFP antibody (Abcam #1218).
Uncropped scans of the most important blots are provided asSupplementary Fig. 10 .
Uncropped scans of the most important blots are provided as
7
Immunofluorescence Staining of Cell Cultures
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Cells were fixed with 4% PFAH for 10 min at RT and simultaneously permeabilized and blocked with 0.3% Triton in 30% horse serum (Gibco) for 10 min at RT. Following antibodies and antibody dilutions were used: JMJD1a (12835-1-AP, Proteintech, 1:100), YAP/TAZ (sc-101199, Santa Cruz Biotechnology, 1:75), JMJD1a (sc-376608, Santa Cruz Biotechnology, 1:100), Atto-Phalloidin-647N (65906, Sigma, 1:200), Collagen I (NB600-408, Novus, 1:100) and Fibronectin (F3648, Sigma, 1:400).
8
Immunofluorescence Analysis of YAP/TAZ
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Immunofluorescence was performed on PFA-fixed cells. Primary antibody was YAP/TAZ (Santa Cruz Biotechnology no. sc-101199). F-actin was stained with Alexa Fluor 488 Phalloidin (Thermo Fisher Scientific). Secondary antibody (1:200) were from Molecular Probes. Samples were counterstained with ProLong-DAPI (Molecular Probes, Life Technologies) to label cell nuclei. Confocal images were obtained with a Leica TCS SP5 equipped with a CCD camera and analysed using Volocity software (PerkinElmer, version 5.5.1).
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