Mitochondrial matrix proteostasis is linked to hereditary paraganglioma: LON‐mediated turnover of the human flavinylation factor SDH5 is regulated by its interaction with SDHA

Abstract

Mutations in succinate dehydrogenase (SDH) subunits and assembly factors cause a range of clinical conditions. One such condition, hereditary paraganglioma 2 (PGL2), is caused by a G78R mutation in the assembly factor SDH5. Although SDH5(G78R) is deficient in its ability to promote SDHA flavinylation, it has remained unclear whether impairment to its import, structure, or stability contributes to its loss of function. Using import-chase analysis in human mitochondria isolated from HeLa cells, we found that the import and maturation of human SDH5(G78R) was normal, while its stability was reduced significantly, with ~25% of the protein remaining after 180 min compared to ~85% for the wild-type protein. Notably, the metabolic stability of SDH5(G78R) was restored to wild-type levels by depleting mitochondrial LON (LONM). Degradation of SDH5(G78R) by LONM was confirmed in vitro; however, in contrast to the in organello analysis, wild-type SDH5 was also rapidly degraded by LONM. SDH5 instability was confirmed in SDHA-depleted mitochondria. Blue native PAGE showed that imported SDH5(G78R) formed a transient complex with SDHA; however, this complex was stabilized in LONM depleted mitochondria. These data demonstrate that SDH5 is protected from LONM-mediated degradation in mitochondria by its stable interaction with SDHA, a state that is dysregulated in PGL2.

Mitochondrial complex II of plants: subunit composition, assembly, and function in respiration and signaling

“…Mutation of the yeast homolog SDHAF1 as well as the expression of variants corresponding to human mutants result in a reduction in SDH activity and the failure of OXPHOS‐dependent growth (Ghezzi et al ., ). Mutations in SDHAF1 affect its interaction with SDH2, resulting in an altered biogenesis of the SDH holo‐enzyme (Bezawork‐Geleta et al ., ; Na et al ., ; Maio et al ., ) (Figure ). Detailed analysis revealed that SDHAF1 transiently binds to aromatic peptides of SDH2 through an arginine‐rich region in its C‐terminus and specifically engages an Fe–S donor complex, consisting of the scaffold, holo‐ISCU and the co‐chaperone–chaperone pair, HSC20–HSPA9, through an LYR motif near its N‐terminal domain (Maio et al ., ).…”

New roles for mitochondrial proteases in health, ageing and disease

“…Also, in mammals, several proteins have been identified that are degraded by LONP under normal conditions, as well as when they are damaged or mutated. These include aconitase, cytochrome c oxidase subunit 4 isoform 1, steroidogenic acute regulatory protein (STAR), succinate dehydrogenase subunit 5, transcription factor A (TFAM) and glutaminase C, as well as some haem-related enzymes such as cystathionine β-synthase, haem oxygenase 1 and 5-aminolevulinate synthase [31][32][33][34][35][36][37][38] . Deficiency in mouse LONP1 induces alterations in mitochondrial respiration and in the OXPHOS system 39 .…”

The Assembly Factor SDHAF2 Is Dispensable for Flavination of the Catalytic Subunit of Mitochondrial Complex II in Breast Cancer Cells

“…The G78R mutation in human SDH5/SDHAF2 gene was shown to be associated with PGL2 (25), and we and others have reported that the mutation affects interaction of SDHAF2/SDH5 with SDHA (25)(26)(27)(28). SDHAF2/SDH5, which is not in complex with SDHA, is degraded by the Lon protease as part of the mitochondrial protein homeostasis (26,28). Initially, it was hypothesized that Sdh5 may be necessary and sufficient for flavination of Sdh1, although an NMR study showed that Sdh5 cannot bind FAD (27).…”

“…Sdh5 has been identified in yeast as the assembly factor responsible for flavination of Sdh1, which involves covalent attachment of the redox cofactor FAD (25). The G78R mutation in human SDH5/SDHAF2 gene was shown to be associated with PGL2 (25), and we and others have reported that the mutation affects interaction of SDHAF2/SDH5 with SDHA (25)(26)(27)(28). SDHAF2/SDH5, which is not in complex with SDHA, is degraded by the Lon protease as part of the mitochondrial protein homeostasis (26,28).…”