The cardiac-enriched microprotein mitolamban regulates mitochondrial respiratory complex assembly and function in mice

Abstract

Emerging evidence indicates that a subset of RNA molecules annotated as noncoding contain short open reading frames that code for small functional proteins called microproteins, which have largely been overlooked due to their small size. To search for cardiac-expressed microproteins, we used a comparative genomics approach and identified mitolamban (Mtlbn) as a highly conserved 47-amino acid transmembrane protein that is abundantly expressed in the heart. Mtlbn localizes specifically to the inner mitochondrial membrane where it interacts with subunits of complex III of the electron transport chain and with mitochondrial respiratory supercomplexes. Genetic deletion of Mtlbn in mice altered complex III assembly dynamics and reduced complex III activity. Unbiased metabolomic analysis of heart tissue from Mtlbn knockout mice further revealed an altered metabolite profile consistent with deficiencies in complex III activity. Cardiac-specific Mtlbn overexpression in transgenic (TG) mice induced cardiomyopathy with histological, biochemical, and ultrastructural pathologic features that contributed to premature death. Metabolomic analysis and biochemical studies indicated that hearts from Mtlbn TG mice exhibited increased oxidative stress and mitochondrial dysfunction. These findings reveal Mtlbn as a cardiac-expressed inner mitochondrial membrane microprotein that contributes to mitochondrial electron transport chain activity through direct association with complex III and the regulation of its assembly and function.

Proteomics‐driven identification of short open reading frame‐encoded peptides

“…In the past decade, SEP identification has changed from accidental discovery to high‐throughput screening, thus finding more critical functional SEPs. Figure 3 summarized the history of high‐throughput identification of SEPs in different species [8, 10–12, 29–30, 38‐71] and several representative SEPs with important biological functions [5, 14, 16, 18–21, 36, 72‐91].…”

“…The history of high‐throughput identification of SEPs in different species by Ribo‐seq [29, 34‐71] and MS‐based proteomics [8, 10‐12, 33, 38‐56], and representative functional SEPs [5, 14, 16, 18‐21, 36, 72‐91]…”

Human LINE-1 retrotransposons: impacts on the genome and regulation by host factors

“…And do such second ORFencoded proteins have any physiological functions, with ORF2p as a precedent? Although several lines of evidence suggest that translation of polycistronic mRNAs, some of which encode "microproteins", occurs more frequently than expected in eukaryotes (Kondo et al, 2007;Brubaker et al, 2014;Karginov et al, 2017;Makarewich et al, 2022), further proteomic studies and functional validation of these products will be needed to elucidate these questions.…”

Mitochondrion‐located peptides and their pleiotropic physiological functions

“…A correlation of Stmp1 with innate immunity was also reported: Stmp1‐deficient macrophages were compromised for the NOD‐like receptor 3 (Nlrp3) inflammasome formation accompanied by a reduction in expression of cleaved Caspase‐1 and IL‐1β at protein levels, suggesting a role of Stmp1 in the activation of the inflammasome but not the priming process [73]. In addition, a recent study reported that cardiac‐specific overexpression of Stmp1 in mice induced cardiomyopathy [72]. These evidence indicates a key role for Stmp1 in diverse physiological functions.…”

“…Except for Stmp1, the origin gene 1810058I24Rik was also predicted to translate into other non-AUG peptides [71]. Stmp1 was also reported to interact with other subunits of CIII [72]. In the mouse heart, its inactivation altered CIII assembly dynamics, reduced CIII activity, and resulted in an altered metabolite profile; while overexpression of Stm-p1 caused increased oxidative stress and mitochondrial dysfunction [72].…”