We made the global transcriptome (global mRNA expression), by RNAseq, from human subcutaneous adipocytes. Subsequently we did the Western blot for some proteins that were determined from the results of RNAseq, however we had a decrease in mRNA due to the treatment and when we analyzed the respective protein it was shown to be increased by the treatment. We know that there are cytoplasmic factors that stabilize mRNA or even enhance the translation of a certain protein, but we need a robust basis to explain this decrease in mRNA and increase in protein.
That can be explained two ways:
1) Your protein and mRNA analyses only represent that very specific moment in time similar to a picture in an entire movie. Therefore, you may have missed (sampled after) the rise in mRNA and only detected the increase in protein abundance.
2) You may have impacted the rate of protein translation from mRNA without increasing the mRNA abundance. Remember the number of ribosomes that are translating mRNA is not constant nor is it always proceeding at the same speed. Therefore, lower mRNA can result in more cognate protein expression if the translation rate per unit mRNA increases.
I hope that helps.
Gordon K Murdoch, I really appreciate your help.
Could you, please, indicate articles that cover your explanation to that I can go deeper into explaining these results. The biological samples that were used for mRNA (RNAseq) and protein (Western blot) came from the same biological aliquot. Thanks.
There are many, I recommend that you search and read articles on "regulation of translation in eukaryote" and the following may be a good start for comparison of mRNA versus protein expression:
Article The utility of protein and mRNA correlation
In addition to the explanation by Gordon K Murdoch, there are more two possible causes that can explain your results.
- As you obtained the samples for RNA-Seq and Western blot from the same biological aliquot, maybe the alterations in transcription don't were reflected in translation yet.
- The same human gene, like other species, can be processed into different transcripts isoforms (alternative splicing). The transcript isoforms can be protein-coding or non-coding. These transcripts can be non-coding even though the original gene be protein-coding. Conducting your analysis at the isoforms level rather than gene level, you will be able to identify whether more than one isoform from the same gene is being expressed. There is a possibility that a non-coding isoform is decreasing the overall expression of the coding gene and the increase in the expression of the protein-coding isoform is being covered up by this. Thus, you are unable to see the increase of protein-coding isoform of the gene.
Hope this helps.
Stellamaris Soares thanks for your explanation. You guys helped me a lot.
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