ProSubzero

I have interesting results and I'm not sure where to go from here.

I have a cell line stably transfected with the wild type gene, and two mutated and constitutively active versions of the gene. The mRNA and protein expression of the gene is very high compared to the naive cells. So all is well with the cell lines.

I identified a gene whose mRNA expression is greatly reduced as a result of the two mutant forms of the gene compared to the transfected wild type and naive cells. It was reduced by at least hundred(s) of folds. It is a significant reduction. This was confirmed via microarray, RT-qPCR and Northern blots. I'm pretty confident in these results.

When I examined the protein expression of this identified gene, the mutated transfected cell lines expressed this gene at a much higher rate than the wild type gene. The wild type transfected cell line had reduced expression of the protein compared to the naive, and the mutated transfection doubled the protein expression. I've repeated these Westerns a few times. Again, I'm pretty confident in this part as well.

So in summary, mRNA indicates the gene is suppressed in mutant cell lines. Protein indicated the gene is increased in the mutant cell lines.

One problem here is that the regulation and signalling upstream of this gene isn't well known at all. I don't exactly have specific targets to go after.

I haven't had experience with this... anybody have any ideas/suggestions/places to look at? Any feed back would be most welcome.

Thanks!