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Regulation of mitochondrial DNA homeostasis by a mitochondrial microprotein

Ajala, I.; Lipuma, D.; Bonnamour, G.; Vanderperre, B.

2026-02-21 biochemistry
10.64898/2026.02.20.706844 bioRxiv
Show abstract

The actin cytoskeleton and non-muscle myosins coordinate a wide array of cellular functions, yet their contribution to mitochondrial genome maintenance remains poorly understood. Here, we identify AltSLC35A4, a conserved protein encoded by an alternative open reading frame (AltORF), as a novel regulator of mitochondrial DNA (mtDNA) homeostasis. Using co-immunoprecipitation and mass spectrometry, we show that AltSLC35A4 interacts with actomyosin cytoskeletal regulators including MYH9 and MYH10, which have been previously implicated in mtDNA regulation. Loss of AltSLC35A4 increases mtDNA copy number and causes a dispersed spatial distribution of mitochondrial nucleoids, leading to the accumulation of extramitochondrial DNA puncta. Theses alterations occur without detectable changes in TFAM expression and mitochondrial membrane potential, suggesting that the mtDNA dysregulation is independent of mtDNA replication and transcription and mitochondrial bioenergetic state. Our findings uncover a previously unrecognized role for AltSLC35A4 in mitochondrial nucleoids regulation and highlight the functional importance of altORF-encoded proteins in mitochondrial biology.

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