Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
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UniProt ID:

#P46937

Entrez-Gene Id:

10413

Product Includes Quantity Reactivity MW(kDa) Isotype
YAP (D8H1X) XP® Rabbit mAb 14074 100 µl H M R Hm Mk 65-78 Rabbit IgG
Phospho-YAP (Ser109) (E5I9G) Rabbit mAb 53749 100 µl H M R Mk 65-78 Rabbit IgG

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

PhosphoPlus® Duets from Cell Signaling Technology (CST) provide a means to assess protein activation status. Each Duet contains an activation-state and total protein antibody to your target of interest. These antibodies have been selected from CST's product offering based upon superior performance in specified applications.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

YAP (Yes-associated protein, YAP65) was first identified based on its ability to associate with the SH3 domain of Yes. It also binds to other SH3 domain-containing proteins such as Nck, Crk, Src, and Abl (1). In addition to the SH3 binding motif, YAP contains a PDZ interaction motif, a coiled-coil domain, and WW domains (2-4). While initial studies of YAP all pointed towards a role in anchoring and targeting to specific subcellular compartments, subsequent studies showed that YAP is a transcriptional co-activator by virtue of its WW domain interacting with the PY motif (PPxY) of the transcription factor PEBP2 and other transcription factors (5). In its capacity as a transcriptional co-activator, YAP is now widely recognized as a central mediator of the Hippo Pathway, which plays a fundamental and widely conserved role in regulating tissue growth and organ size (6-8). Phosphorylation at multiple sites (e.g., Ser109, Ser127) by LATS kinases promotes YAP translocation from the nucleus to the cytoplasm, where it is sequestered through association with 14-3-3 proteins (7-9). These LATS-driven phosphorylation events serve to prime YAP for subsequent phosphorylation by CK1δ/ε in an adjacent phosphodegron, triggering proteasomal degradation of YAP (10).

  1. Sudol, M. (1994) Oncogene 9, 2145-52.
  2. Mohler, P.J. et al. (1999) J Cell Biol 147, 879-90.
  3. Espanel, X. and Sudol, M. (2001) J Biol Chem 276, 14514-23.
  4. Sudol, M. et al. (1995) FEBS Lett 369, 67-71.
  5. Yagi, R. et al. (1999) EMBO J 18, 2551-62.
  6. Dong, J. et al. (2007) Cell 130, 1120-33.
  7. Zhao, B. et al. (2010) Genes Dev 24, 862-74.
  8. Zhao, B. et al. (2007) Genes Dev 21, 2747-61.
  9. Yu, F.X. et al. (2012) Cell 150, 780-91.
  10. Zhao, B. et al. (2010) Genes Dev 24, 72-85.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    PhosphoPlus is a registered trademark of Cell Signaling Technology, Inc.
    XP is a registered trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    使用に関する制限

    法的な権限を与えられたCSTの担当者が署名した書面によって別途明示的に合意された場合を除き、 CST、その関連会社または代理店が提供する製品には以下の条件が適用されます。お客様が定める条件でここに定められた条件に含まれるものを超えるもの、 または、ここに定められた条件と異なるものは、法的な権限を与えられたCSTの担当者が別途書面にて受諾した場合を除き、拒絶され、 いかなる効力も効果も有しません。

    研究専用 (For Research Use Only) またはこれに類似する表示がされた製品は、 いかなる目的についても FDA または外国もしくは国内のその他の規制機関により承認、認可または許可を受けていません。 お客様は製品を診断もしくは治療目的で使用してはならず、また、製品に表示された内容に違反する方法で使用してはなりません。 CST が販売または使用許諾する製品は、エンドユーザーであるお客様に対し、使途を研究および開発のみに限定して提供されるものです。 診断、予防もしくは治療目的で製品を使用することまたは製品を再販売 (単独であるか他の製品等の一部であるかを問いません) もしくはその他の商業的利用の目的で購入することについては、CST から別途許諾を得る必要があります。 お客様は以下の事項を遵守しなければなりません。(a) CST の製品 (単独であるか他の資材と一緒であるかを問いません) を販売、使用許諾、貸与、寄付もしくはその他の態様で第三者に譲渡したり使用させたりしてはなりません。また、商用の製品を製造するために CST の製品を使用してはなりません。(b) 複製、改変、リバースエンジニアリング、逆コンパイル、 分解または他の方法により製品の構造または技術を解明しようとしてはなりません。また、 CST の製品またはサービスと競合する製品またはサービスを開発する目的で CST の製品を使用してはなりません。(c) CST の製品の商標、商号、ロゴ、特許または著作権に関する通知または表示を除去したり改変したりしてはなりません。(d) CST の製品をCST 製品販売条件(CST’s Product Terms of Sale) および該当する書面のみに従って使用しなければなりません。(e) CST の製品に関連してお客様が使用する第三者の製品またはサービスに関する使用許諾条件、 サービス提供条件またはこれに類する合意事項を遵守しなければなりません。

    Revision 1
    #26699

    PhosphoPlus® YAP (Ser109) Antibody Duet

    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 1 Expand Image
    Flow cytometric analysis of RL cells (blue) and A-204 cells (green) using YAP (D8H1X) XP® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. 
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 2 Expand Image
    Simple Western™ analysis of lysates (1 mg/mL) from MCF-7 cells using YAP (D8H1X) XP® Rabbit mAb #14074. The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 3 Expand Image
    Confocal immunofluorescent analysis of fixed frozen mouse small intestine labeled with YAP (D8H1X) XP® Rabbit mAb (left, green) and co-labeled with ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 4 Expand Image
    Confocal immunofluorescent analysis of fixed frozen mouse kidney labeled with YAP (D8H1X) XP® Rabbit mAb (left, green) and co-labeled with ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 5 Expand Image
    Confocal immunofluorescent analysis of fixed frozen mouse cerebellum labeled with YAP (D8H1X) XP® Rabbit mAb (left, green) and co-labeled with ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 6 Expand Image
    Western blot analysis of extracts from control HeLa cells (Lane 1) or YAP knockout HeLa cells (Lane 2) using YAP (D8H1X) XP® Rabbit mAb (upper) or β-actin (13E5) Rabbit mAb #4970 (lower). The absence of signal in the YAP knockout HeLa cells confirms the specificity of the antibody for YAP.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 7 Expand Image
    Western blot analysis of extracts from HeLa cells (lane 1) or YAP knock-out cells (lane 2) using Phospho-YAP (Ser109) (E5I9G) Rabbit mAb #53749 (upper), and GAPDH (D6H11) XP® Rabbit mAb #5174 (lower). The absence of signal in the YAP knock-out HeLa cells confirms specificity of the antibody for YAP.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 8 Expand Image
    Western blot analysis of extracts from various cell lines using YAP (D8H1X) XP® Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). As expected, RL cells are negative for YAP protein expression.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 9 Expand Image
    Western blot analysis of extracts from PANC-1 cells and HeLa cells, untreated (-) or treated with calf intestinal phosphatase (CIP) and λ-phosphatase (+), using Phospho-YAP (Ser109) (E5I9G) Rabbit mAb (upper), YAP (D8H1X) XP® Rabbit mAb #14074 (middle), and β-Actin (D6A8) Rabbit mAb #8457 (lower).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 10 Expand Image
    Immunoprecipitation of YAP protein from A-204 cell extracts using Rabbit (DA1E) mAb XP® Isotype Control #3900 (lane 2) or YAP (D8H1X) XP® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using YAP (D8H1X) XP® Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb #3678 was used as a secondary antibody.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 11 Expand Image
    Western blot analysis of extracts from various cell lines using using Phospho-YAP (Ser109) (E5I9G) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). RL cells are negative for YAP expression, confirming specificity of the antibody. YAP expression is undetectable in RL extracts, as predicted from published gene expression databases and as confirmed with other YAP antibodies.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 12 Expand Image
    Immunohistochemical analysis of paraffin-embedded human non-Hodgkin lymphoma using YAP (D8H1X) XP® Rabbit mAb performed on the Leica® BOND Rx.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 13 Expand Image
    Immunoprecipitation of Phospho-YAP (Ser109) protein from PANC-1 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-YAP (Ser109) (E5I9G) Rabbit mAb. Western blot analysis was performed using Phospho-YAP (Ser109) Antibody #46931. Mouse anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127 was used for detection to avoid cross-reactivity with IgG.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 14 Expand Image
    Immunohistochemical analysis of paraffin-embedded human endometrioid adenocarcinoma using YAP (D8H1X) XP® Rabbit mAb performed on the Leica® BOND Rx.  
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 15 Expand Image
    Immunohistochemical analysis of paraffin-embedded human breast adenocarcinoma using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 16 Expand Image
    Immunohistochemical analysis of paraffin-embedded human breast adenocarcinoma using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 17 Expand Image
    Immunohistochemical analysis of paraffin-embedded human ovarian serous carcinoma using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 18 Expand Image
    Immunohistochemical analysis of paraffin-embedded human non-small cell lung carcinoma (left), colon carcinoma case 1 (middle) or colon carcinoma case 2 (right), using YAP (D8H1X) XP® Rabbit mAb (top) or TAZ (E9J5A) XP® Rabbit mAb #72804 (bottom).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 19 Expand Image
    Immunohistochemical analysis of paraffin-embedded human benign prostatic hyperplasia using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 20 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse small intesine using YAP (D8H1X) XP® Rabbit mAb (left) or YAP Antibody (right). These two antibodies detect unique, non-overlapping epitopes on YAP protein. The similar staining patterns obtained with both antibodies help to confirm the specificity of the staining.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 21 Expand Image
    Immunohistochemical analysis of paraffin-embedded BEN cell pellet, untransfected (left, YAP/TAZ low), YAP-transfected (middle) or TAZ-transfected (right), using YAP (D8H1X) XP® Rabbit mAb (top) or TAZ (E9J5A) XP® Rabbit mAb #72804 (bottom).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 22 Expand Image
    Immunohistochemical analysis of paraffin-embedded 293T cell pellet, wild-type (left, positive) or YAP/TAZ knockout (right, negative), using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 23 Expand Image
    Immunohistochemical analysis of paraffin-embedded PANC-1 (left) and RL (right) cell pellets using YAP (D8H1X) XP® Rabbit mAb.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 24 Expand Image
    Confocal immunofluorescent analysis of low confluence MCF 10A cells (left), high confluence MCF 10A (center), and YAP negative RL cells (right) using YAP (D8H1X) XP® Rabbit mAb (green). Blue pseudocolor in lower images = DRAQ5® #4084 (fluorescent DNA dye). Increased nuclear localization of YAP protein is seen in low confluence (proliferating) cells.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 25 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from NCI-H2052 cells and YAP (D8H1X) XP® Rabbit mAb, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across CTGF, a known target gene of YAP (see additional figure containing ChIP-qPCR data).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 26 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from NCI-H2052 cells and YAP (D8H1X) XP® Rabbit mAb, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across chromosome 6 (upper), including CTGF (lower), a known target gene of YAP (see additional figure containing ChIP-qPCR data).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 27 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from NCI-2052 cells and either YAP (D8H1X) XP® Rabbit mAb, or Normal Rabbit IgG #2729, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human CTGF Promoter Primers #14927, human SMYD3 intron 2 primers, and SimpleChIP® Human CTGF Upstream Primers #14928. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 28 Expand Image
    CUT&RUN was performed with NCI-H2052 cells and YAP (D8H1X) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding around TSPAN2, a known target gene of YAP (see additional figure containing CUT&RUN-qPCR data).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 29 Expand Image
    CUT&RUN was performed with NCI-H2052 cells and YAP (D8H1X) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across across chromosome 1 (upper), including TSPAN2 (lower), a known target gene of YAP (see additional figure containing CUT&RUN-qPCR data).
    PhosphoPlus® YAP (Ser109) Antibody Duet: Image 30 Expand Image
    CUT&RUN was performed with NCI-H2052 cells and either YAP (D8H1X) XP® Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human C1D downstream primers, human TSPAN2 upstream primers, and human ITM2A upstream primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.