Transcription Factors

Daniel DurochD at IRCM.UMontreal.CA
Sat Mar 16 16:11:42 EST 1996


Brent and Julaine Bilhartz wrote:
> 
> If you had a transcription factor in hand how would you hunt down novel
> genes that it regulates. The consensus binding sequence is known. Any and
> all ideas welcome. Please respond by email bilha001 at gold.tc.umn.edu
> 
>             Thanks,
>                Brent

Brent,

Conceptually, I think the best way to identify novel targets is via 
differential display.  First, you transfect a cell line with an inducible 
expression system (e.g. tet repressor) or with a ts mutant of your 
transcription factor (if you have one).  You establish at least two 
different cell lines.  You only pick bands on your dd-PCR gels that are 
not present in the absence of inducer and that are consistently present 
in different lines upon induction.  You then use those bands as probes to 
perform Northern blots to confirm your results and then use them to 
screen genomic libraries to get the promoter. Then it is classical 
transcription work.

A very good illustration of this scheme to clone targets for 
transcription factors is published in a paper in EMBO J. about the 
identification of cyclin D as a transcription target for p53 (I am not 
sure).  It was published less than a year ago and you can write to me if 
you need the detailed reference I will look in my files.

I hope it helped you.

Daniel



More information about the Proteins mailing list