Dataset Information

Impaired protein import drives mitochondrial dysfunction upon mutation of TIMM50 in mitochondrial disease

ABSTRACT: TIMM50 is a core subunit of the TIM23 complex, the mitochondrial inner membrane translocase responsible for the import of presequence-containing precursors into the matrix or inner membrane. Here we describe a mitochondrial disease patient that is homozygous for a novel variant in TIMM50 and establish the first proteomic profile of mitochondrial disease associated with TIMM50 mutation. We demonstrate that TIMM50 pathogenic variants reduce the levels and activity of endogenous TIM23 complex, which significantly impacts the mitochondrial proteome, resulting in a combined oxidative phosphorylation (OXPHOS) defect and changes to mitochondrial ultrastructure. Using proteomic data sets from TIMM50 patient cells and a TIMM50 HEK293T cell model of disease, we reveal that laterally inserted substrates imported via the TIM23SORT complex are more sensitive to loss of TIMM50. Indeed, proteins involved in OXPHOS and mitochondrial ultrastructure are enriched in laterally-released TIM23SORT substrates, providing a biochemical mechanism for the specific defects in TIMM50 patients. These results highlight the power of using proteomics for elucidating molecular mechanisms of disease and uncovering novel features of fundamental biology.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Kidney Cell, Fibroblast

DISEASE(S): 3-methylglutaconic Aciduria Type 9

SUBMITTER: Jordan Crameri

LAB HEAD: Diana Stojanovski

PROVIDER: PXD047774 | Pride | 2024-07-03

REPOSITORIES: Pride

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			Action	DRS
	220207_Jordan_JC-5756_1.raw	Raw
	220207_Jordan_JC-5756_2.raw	Raw
	220207_Jordan_JC-5756_3.raw	Raw
	220207_Jordan_JC-5756_4.raw	Raw
	220207_Jordan_JC-5756_5.raw	Raw

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Publications

Reduced Protein Import via TIM23 SORT Drives Disease Pathology in TIMM50-Associated Mitochondrial Disease.

Crameri Jordan J JJ Palmer Catherine S CS Stait Tegan T Jackson Thomas D TD Lynch Matthew M Sinclair Adriane A Frajman Leah E LE Compton Alison G AG Coman David D Thorburn David R DR Frazier Ann E AE Stojanovski Diana D

Molecular and cellular biology 20240603 6

TIMM50 is a core subunit of the TIM23 complex, the mitochondrial inner membrane translocase responsible for the import of pre-sequence-containing precursors into the mitochondrial matrix and inner membrane. Here we describe a mitochondrial disease patient who is homozygous for a novel variant in <i>TIMM50</i> and establish the first proteomic map of mitochondrial disease associated with TIMM50 dysfunction. We demonstrate that TIMM50 pathogenic variants reduce the levels and activity of endogenou ...[more]

PMID: 38828998

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Project description:Background: Transcription control of mitochondrial metabolism is essential for cellular function. A better understanding of this process will aid the elucidation of mitochondrial disorders, in particular of the many genetically unsolved cases of oxidative phosphorylation (OXPHOS) deficiency. Yet, to date only few studies have investigated nuclear gene regulation in the context of OXPHOS deficiency. In this study, we combined RNA sequencing of human complex I-deficient patient cells across 32 conditions of perturbed mitochondrial metabolism, with a comprehensive analysis of gene expression patterns, co-expression calculations and transcription factor binding sites. Results: Our analysis shows that OXPHOS genes have a significantly higher co-expression with each other than with other genes, including mitochondrial genes. We found no evidence for complex-specific mRNA expression regulation in the tested cell types and conditions: subunits of different OXPHOS complexes are similarly (co-)expressed and regulated by a common set of transcription factors. However, we did observe significant differences between the expression of OXPHOS complex subunits compared to assembly factors, suggesting divergent transcription programs. Furthermore, complex I co-expression calculations identified 684 genes with a likely role in OXPHOS biogenesis and function. Analysis of evolutionarily conserved transcription factor binding sites in the promoters of these genes revealed almost all known OXPHOS regulators (including GABP, NRF1/2, SP1, YY1, E-box factors) and a set of six yet uncharacterized candidate transcription factors (ELK1, KLF7, SP4, EHF, ZNF143, and EL2). Conclusions: OXPHOS genes share an expression program distinct from other mitochondrial genes, indicative of targeted regulation of this mitochondrial sub-process. Within the subset of OXPHOS genes we established a difference in expression between subunits and assembly factors. Most transcription regulators of genes that co-express with complex I are well-established factors for OXPHOS biogenesis. For the remaining six factors we here suggest for the first time a link with transcription regulation in OXPHOS deficiency. RNA-SEQ of whole cell RNA in 2 control and 2 complex I deficient patient fibroblast cell lines treated with 4 compounds in duplicate, resulting in a total of 2x2x4x2=32 samples