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![Fcj1 is enriched at CJs. (A) Immunogold labeling of Fcj1 in wild-type cells. (B) Representation of gold particles after immunogold labeling of Fcj1 plotted on a model of CM, IBM, and OM (Vogel et al., 2006). (C and D) Quantification of protein densities in the IM. The number of gold particles occurring within a 14-nm distance from the IM was determined by moving a sliding window in silico along the IM in the model from bottom to top (raw data from C and from Vogel et al. [2006]). Gray boxes indicate the CJ region. The protein densities of Fcj1 and Cox2 (C) and of Su g and Su e (D) are shown. Bars, 100 nm.](publication/26293816/figure/fig2/AS:11431281177874473@1690636022564/Fcj1-is-enriched-at-CJs-A-Immunogold-labeling-of-Fcj1-in-wild-type-cells-B.jpeg)
Fcj1 is enriched at CJs. (A) Immunogold labeling of Fcj1 in wild-type cells. (B) Representation of gold particles after immunogold labeling of Fcj1 plotted on a model of CM, IBM, and OM (Vogel et al., 2006). (C and D) Quantification of protein densities in the IM. The number of gold particles occurring within a 14-nm distance from the IM was determined by moving a sliding window in silico along the IM in the model from bottom to top (raw data from C and from Vogel et al. [2006]). Gray boxes indicate the CJ region. The protein densities of Fcj1 and Cox2 (C) and of Su g and Su e (D) are shown. Bars, 100 nm.
Source publication
Crista junctions (CJs) are important for mitochondrial organization and function, but the molecular basis of their formation and architecture is obscure. We have identified and characterized a mitochondrial membrane protein in yeast, Fcj1 (formation of CJ protein 1), which is specifically enriched in CJs. Cells lacking Fcj1 lack CJs, exhibit concen...
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Citations
... Despite extensive study, the definitions of Mic60's C-terminal Mitofilin domain boundary vary. While it has been established that the Mitofilin domain extends to the C terminus, multiple N-terminal boundaries have been proposed 10,26,34,37,38 . We superimposed AlphaFold2 predictions of Mic60 from H. sapiens, S. cerevisiae and the alphaproteobacterium C. sphaeroides and observed high structural conservation for a C-terminal region ranging from residues 318-540 (S. cerevisiae numbering; Figure 1A ...
In eukaryotes, the essential process of cellular respiration takes place in the cristae of mitochondria. The protein Mic60 is known to stabilize crista junctions; however, how the C-terminal Mitofilin domain of Mic60 mediates cristae-supported respiration remains elusive. Here, we used ancestral sequence reconstruction to generate Mitofilin ancestors up to and including the last opisthokont common ancestor (LOCA). We found that yeast-lineage derived Mitofilin ancestors as far back as the LOCA rescue respiration. By comparing Mitofilin ancestors with different respiratory phenotypes, we identify four residues that explain the difference between respiration functional yeast- and non-functional animal-derived common Mitofilin ancestors. Our results imply that Mitofilin-supported respiration in yeast stems from a conserved mechanism, and provide a foundation for investigating the divergence of candidate crista junction interactions present during the emergence of eukaryotes.
... MIC60 (encoded by the gene Immt) is the largest member of the MICOS complex, and its presence at CJs stabilizes the expression and organization of other MICOS family members (5,8,9,10). Many studies have detailed the disrupted cristae morphology and reduced mitochondrial respiration after MIC60 expression loss using yeast models or mammalian cell culture models via siRNA-mediated knockdown (8,9,11,12,13,14). More recently, structural work has characterized the close interactions between MIC60 subdomains, MIC19, and SAM50, while also assessing their organization at CJs in yeast (15). ...
... As a core component of the MICOS complex, the loss of MIC60 expression would be expected to affect mitochondrial morphology (8,9,11,12,13,14). Using immunohistochemical staining of COX1 (gene name Mtco1), a subunit of complex IV of the electron transport chain (21), we found that mitochondrial staining appeared disorganized in the small intestine, colon, and cecum but was unchanged in the liver or kidney (Figs 2C and S2A). ...
... We also identified significantly increased mitochondrial perimeter, major axis length, minor axis length, and circularity in Immt F/F small intestine and colon, relative to WT tissues (Fig S2B and C). Altogether, these results demonstrate that conditional Immt deletion in vivo recapitulates the effects on MICOS expression and mitochondrial morphology that have been previously reported from in vitro studies (8,9,11,12,13,14). ...
The mitochondrial contact site and cristae organizing system (MICOS) is important for crista junction formation and for maintaining inner mitochondrial membrane architecture. A key component of the MICOS complex is MIC60, which has been well studied in yeast and cell culture models. However, only one recent study has demonstrated the embryonic lethality of losing Immt (the gene encoding MIC60) expression. Tamoxifen-inducible ROSA-CreER T2 –mediated deletion of Immt in adult mice disrupted the MICOS complex, increased mitochondria size, altered cristae morphology, and was lethal within 12 d. Pathologically, these mice displayed defective intestinal muscle function (paralytic ileus) culminating in dehydration. We also identified bone marrow (BM) hypocellularity in Immt -deleted mice, although BM transplants from wild-type mice did not improve survival. Altogether, this inducible mouse model demonstrates the importance of MIC60 in vivo, in both hematopoietic and non-hematopoietic tissues, and provides a valuable resource for future mechanistic investigations into the MICOS complex.
... Both MICOS subcomplexes are necessary for the formation of crista junctions and their physical coupling is largely mediated by Mic12/QIL1 (Guarani et al, 2015;Zerbes et al, 2016). MICOS deficiency causes the loss of crista junctions and the detachment of cristae from the inner boundary membrane, and loss of function mutations in human patients cause severe mitochondrial pathologies, including a fatal encephalopathy (John et al, 2005;Rabl et al, 2009;Mun et al, 2010;Harner et al, 2011;Hoppins et al, 2011;Malsburg et al, 2011;Guarani et al, 2016;Zeharia et al, 2016;Benincá et al, 2021;Peifer-Weiß et al, 2023). ...
The boundary and cristae domains of the mitochondrial inner membrane are connected by crista junctions. Most cristae membrane proteins are nuclear-encoded and inserted by the mitochondrial protein import machinery into the inner boundary membrane. Thus, they must overcome the diffusion barrier imposed by crista junctions to reach their final location. Here, we show that respiratory chain complexes and assembly intermediates are physically connected to the mitochondrial contact site and cristae organizing system (MICOS) that is essential for formation and stability of crista junctions. We identify the inner membrane protein Mar26 (Fmp10) as determinant in the biogenesis of the cytochrome bc 1 complex (complex III). Mar26 couples a Rieske Fe/S protein-containing assembly intermediate to MICOS. Our data indicate that Mar26 maintains an assembly-competent Rip1 pool at crista junctions where complex III maturation likely occurs. MICOS facilitates efficient Rip1 assembly by recruitment of complex III assembly intermediates to crista junctions. We propose that MICOS, via interaction with assembly factors such as Mar26, directly contributes to the spatial and temporal coordination of respiratory chain biogenesis.
... Instead, the loss of cardiolipin, an IMM-specific phospholipid, caused a partial reduction of the IMM diffusion barrier (Fig S4 F). In contrast, the loss of a mitochondrial contact site and the cristae organizing system (MICOS) complex component, Mic60 (Rabl et al., 2009), did not have a large impact on the diffusion pattern of Yta12-GFP (Fig. S4 G), suggesting that the changes observed in the crd1Δ cells are not simply due to the altered cristae structure. These data suggest that the mitochondrial barrier involves distinct molecular mechanisms than the ER diffusion barrier, which is composed of the local accumulation of ceramide in the membrane bilayer at the site of the barrier. ...
Lateral diffusion barriers compartmentalize membranes to generate polarity or asymmetrically partition membrane-associated macromolecules. Budding yeasts assemble such barriers in the endoplasmic reticulum (ER) and the outer nuclear envelope at the bud neck to retain aging factors in the mother cell and generate naïve and rejuvenated daughter cells. However, little is known about whether other organelles are similarly compartmentalized. Here, we show that the membranes of mitochondria are laterally compartmentalized at the bud neck and near the cell poles. The barriers in the inner mitochondrial membrane are constitutive, whereas those in the outer membrane form in response to stresses. The strength of mitochondrial diffusion barriers is regulated positively by spatial cues from the septin axis and negatively by retrograde (RTG) signaling. These data indicate that mitochondria are compartmentalized in a fission-independent manner. We propose that these diffusion barriers promote mitochondrial polarity and contribute to mitochondrial quality control.
... In S. cerevisiae, it was shown that Mic10 has a conserved GxGxGxG motif that is important for homo-oligomerization and, consequently, for inducing membrane curvature (7,14). The formation of cristae and CJs is coordinated by a delicate balance of Mic60 and the assembly state of the F 1 F 0 ATP synthase dimers (106). In S. cerevisiae, Mic60, also known as Formation of CJ protein 1 (Fcj1), was the first protein shown to be required for CJ formation and to be enriched at CJs (106). ...
... The formation of cristae and CJs is coordinated by a delicate balance of Mic60 and the assembly state of the F 1 F 0 ATP synthase dimers (106). In S. cerevisiae, Mic60, also known as Formation of CJ protein 1 (Fcj1), was the first protein shown to be required for CJ formation and to be enriched at CJs (106). Cells lacking Mic60 (Fcj1) showed fragmented mitochondrial morphology with partial loss of mtDNA. ...
... Overall, it is not entirely clear whether, or when, CHCHD2 and CHCHD10 are bona fide subunits of the MICOS complex enriched at CJs (59,121). The first MICOS subunit, Mic60 (Fcj1), was discovered and characterized more than 14 years ago (106), and since then, it has been implicated in a wide range of functions in different organisms. The role of the different MICOS subunits is well documented in physiological processes such as shaping the IM, mitochondrial import pathways, mtDNA maintenance and nucleoid morphology, m maintenance, lipid transport, respiratory chain complex (RCC) stability, and mitophagy (72,78). ...
Mitochondria are essential organelles performing important cellular functions ranging from bioenergetics and metabolism to apoptotic signaling and immune responses. They are highly dynamic at different structural and functional levels. Mitochondria have been shown to constantly undergo fusion and fission processes and dynamically interact with other organelles such as the endoplasmic reticulum, peroxisomes, and lipid droplets. The field of mitochondrial dynamics has evolved hand in hand with technological achievements including advanced fluorescence super-resolution nanoscopy. Dynamic remodeling of the cristae membrane within individual mitochondria, discovered very recently, opens up a further exciting layer of mitochondrial dynamics. In this review, we discuss mitochondrial dynamics at the following levels: ( a) within an individual mitochondrion, ( b) among mitochondria, and ( c) between mitochondria and other organelles. Although the three tiers of mitochondrial dynamics have in the past been classified in a hierarchical manner, they are functionally connected and must act in a coordinated manner to maintain cellular functions and thus prevent various human diseases.
Expected final online publication date for the Annual Review of Biophysics, Volume 53 is May 2024. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... Related to mitochondrial volume, OPA1 expression in fast-twitch fibers was accentuated even more, with levels 10-fold over that found in mitochondria of slowtwitch fibers ( Figure 3A). Likewise, MIC60, a key component in the mitochondrial inner membrane organizing system (MINOS) [30,31], was also expressed in a fiber type-dependent manner with nearly three-fold higher expression in fast-twitch fiber mitochondria ( Figure 3B). Additionally, fast-twitch mitochondria expressed double the ATP-synthase (Complex V; CV) levels of mitochondria in slow- twitch fibers ( Figure 3C). ...
Objective
Human skeletal muscle consists of a mixture of slow- and fast-twitch fibers with distinct capacities for contraction mechanics, fermentation, and oxidative phosphorylation. While the divergence in mitochondrial volume favoring slow-twitch fibers is well established, data on the fiber type-specific intrinsic mitochondrial function and morphology are highly limited with existing data mainly being generated in animal models. This highlights the need for more human data on the topic.
Methods
Here, we utilized THRIFTY, a rapid fiber type identification protocol to detect, sort, and pool fast- and slow-twitch fibers within 6 h of muscle biopsy sampling. Respiration of permeabilized fast- and slow-twitch fiber pools was then analyzed with high-resolution respirometry. Using standardized western blot procedures, muscle fiber pools were subsequently analyzed for control proteins and key proteins related to respiratory capacity.
Results
Maximal complex I+II respiration was 25% higher in human slow-twitch fibers compared to fast-twitch fibers. However, per mitochondrial volume, the respiratory rate of mitochondria in fast-twitch fibers was approximately 50% higher for complex I+II, which was primarily mediated through elevated complex II respiration. Furthermore, the abundance of complex II protein and proteins regulating cristae structure were disproportionally elevated in mitochondria of the fast-twitch fibers. The difference in intrinsic respiratory rate was not reflected in fatty acid–or complex I respiration.
Conclusion
Mitochondria of human fast-twitch muscle fibers compensate for their lack of volume by substantially elevating intrinsic respiratory rate through increased reliance on complex II.
... The molecular determinants of CM are best understood in Saccharomyces cerevisiae, where ATP synthases form ribbon-like rows of dimers, which induce curvature along tubule and lamellar rims (Dudkina et al, 2005;Strauss et al, 2008;Davies et al, 2011;Blum et al, 2019). Loss of the dimerization subunit g, Atp20p, results in monomeric ATP synthases and onion-like mitochondria with flat layers of IMM that run parallel with the OMM (Arnold et al, 1998;Paumard et al, 2002;Arselin et al, 2004;Rabl et al, 2009). Reconstituted ATP synthase dimers spontaneously assemble into rows driven by changes to elastic membrane bending energies (Anselmi et al, 2018) and are sufficient to form tubular liposomes (Blum et al, 2019). ...
Cristae are high‐curvature structures in the inner mitochondrial membrane (IMM) that are crucial for ATP production. While cristae‐shaping proteins have been defined, analogous lipid‐based mechanisms have yet to be elucidated. Here, we combine experimental lipidome dissection with multi‐scale modeling to investigate how lipid interactions dictate IMM morphology and ATP generation. When modulating phospholipid (PL) saturation in engineered yeast strains, we observed a surprisingly abrupt breakpoint in IMM topology driven by a continuous loss of ATP synthase organization at cristae ridges. We found that cardiolipin (CL) specifically buffers the inner mitochondrial membrane against curvature loss, an effect that is independent of ATP synthase dimerization. To explain this interaction, we developed a continuum model for cristae tubule formation that integrates both lipid and protein‐mediated curvatures. This model highlighted a snapthrough instability, which drives IMM collapse upon small changes in membrane properties. We also showed that cardiolipin is essential in low‐oxygen conditions that promote PL saturation. These results demonstrate that the mechanical function of cardiolipin is dependent on the surrounding lipid and protein components of the IMM.
... Multiple studies have characterized disrupted cristae morphology and reduced mitochondrial respiration following MIC60 expression loss (7,8,9,10,11,12); however, these studies were all completed using yeast models or mammalian cell culture models via siRNAmediated knockdown. To date, the in vivo impact of MIC60 loss has not been documented. ...
The mitochondrial contact site and cristae organizing system (MICOS) is important for cristae junctions (CJ) formation and for maintaining inner mitochondrial membrane (IMM) architecture. As the largest member, MIC60 is the primary scaffold protein for this complex. While MIC60 has been well studied in yeast and cell culture models, its function in mammals is poorly understood. To address this, we developed a mouse model conditionally deleting Immt (which encodes MIC60) and found that global Immt deletion disrupted the MICOS complex and resulted in lethality within 9 days of tamoxifen treatment. Pathologically, these mice display intestinal defects consistent with paralytic ileus, resulting in dehydration. We also identified bone marrow hypocellularity in tamoxifen-treated mice. However, bone marrow transplants from ImmtWT mice failed to rescue survival. Altogether, this novel mouse model demonstrates the importance of MIC60 in vivo, in both hematopoietic and non-hematopoietic tissues, and provides a valuable resource for future mechanistic investigations into the MICOS complex. Such investigations could include an in vivo structure-function analysis of MIC60 functional domains, with characterizations that are relevant to human diseases.
... The CM is enriched with respiratory chain complexes, iron-sulfur cluster assembly proteins, and selective channels/transporters while the IBM contains the protein translocation and membrane fusion machinery (Vogel et al., 2006;Suppanz et al., 2009). Cristae junctions are tube-like membrane invaginations of~25 nm diameter that separate IBM and CM (Rabl et al., 2009;Davies et al., 2012;Busch, 2020). The cristae occupy most of the inner membrane surface, indicating their crucial role in cellular physiology. ...
Mitochondria play a critical role in energy metabolism and signal transduction, which is tightly regulated by proteins, metabolites, and ion fluxes. Metabolites and ion homeostasis are mainly mediated by channels and transporters present on mitochondrial membranes. Mitochondria comprise two distinct compartments, the outer mitochondrial membrane (OMM) and the inner mitochondrial membrane (IMM), which have differing permeabilities to ions and metabolites. The OMM is semipermeable due to the presence of non-selective molecular pores, while the IMM is highly selective and impermeable due to the presence of specialized channels and transporters which regulate ion and metabolite fluxes. These channels and transporters are modulated by various post-translational modifications (PTMs), including phosphorylation, oxidative modifications, ions, and metabolites binding, glycosylation, acetylation, and others. Additionally, the mitochondrial protein quality control (MPQC) system plays a crucial role in ensuring efficient molecular flux through the mitochondrial membranes by selectively removing mistargeted or defective proteins. Inefficient functioning of the transporters and channels in mitochondria can disrupt cellular homeostasis, leading to the onset of various pathological conditions. In this review, we provide a comprehensive overview of the current understanding of mitochondrial channels and transporters in terms of their functions, PTMs, and quality control mechanisms.
... MA-5 also improves distended cristae in C. elegans immt-1 single mutants 12 . Mitofilin/Mic60 depletion led to a loss of cristae junctions (CJs) 25 . Intriguingly, MICU-1, one of the components of the MCU complex, was also recently reported to be important not only for Ca 2+ transport but also for maintenance of CJs width 26 . ...
Mitochonic acid-5 ameliorates the pathophysiology of human mitochondrial-disease fibroblasts and Caenorhabditis elegans Duchenne muscular dystrophy and Parkinson's disease models. Here, we found that 10 μM MA-5 attenuates the age-related decline in motor performance, loss of muscle mitochondria, and degeneration of dopaminergic neurons associated with mitochondrial Ca2+ overload in C. elegans. These findings suggest that MA-5 may act as an anti-aging agent against a wide range of neuromuscular dysfunctions in metazoans.
