I have these samples where a few of them have really high expression of the TP53 gene in exons 8-10. Overall the expression is higher for these samples but its mainly coming from these exons and I'm just wondering how to interpret this? And what if it was reverse, that is instead these sample had lower overall expression but it was due to the last few exons being really low? thanks in advance.
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simplitia •
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I don't think it means anything biologically, as exons are not transcribed individually. One option is that the transcript is degraded unevenly, giving rise to higher amount of sequence preserved in those regions. This is either because that gene region is furthest from ends (most RNases are exos rather than endos), or maybe it has a stable secondary structure that resists degradation. It could also be some kind of sequencing artifact. Either way, I don't think there is any biological relevance to it, as it would be extremely unlikely that there is a functional protein isoform that is made only from those exons.
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What method are you using to determine exon expression levels?
rpolicastro I'm using, subread featureCounts
featureCounts should only be used for gene level quantification, since there are a few challenges in exon and transcript level quantification that it's not designed to handle. I would instead recommend [Salmon]https://salmon.readthedocs.io/en/latest/salmon.html) for transcript level quantification, and fishpond if you want to do differential isoform usage. In most common use cases exon quantification itself is not informative since the functional unit of gene expression is the transcript, which is a collection of exons.