how to determine whether protein level increase is resulted from mRNA stability - (Aug/26/2006 )

Hi,

I am wondering what kind of technology I can use to determine whether protein level increase is resulted from the enhancement of mRNA stability or posttranslation modification if it is not caused by the up-regulation in transcription level?

Anyone can help? Thanks.

If you think that mRNA stability is enhanced I would conduct a stability assay. Basically use a global transcritpional inhibitor such as actinomycin D and quantitatively assess how fast your mRNA of choice degades over time between control and experimental samples. This will associate mRNA stability with your increase in translation, but to really nail it, you should probably do a polysome analysis to see if your mRNA is more represented in polyribosome fractions compared to monoribosomes in your experimental conditions.