Supplementary Figure 4: Analysis of HLA ligand cleavage signatures.

(a, b) Cleavage signature analysis for MCL patient HLA-I ligands. Frequencies of 20 amino acids plus protein terminal position (-) in for 6 amino acids upstream of the C-terminus of HLA-I ligand (-6 to -1) and 6 amino acids downstream of HLA-I ligands (+1 to +6) are compared to the background distribution (n=22100 and n=42906) to determine amino acid enrichment and depletion surrounding HLA-DR ligands. Colors of the heat-map (a) and sizes of the IceLogo plot (b) letters indicate fold-change. The logo plot only includes statistically significant enrichment (P<0.0001). Compared to HLA-DR, HLA-I showed preferences for presences of tryptophan in both upstream and downstream sequences. (c) Comparison of cleavage signatures distinguishing flanking sequences of HLA-I and HLA-DR ligands. The IceLogo plot only includes statistically significant enrichments/depletions (P<0.001, two-tailed independent t-test by IceLogo⁶⁰). Methionines, lysines, glutamines, and histidines are notably enriched in sequences flanking HLA-I ligands (n=22100) compared to the same regions of HLA-DR ligands (n=12150). In contrast, tryptophans, glycines, and tyrosines appear depleted from HLA-I flanks relative to HLA-DR. Cleavage signatures of (d) JeKo-1 mantle cell lymphoma cell lines, (e) L128 mantle cell lymphoma cell lines, (f) patient melanoma tissues, and (g) MUTZ3 dendritic cell lines. Each cell or tissue type exhibits variable cleavage signatures, but their HLA-DR ligands consistently show enrichment for peptides at the tails of protein (-) and depletion for proline (P) and histidine (H) flanking regions. Cleavage signatures of our lymphoma HLA-I and HLA-II ligands compared to two previous studies. (h) HLA-II ligands from dendritic cell line MUTZ3 (HLA-DRB1*10, HLA-DRB1*11, n=11419)¹ were profiled by Mommen et al. 2016 Amino acid frequencies at six amino acids upstream and downstream of the presented ligands were compared to their counterparts from our MCL HLA-II ligands. ICE logo plots showed statistically significant enrichment or depletion in each position (p-value cut-off = 0.001). Methionine showed enrichment in dendritic cell cleavage signatures, and prolines showed stronger depletions at the majority of positions. (i) Similarly HLA-I ligands from B-lymphoblastoid cells profiled by Abelin et al. 2017 (HLA-A*01:01, HLA-A*02:01, HLA-B*35:01, n=4857)² were compared to our MCL HLA-I ligands. Fewer significant differences were observed. Phenylalanines were enriched in B-lymphoblastoid cleavage signatures.