Mitochondrial protein quality control is crucial for the maintenance of correct mitochondrial homeostasis. It is ensured by several specific mitochondrial proteases located across the various mitochondrial subcompartments. Here, we focused on characterization of functional overlap and cooperativity of proteolytic subunits AFG3L2 (AFG3 Like Matrix AAA Peptidase Subunit 2) and YME1L (YME1 like ATPase) of mitochondrial inner membrane AAA (ATPases Associated with diverse cellular Activities) complexes in the maintenance of mitochondrial structure and respiratory chain integrity. We demonstrate that loss of AFG3L2 and YME1L, both alone and in combination, results in diminished cell proliferation, fragmentation of mitochondrial reticulum, altered cristae morphogenesis, and defective respiratory chain biogenesis. The double AFG3L2/YME1L knockdown cells showed marked upregulation of OPA1 protein forms, with the most prominent increase in short OPA1 (optic atrophy 1). Loss of either protease led to marked elevation in OMA1 (OMA1 zinc metallopeptidase) (60 kDa) and severe reduction in the SPG7 (paraplegin) subunit of the m-AAA complex. Loss of the YME1L subunit led to an increased Drp1 level in mitochondrial fractions. While loss of YME1L impaired biogenesis and function of complex I, knockdown of AFG3L2 mainly affected the assembly and function of complex IV. Our results suggest cooperative and partly redundant functions of AFG3L2 and YME1L in the maintenance of mitochondrial structure and respiratory chain biogenesis and stress the importance of correct proteostasis for mitochondrial integrity.

• Klíčová slova
• AAA complex, AFG3L2, YME1L, mitochondria, protease,
• MeSH
• ATPázy spojené s různými buněčnými aktivitami genetika   metabolismus   MeSH
• HEK293 buňky MeSH
• lidé MeSH
• metaloendopeptidasy genetika   metabolismus   MeSH
• mitochondriální membrány metabolismus   MeSH
• mitochondriální proteiny genetika   metabolismus   MeSH
• mitochondrie metabolismus   ultrastruktura   MeSH
• proliferace buněk genetika   fyziologie   MeSH
• proteasy závislé na ATP genetika   metabolismus   MeSH
• transmisní elektronová mikroskopie MeSH
• western blotting MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• AFG3L2 protein, human MeSH Prohlížeč
• ATPázy spojené s různými buněčnými aktivitami MeSH
• metaloendopeptidasy MeSH
• mitochondriální proteiny MeSH
• proteasy závislé na ATP MeSH
• YME1L1 protein, human MeSH Prohlížeč

p53 is a major cellular tumor suppressor that in addition to its nuclear, transcription-dependent activity is also known to function extranuclearly. Cellular stressors such as reactive oxygen species can promote translocation of p53 into mitochondria where it acts to protect mitochondrial genome or trigger cell death via transcription-independent manner. Here we report that the mammalian homologue of yeast mitochondrial Afg1 ATPase (LACE1) promotes translocation of p53 into mitochondria. We further show that LACE1 exhibits significant pro-apoptotic activity, which is dependent on p53, and that the protein is required for normal mitochondrial respiratory function. LACE1 physically interacts with p53 and is necessary for mitomycin c-induced translocation of p53 into mitochondria. Conversely, increased expression of LACE1 partitions p53 to mitochondria, causes reduction in nuclear p53 content and induces apoptosis. Thus, LACE1 mediates mitochondrial translocation of p53 and its transcription-independent apoptosis.

• Klíčová slova
• LACE1, apoptosis, mitochondria, p53, translocation,
• MeSH
• adenosintrifosfatasy metabolismus   MeSH
• apoptóza fyziologie   MeSH
• HEK293 buňky MeSH
• lidé MeSH
• mitochondriální proteiny metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• transfekce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adenosintrifosfatasy MeSH
• AFG1L protein, human MeSH Prohlížeč
• mitochondriální proteiny MeSH
• nádorový supresorový protein p53 MeSH
• TP53 protein, human MeSH Prohlížeč

Mitochondrial ATPases associated with diverse cellular activities (AAA) proteases are involved in the quality control and processing of inner-membrane proteins. Here we investigate the cellular activities of YME1L, the human orthologue of the Yme1 subunit of the yeast i-AAA complex, using stable short hairpin RNA knockdown and expression experiments. Human YME1L is shown to be an integral membrane protein that exposes its carboxy-terminus to the intermembrane space and exists in several complexes of 600-1100 kDa. The stable knockdown of YME1L in human embryonic kidney 293 cells led to impaired cell proliferation and apoptotic resistance, altered cristae morphology, diminished rotenone-sensitive respiration, and increased susceptibility to mitochondrial membrane protein carbonylation. Depletion of YME1L led to excessive accumulation of nonassembled respiratory chain subunits (Ndufb6, ND1, and Cox4) in the inner membrane. This was due to a lack of YME1L proteolytic activity, since the excessive accumulation of subunits was reversed by overexpression of wild-type YME1L but not a proteolytically inactive YME1L variant. Similarly, the expression of wild-type YME1L restored the lamellar cristae morphology of YME1L-deficient mitochondria. Our results demonstrate the importance of mitochondrial inner-membrane proteostasis to both mitochondrial and cellular function and integrity and reveal a novel role for YME1L in the proteolytic regulation of respiratory chain biogenesis.

• MeSH
• apoptóza MeSH
• ATPázy spojené s různými buněčnými aktivitami MeSH
• genový knockdown MeSH
• GTP-fosfohydrolasy metabolismus   MeSH
• lidé MeSH
• metaloendopeptidasy metabolismus   MeSH
• mitochondriální membrány metabolismus   MeSH
• mitochondriální proteiny MeSH
• mitochondrie metabolismus   MeSH
• NADH, NADPH oxidoreduktasy metabolismus   MeSH
• proliferace buněk * MeSH
• proteasy závislé na ATP metabolismus   MeSH
• proteasy metabolismus   MeSH
• protein - isoformy metabolismus   MeSH
• respirační komplex I MeSH
• respirační komplex IV metabolismus   MeSH
• Saccharomyces cerevisiae - proteiny metabolismus   MeSH
• Saccharomyces cerevisiae cytologie   metabolismus   MeSH
• transport elektronů * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ATPázy spojené s různými buněčnými aktivitami MeSH
• GTP-fosfohydrolasy MeSH
• metaloendopeptidasy MeSH
• mitochondriální proteiny MeSH
• NADH, NADPH oxidoreduktasy MeSH
• NDUFB6 protein, human MeSH Prohlížeč
• OPA1 protein, human MeSH Prohlížeč
• proteasy závislé na ATP MeSH
• proteasy MeSH
• protein - isoformy MeSH
• respirační komplex I MeSH
• respirační komplex IV MeSH
• Saccharomyces cerevisiae - proteiny MeSH
• YME1 protein, S cerevisiae MeSH Prohlížeč
• YME1L1 protein, human MeSH Prohlížeč