Hello. I am using a tool called MOODS to scan some gene sequences for the TATA box motifs. A sample output from MOODS looks like this:
seq_1,MA0001.pfm,13329,+,5.23551915867,CCATAATTGC,
where each field is the sequence name, Position Frequency Matrix file name, position of the motif hit, strandedness indicates whether the match is versus the input strand (+) or the reverse complement (-), match score and sequence of the hit in the original sequence respectively
I want to know if the strand match + or - here is important for promoter analysis. For instance for promoter motif analysis in a + strand gene sequence, will the + hits from MOODS be more important or relevant than the - hits? Similarly, for promoter motif analysis in a - strand gene, will - hits from MOODS be more relevant than the + hits? Or does the strand match not matter, and only the position of the hit with respect to the predicted TSS matters?
I want to understand this better because for some sequences from a + strand gene, I get only one hit in MOODS and the strand match is - I am not sure how to consider this for obtaining a set of high relevance hits.
It would be great if someone could help me with this.
Thank you!
1 answer
MOODS scans both strands by default, so a - hit just means the PWM matched the reverse complement at that position. It's a real occurrence, not an artefact.
Whether it matters comes down to your input orientation, which I suspect is the actual source of the confusion. If you reoriented each promoter to run 5'->3' in the gene's direction before scanning, then for something directional like TATA the + hits are the ones that could serve that gene and a - hit is antisense. If you scanned raw genomic plus strand without reorienting, then a minus-strand gene's genuine TATA legitimately turns up as a - hit and that one is the relevant one. So "plus strand gene but the only hit is minus" is completely normal in the second case and a warning sign in the first.
Quickest way to work out which situation you're in: take one promoter you're sure about and search it for the TATA consensus TATAWAWR on both strands, then look at whether the plus or the minus hits land ~25-35 bp upstream of your TSS. That's consensus matching rather than PWM scoring so it doesn't replace MOODS, it just tells you which way your input is facing. I work on SeqBench so discount accordingly, https://seqbench.com/tools/motif-finder does IUPAC codes with both-strand scanning; any pattern searcher does the same job.
One other thing worth flagging -- MA0001 is AGL3 in JASPAR, a plant MADS-box factor, not TBP. TBP is MA0108. If TATA boxes are what you're after then you're scanning with the wrong matrix and the strand question is moot until you swap it.
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