IRFinder Diff output interpretations

[rezarahman12]

Hi Claudio I've just got question to clarify the IRFinder Diff interpretation.

the log2FoldChange values reported in the "CTR_TTX_DESeq2_WT.tsv" file would indicate the log2-fold change in intron retention levels between the CTR and TTX conditions. So, a positive log2-fold change value refers to high IRratio in CTR than TTX. Similarly, a negative log2-fold change value refers to a lower IRratio in CTR than TTX. Are the interpretations correct? I'm asking this because in .out file after running the IRFinder Diff- I see the below which denotes TTX versus CTR but in the results file I see the opposite CTR versis TTX. Could you please clarify this point?

I appreciate your time.

[1] "cooks_cutoff disabled" [1] "independentFiltering disabled" [1] "processing file 1 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr1/IRFinder-IR-nondir.txt" [1] "processing file 2 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr2/IRFinder-IR-nondir.txt" [1] "processing file 3 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr3/IRFinder-IR-nondir.txt" [1] "processing file 4 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX1/IRFinder-IR-nondir.txt" [1] "processing file 5 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX2/IRFinder-IR-nondir.txt" [1] "processing file 6 at /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX3/IRFinder-IR-nondir.txt" [1] "LowCover" [1] 0.1 [1] "Warning removed: 137410" [1] "Ratio removed: 180770" [1] "Combined removed: 200323" [1] "Intercept" "Condition_TTX_vs_CTR"
[3] "ConditionCTR.IRFinderIR" "ConditionTTX.IRFinderIR"

[CloXD]

Dear Reza, The wrapper of DESeq2 can take as input multiple groups, and it computes the differential IR for each pairwise comparison. Let's say you have 3 groups: A, B, and C, it will compare A_B, A_C and B_C. For each of the comparisons, you get a file A_B_DESeq2.tsv containing the comparisons of the groups in A using B as control ( positive Log2FoldChange are higher in A ).

You can check in all_results_DESeq2.tsv, which contains the information of all the comparisons, what is the average IRratio in each group for each intron. The output you showed in bold is part of the standard output of DESeq2. Cheers, Claudio

[rezarahman12]

Hi Claudio Thank you so much for your quick response. I'm sorry I still ask you some clarification on this. Since the file paths I gave in IRFinder Diff as below- singularity exec -e /scratch/project_mnt/S0077/IRFinder/IRFinder_v_2.0.1/IRFinder IRFinder Diff \ -g:CTR /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr1/IRFinder-IR-nondir.txt \ -g:CTR /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr2/IRFinder-IR-nondir.txt \ -g:CTR /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/CTR/Ctr3/IRFinder-IR-nondir.txt \ -g:TTX /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX1/IRFinder-IR-nondir.txt \ -g:TTX /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX2/IRFinder-IR-nondir.txt \ -g:TTX /scratch/project_mnt/S0077/scaling_IRFinder_v_2.0.1/TTX/TTX3/IRFinder-IR-nondir.txt \ Here is two condition one is CTR (control) and TTX (treatment). So, the log2foldchange in IRFinder reported "CTR_TTX_DESeq2_WTneurons" file is based on CTR compared to TTX, so positive value denotes downregulation in TTX (treatment) while negative log2foldchange refers to upregulated in TTX(treatment). Is that correct? I'm sorry for taking your time. Best regards Reza

[CloXD]

Hello Reza, Yes, it's correct. The first group in the file's name ( CTR in your case ) is the target, and the second is the control ( TTX ). To verify that it's correct, check in the file "all_results_DESeq.tsv" an event with positive log2FoldChange, the column CTR mean should be higher than the column TTX mean. If you reverse the sign of the log2FoldChange, you can obtain the same result as you were using TTX as target and CTR as control. Cheers, Claudio

[rezarahman12]

thanks