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The human leukocyte antigen (HLA) class II proteins present peptides to CD4+T cells through an interaction with T cell receptors (TCRs). Thus, HLA proteins are key players in shaping immunogenicity and immunodominance. Nevertheless, factors governing peptide presentation by HLA-II proteins are still poorly understood. To address this problem, we profiled the blood transcriptome and immunopeptidome of 20 healthy individuals and integrated the profiles with publicly available immunopeptidomics datasets. In depth multi-omics analysis identified expression levels and subcellular locations as import sequence-independent features governing presentation. Levering this knowledge, we developed the Peptide Immune Annotator Multimodal (PIA-M) tool, as a novel pan multimodal transformer-based framework that utilises sequence-dependent along with sequence-independent features to model presentation by HLA-II proteins.PIA-Millustrated a consistently superior performance relative to existing tools across two independent test datasets (area under the curve: 0.93 vs. 0.84 and 0.95 vs. 0.86), respectively. Besides achieving a higher predictive accuracy,PIA-Mwith its Rust-based pre-processing engine, had significantly shorter runtimes.PIA-Mis freely available with a permissive licence as a standalone pipeline and as a webserver (https://hybridcomputing.ikmb.uni-kiel.de/pia). In conclusion,PIA-Menables a new state-of-the-art accuracy in predicting peptide presentation by HLA-II proteinsin vivo.

Detyrosination is a major post-translational modification of microtubules (MT), which has significant impact on MT function in cell division, differentiation, growth, migration, polarity, and intracellular trafficking. Detyrosination of α-tubulin occurs via the recently identified complex of vasohibin 1/2 (vash1/2) and small vasohibin binding protein (SVBP). However, there is still remaining detyrosinating activity in the absence of vash1/2/SVBP, and little is known about the regulation of detyrosination. Using cellular and cell-free assays we showed that the calcium-dependent proteases calpains 1 and 2 regulate MT detyrosination. We identified new calpain cleavage sites in the N-terminal disordered region of vash1 using in vitro proteolysis followed by mass spectrometry. However, this cleavage did not affect the detyrosination activity of vasohibin. In conclusion, the regulation of MT detyrosination by calpains occurs via another yet unknown tubulin carboxypeptidase. Importantly, calpains’ calcium dependency could allow a fine regulation of MT detyrosination. Thus, identifying the calpain-regulated pathway of MT detyrosination can be of major importance for several basic and clinical research and should be focused on in future studies.Summary StatementThe conventional calpains 1 and 2 play an important role in the regulation of microtubule detyrosination in a vasohibin independent way. Thus, they possibly control another still unknown tubulin carboxypeptidase.