I have a transcription factor protein X and X has a co-activator protein Y. XY bind to the DNA and downstream expression of proteins occur. Now I block the activity of X through inhibitor but dont see any change in the binding of Y. When i check for downstream proteins, I observe low/reduced expression of downstream proteins. How is that possible if the XY binds normally? What could be the reasons? I am assuming there's change in the binding pattern of Y? Or the XY complex bind to the DNA in different pattern?
Hi,
To access this question, you might do the ChIP-seq on X & Y on untreated sample, then do the same on the sample after inhibition. If your inhibitor is blocking the binding of X, your X-peak on the enhancer of your protein of interest will disappear, while the peak Y will still be there. If the inhibitor is blocking the activity, but not the binding, the X-peak will still be present. With Y-antibody ChIP you may also check if the Y-binding pattern changed after the X-inhibition.
Best,
Michail
and also think that your X-Y model is very likely much tooo simple:
1) only a two component system?
2) what about other factors replacing the X-Y complex?
3) what about other pathways compensating for your pathway involving X-Y ???
I know questions over questions, hard to answer, but perhaps worthwhile doing...
Tobias Aurelius Knoch You are right! Indeed. I was searching for the whole interaction pathway and read articles to find that one of the other protein was competing for the X.
Thank you everyone for adding your views.
I would like to express my opinion. The gene expression is under the control of virus.
Binding-protein and/or Enzyme has many substrates (please see file; Multiple Hydrolase LIP). Avidin has recently turned out to be biotin/lipoic acid/amino acids-binding protein (please see file; Lipoic acid Avidin).
I have recently searched for Wnt signaling-related proteins in my protein database (please see file; HepG2 Fucoidan).
Normal liver tissue (with pseudo-liver cancer) has T-cell surface glycoprotein CD8 alpha chain at 0.64 μg/mg tissue protein.
LC tissue (with leprosy) has T-cell surface glycoprotein CD8 beta chain/CD8b at 2.1, Beta-Catenin/Catenin beta-1 at 3.2, Alpha-Catenin/Alpha E-catenin at 5.0, and Adenomatous polyposis coli protein/APC protein/Deleted in polyposis 2.5 at 26.0 μg/mg tissue protein, respectively. Total becomes to be 36.3 μg/mg tissue protein.
HCC tissue (with PBC) has CD30L receptor/Ki-1 antigen/Tumor necrosis factor (TNF) receptor superfamily member 8 at 3.8/CD40 ligand/T cell antigen GP39/TNF-related activation protein at 3.2, and Adenomatous polyposis coli protein/APC protein at 6.4 μg/mg tissue protein, respectively. Total becomes to be 13.4 μg/mg tissue protein.
LC tissue (named as No.6) has Adenomatous polyposis coli protein/APC protein at 21.2 μg/mg tissue protein.
HCC tissue (named as No.6) has CD99 antigen/T-cell surface glycoprotein E2 at 0.81, Adenosine deaminase at 0.11, and B-cell lymphoma 3 protein/BCL-3 protein/Proto-oncogene BCL3 at 1.2 μg/mg tissue protein, respectively. Total becomes to be 2.12 μg/mg tissue protein.
Hepatoma HepG2 (cultured without fucoidan) has CD5 antigen-like/IgM-associated peptide at 1.3, B-lymphocyte activator macrophage expressed/SLAM family member 8 at 0.19, and Axin-1/Axis inhibition protein 1 at 2.1 μg/mg cell protein, respectively. Total becomes to be 3.59 μg/mg cell protein.
Healed hepatocyte HepG2 (cultured with fucoidan) has T-cell leukemia homeobox protein 2 at 0.04, and T-lymphoma invasion and metastasis-inducing protein 1 at 2.7 μg/mg cell protein, respectively. Total becomes to be 2.74 μg/mg cell protein.
Fetal hepatocyte Hc has T-cell-specific kinase/Tyrosine-protein kinase ITK/TSK at 10.3 μg/mg cell protein.
Serum of Gait disorder patient has Alpha-2-macroglobulin receptor-associated protein/Low density lipoprotein (LDL) receptor-related protein associated protein 1/RAP at 4.6 μg/mg serum protein.
Healthy serum (33y, male) has Antigen-presenting glycoprotein CD1d/T-cell surface glycoprotein CD1d at 6.6, B-cell linker protein at 2.2, and Protein Wnt5a at 9.1 μg/mg serum protein, respectively.
The gene expression of Wnt signaling-related proteins is not linked to diseases.
Interestingly, these Wnt signaling-related proteins are up-regulated by Ebolavirus/EBOV, Respirovirus of Sendai virus/SeV, Rubulavirus of Simian virus 41/SV41 and Mumps virus/MuV, and Avulavirus of Newcastle disease virus/NDV (enveloped virus; -ssRNA).
Hi
Once you talk about a transcription factor, u need to consider it may not be absolute in effect and is generally supported by cascade of other proteins and factors to carry out any downstream activity. The factors can be pleiotropic and vice versa.
Following explanations:
1- In you case XY combination (as TF) exerted in an exaggerated/enhanced response, but similar response may not be solely dependent upon that TF only. Other proteins/factors may replace the activity.
2- Secondly, its never an ALL or NONE phenomena, eliminating the TF only doesn't eliminate the downstream activity, it sometimes just slows down. So this seems more likely in your X-Y combination case.
3- You need to focus on the target receptor as XY may not be the only factor stimulating/inhibiting them; other culprits may be there.
Regards
Dr Sikandar
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