Signaling Pathway Analysis of CRTC Proteins

All chemicals were obtained from Sigma unless otherwise stated. Antibodies to CREB (#9104), phospho S133 CREB (#9196), VASP (#3132), CRTC3 (#2720) and Lamin A/C (#4777) were from Cell Signaling Technologies. Antibodies to GAPDH (#MAB374) and CRTC2 (#ST1099) were from Merck-Millipore. Anti-BrdU antibody (B2531) was from Sigma Aldrich. Silencer® Select siRNA targeting CRCT2 and CRTC3 (#4390791 and #4390771 respectively) and negative control (#4390843) were purchased from Thermo Fisher. ON-TARGETplus SMARTpool CREB siRNA (#L-092995-02-0005) and ON-TARGETplus Non-targeting Control Pool (#D-001810-10-05) were purchased from Dharmacon. UO126, LY294002, SB 203580 were from Merck-Calbiochem. Bisindolylmaleimide I (BIS1, GF 109203×) was purchased from Tocris Bioscience. MK-2206 was from Selleckchem. HG-9-91-01 was a gift from Dr Kris Clark (University of Dundee)^{37 (link)}. Forskolin was purchased from Sigma Aldrich. BAY60-6583^{50 (link)} and Cicaprost were purchased from Caymen Chemical. PDGF-BB was purchased from R&D systems. 6-BNZ-cAMP-AM^{35 (link)}, a cell permeable selective PKA agonist, and 8-pCPT-2′-O-Me-cAMP-AM, a cell permeable selective EPAC super-agonist^{36 (link)}, were purchased from Biolog.

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Hudson C., Kimura T.E., Duggirala A., Sala-Newby G.B., Newby A.C, & Bond M. (2018). Dual Role of CREB in The Regulation of VSMC Proliferation: Mode of Activation Determines Pro- or Anti-Mitogenic Function. Scientific Reports, 8, 4904.

Publication 2018

Lamin A/C Anti-BrdU antibody (B2531) Silencer® Select siRNA targeting ON-TARGETplus Non-targeting Control Pool UO126 LY294002 SB 203580 MK-2206 Forskolin PDGF-BB 8-pCPT-2′-O-Me-cAMP-AM

Anti antibody Antibodies Bisindolylmaleimide i Brdu Cell Cicaprost Crtc2 Epac Forskolin Gapdh Hg 9 91 01 Lamin a c Ly294002 Mk 2206 Pdgf bb Permeable Sb 203580 Sirna Uo126 Vasp

Corresponding Organization :

Other organizations : Bristol Royal Infirmary, University of Bristol

Top 5 similar protocols

Variable analysis

independent variables

UO126
LY294002
SB 203580
Bisindolylmaleimide I (BIS1, GF 109203×)
MK-2206
HG-9-91-01
Forskolin
BAY60-6583
Cicaprost
PDGF-BB
6-BNZ-cAMP-AM
8-pCPT-2′-O-Me-cAMP-AM

dependent variables

CREB phosphorylation
CRTC2
CRTC3
BrdU incorporation

control variables

GAPDH
Lamin A/C

positive controls

Forskolin
6-BNZ-cAMP-AM
8-pCPT-2′-O-Me-cAMP-AM

negative controls

Silencer® Select siRNA targeting CRCT2 and CRTC3
ON-TARGETplus Non-targeting Control Pool

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