Fig. 1: Proteogenomic screen in human endothelial cells (HAECs) identifies MOCCI as an inflammatory Mito-SEP (i-Mito-SEP).

a WGCNA-GSEA analysis of mito versus non-mito-SEPs in human failing heart reveals strong anti-correlation of mito-SEPs with inflammation. Each row represents one pathway and the pathways are separated into those that are involved in metabolism and those that are involved in inflammation. Color scale = NES score. Dataset used: European Genome-Phenome Archive EGAS00001002454¹². b HAECs from two healthy male donors were treated with 1 ng/mL IL-1β for 45 min, 12 and 24 h. All timepoints including untreated controls were analyzed by RNA-seq and Ribo-seq in triplicate. NF-kb reporter activity, ICAM-1, and VCAM-1 expression illustrate the dynamics of the HAEC response: an increase in inflammation up to 12 h followed by resolution thereafter. Data for NF-kb reporter activity are presented as mean ±  SEM. n = 4 biological replicates for NF-kb reporter activity. c Workflow to identify i-Mito-SEPs. Blue numbers indicate the number of SEPs called by RiboTaper and black numbers indicate the number of genes encoding those SEPs (see methods). d Summary of the mitochondrial gene signature workflow. (Bottom) UMAP of gene module association determination (G-MAD) scores of MitoCarta and random genes in human colon, skin, skeletal muscle (SKM), and PBMCs undergoing inflammation. Color scale indicates the percentage of mitochondrial genes in each cluster. Based on their G-MAD, the 240 candidate genes from c were assigned to a cluster in each dataset. (Top) Heatmap depicting the percentage of mitochondrial genes in a candidate’s cluster in each of the four datasets. Each column represents one candidate. Top scoring candidates were returned to the pipeline in c for further consideration. Datasets used were: GSE11223⁸⁴, GSE14905⁸³, GSE120862⁸², and GSE9820⁸¹. e Volcano plot of the maximal fold change of the 240 candidate genes during the IL-1β treatment compared to untreated cells. The size of each dot represent the highest TPM reached at any timepoint, while its color indicates if candidate SEP was predicted to localize to the mitochondria by motif, G-MAD or both. The p adjusted values were attained by applying the Wald chi-squared test, followed by correction for multiple testing using the Benjamini and Hochberg method through the DESeq2 pipeline. f MOCCI transcript and translation levels increase by 1000-fold after IL-1β treatment. TE translational efficiency. g Ribo-seq derived p-site read coverage across the MOCCI locus. In-frame (0) and out-of-frame (+1,+2) colored in orange, green, and blue respectively. Inset on the left shows that p-site reads are detected only at the start codon. Inset on the right summarizes the total number of in-frame (0) and out-of frame (+1,+2) reads in dataset.