OPA1 processing controls mitochondrial fusion and is regulated by mRNA splicing, membrane potential, and Yme1L

OPA1, a dynamin-related guanosine triphosphatase mutated in dominant optic atrophy, is required for the fusion of mitochondria. Proteolytic cleavage by the mitochondrial processing peptidase generates long isoforms from eight messenger RNA (mRNA) splice forms, whereas further cleavages at protease sites S1 and S2 generate short forms. Using OPA1-null cells, we developed a cellular system to study how individual OPA1 splice forms function in mitochondrial fusion. Only mRNA splice forms that generate a long isoform in addition to one or more short isoforms support substantial mitochondrial fusion activity. On their own, long and short OPA1 isoforms have little activity, but, when coexpressed, they functionally complement each other. Loss of mitochondrial membrane potential destabilizes the long isoforms and enhances the cleavage of OPA1 at S1 but not S2. Cleavage at S2 is regulated by the i-AAA protease Yme1L. Our results suggest that mammalian cells have multiple pathways to control mitochondrial fusion through regulation of the spectrum of OPA1 isoforms.

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Identification of new OPA1 cleavage site reveals that short isoforms regulate mitochondrial fusion

Molecular Biology of the Cell ◽

10.1091/mbc.e20-09-0605 ◽

2020 ◽

pp. mbc.E20-09-0605

Author(s):

Ruohan Wang ◽

Prashant Mishra ◽

Spiros D. Garbis ◽

Annie Moradian ◽

Michael J. Sweredoski ◽

...

Keyword(s):

Respiratory Chain ◽

Cleavage Site ◽

Mitochondrial Fusion ◽

Mitochondrial Morphology ◽

Cleavage Sites ◽

Mrna Splice ◽

Short Forms ◽

Highly Sensitive ◽

Dependent Site ◽

Splice Forms

OPA1, a large GTPase of the dynamin superfamily, mediates fusion of the mitochondrial inner membranes, regulates cristae morphology, and maintains respiratory chain function. Inner-membrane-anchored long forms of OPA1 (l-OPA1) are proteolytically processed by the OMA1 or YME1L proteases, acting at cleavage sites S1 and S2 respectively, to produce short forms (s-OPA1). In both mouse and human, half of the mRNA splice forms of Opa1 are constitutively processed to yield exclusively s-OPA1. However, the function of s-OPA1 in mitochondrial fusion has been debated, because in some stress conditions, s-OPA1 is dispensable for fusion. By constructing cells in which the Opa1 locus no longer produces transcripts with S2 cleavage sites, we generated a simplified system to identify the new YME1L-dependent site S3 that mediates constitutive and complete cleavage of OPA1. We show that mitochondrial morphology is highly sensitive to the ratio of l-OPA1 to s-OPA1, indicating that s-OPA1 regulates mitochondrial fusion.

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DEGRADATION OF MESSENGER RNA IN MAMMALIAN CELLS

Acta Endocrinologica ◽

10.1530/acta.0.071s369 ◽

1972 ◽

Vol 71 (2_Suppla) ◽

pp. S369-S380 ◽

Cited By ~ 7

Author(s):

Francis T. Kenney ◽

Kai-Lin Lee ◽

Charles D. Stiles

Keyword(s):

Messenger Rna ◽

Mammalian Cells ◽

Messenger Rnas ◽

Tyrosine Transaminase

ABSTRACT Analyses of the response of hydrocortisone-induced tyrosine transaminase in cultured H-35 cells to inhibitors of translation (cycloheximide, puromycin) suggest: (1) that bound ribosomes stabilize messenger RNA in vivo; (2) that messenger is degraded at a rate determined by the rate of translation. Since specific messenger RNAs of mammalian cells are degraded at quite different rates, there may be extensive heterogeneity either in the rate at which ribosomes traverse different messengers or in the number of ribosomes which translate specific messenger RNAs.

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Faculty Opinions recommendation of Cotranscriptional coupling of splicing factor recruitment and precursor messenger RNA splicing in mammalian cells.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1040212.489144 ◽

2006 ◽

Author(s):

Rozanne Sandri-Goldin

Keyword(s):

Rna Splicing ◽

Messenger Rna ◽

Mammalian Cells ◽

Splicing Factor

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Maturation of Mitochondrially Targeted Prx V Involves a Second Cleavage by Mitochondrial Intermediate Peptidase That Is Sensitive to Inhibition by H2O2

Antioxidants ◽

10.3390/antiox10030346 ◽

2021 ◽

Vol 10 (3) ◽

pp. 346

Author(s):

Juhyun Sim ◽

Jiyoung Park ◽

Hyun Ae Woo ◽

Sue Goo Rhee

Keyword(s):

Short Form ◽

Mitochondrial Matrix ◽

Mitochondrial Processing Peptidase ◽

Mouse Tissues ◽

N Terminus ◽

Mitochondrial Intermediate Peptidase ◽

Subsequent Degradation ◽

Processing Peptidase ◽

Mitochondria Targeting ◽

Over Time

Prx V mRNA contains two in-frame AUG codons, producing a long (L-Prx V) and short form of Prx V (S-Prx V), and mouse L-Prx V is expressed as a precursor protein containing a 49-amino acid N-terminal mitochondria targeting sequence. Here, we show that the N-terminal 41-residue sequence of L-Prx V is cleaved by mitochondrial processing peptidase (MPP) in the mitochondrial matrix to produce an intermediate Prx V (I-Prx V) with a destabilizing phenylalanine at its N-terminus, and further, that the next 8-residue sequence is cleaved by mitochondrial intermediate peptidase (MIP) to convert I-Prx V to a stabilized mature form that is identical to S-Prx V. Further, we show that when mitochondrial H2O2 levels are increased in HeLa cells using rotenone, in several mouse tissues by deleting Prx III, and in the adrenal gland by deleting Srx or by exposing mice to immobilized stress, I-Prx V accumulates transiently and mature S-Prx V levels decrease in mitochondria over time. These findings support the view that MIP is inhibited by H2O2, resulting in the accumulation and subsequent degradation of I-Prx V, identifying a role for redox mediated regulation of Prx V proteolytic maturation and expression in mitochondria.

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Review of T-2307, an Investigational Agent That Causes Collapse of Fungal Mitochondrial Membrane Potential

Journal of Fungi ◽

10.3390/jof7020130 ◽

2021 ◽

Vol 7 (2) ◽

pp. 130

Author(s):

Nathan P. Wiederhold

Keyword(s):

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Candida Species ◽

Mammalian Cells ◽

Mitochondrial Membrane ◽

Fungal Infections ◽

Published Data ◽

Candida Auris

Invasive infections caused by Candida that are resistant to clinically available antifungals are of increasing concern. Increasing rates of fluconazole resistance in non-albicans Candida species have been documented in multiple countries on several continents. This situation has been further exacerbated over the last several years by Candida auris, as isolates of this emerging pathogen that are often resistant to multiple antifungals. T-2307 is an aromatic diamidine currently in development for the treatment of invasive fungal infections. This agent has been shown to selectively cause the collapse of the mitochondrial membrane potential in yeasts when compared to mammalian cells. In vitro activity has been demonstrated against Candida species, including C. albicans, C. glabrata, and C. auris strains, which are resistant to azole and echinocandin antifungals. Activity has also been reported against Cryptococcus species, and this has translated into in vivo efficacy in experimental models of invasive candidiasis and cryptococcosis. However, little is known regarding the clinical efficacy and safety of this agent, as published data from studies involving humans are not currently available.

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Trypanosoma brucei has a canonical mitochondrial processing peptidase

Molecular and Biochemical Parasitology ◽

10.1016/j.molbiopara.2012.07.005 ◽

2012 ◽

Vol 185 (2) ◽

pp. 161-164 ◽

Cited By ~ 12

Author(s):

Silvia Desy ◽

André Schneider ◽

Jan Mani

Keyword(s):

Trypanosoma Brucei ◽

Mitochondrial Processing Peptidase ◽

Processing Peptidase

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A Functional Interaction between the Carboxy-Terminal Domain of RNA Polymerase II and Pre-mRNA Splicing

The Journal of Cell Biology ◽

10.1083/jcb.136.1.5 ◽

1997 ◽

Vol 136 (1) ◽

pp. 5-18 ◽

Cited By ~ 91

Author(s):

Lei Du ◽

Stephen L. Warren

Keyword(s):

Rna Polymerase Ii ◽

Mammalian Cells ◽

Mrna Splicing ◽

Splicing Factors ◽

Functional Interaction ◽

Carboxy Terminal Domain ◽

Localization Signal ◽

Nuclear Domains ◽

Carboxy Terminal

In the preceding study we found that Sm snRNPs and SerArg (SR) family proteins co-immunoprecipitate with Pol II molecules containing a hyperphosphorylated CTD (Kim et al., 1997). The association between Pol IIo and splicing factors is maintained in the absence of pre-mRNA, and the polymerase need not be transcriptionally engaged (Kim et al., 1997). The latter findings led us to hypothesize that a phosphorylated form of the CTD interacts with pre-mRNA splicing components in vivo. To test this idea, a nested set of CTD-derived proteins was assayed for the ability to alter the nuclear distribution of splicing factors, and to interfere with splicing in vivo. Proteins containing heptapeptides 1-52 (CTD52), 1-32 (CTD32), 1-26 (CTD26), 1-13 (CTD13), 1-6 (CTD6), 1-3 (CTD3), or 1 (CTD1) were expressed in mammalian cells. The CTD-derived proteins become phosphorylated in vivo, and accumulate in the nucleus even though they lack a conventional nuclear localization signal. CTD52 induces a selective reorganization of splicing factors from discrete nuclear domains to the diffuse nucleoplasm, and significantly, it blocks the accumulation of spliced, but not unspliced, human β-globin transcripts. The extent of splicing factor disruption, and the degree of inhibition of splicing, are proportional to the number of heptapeptides added to the protein. The above results indicate a functional interaction between Pol II's CTD and pre-mRNA splicing.

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Characterization of the mitochondrial processing peptidase of Neurospora crassa.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(17)37639-1 ◽

1994 ◽

Vol 269 (7) ◽

pp. 4959-4967 ◽

Cited By ~ 1

Author(s):

M. Arretz ◽

H. Schneider ◽

B. Guiard ◽

M. Brunner ◽

W. Neupert

Keyword(s):

Neurospora Crassa ◽

Mitochondrial Processing Peptidase ◽

Processing Peptidase

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The effects of D- and L-threo-chloramphenicol on the early development of the chick embryo

Development ◽

10.1242/dev.13.3.341 ◽

1965 ◽

Vol 13 (3) ◽

pp. 341-356

Author(s):

F. S. Billett ◽

Rosalba Collini ◽

Louie Hamilton

Keyword(s):

Protein Synthesis ◽

Amino Acid ◽

Chick Embryo ◽

Messenger Rna ◽

Mammalian Cells ◽

Animal Cells ◽

Inhibit Protein Synthesis ◽

Acid Complex ◽

Amino Acid Complex ◽

Bacterial Systems

In many bacterial systems chloramphenicol has been shown to inhibit protein synthesis (Hahn & Wisseman, 1951; Gale & Folkes, 1953). The precise mechanism of this inhibition is not clear, although the evidence suggests that the interaction of the soluble RNA-amino acid complex with the ribosomes is prevented because the attachment of the messenger RNA to the ribosomes is itself impaired (Lacks & Gros, 1959; Nathans & Lipman, 1961; Jardetsky & Julian, 1964; Julian & Jardetsky, 1964). In contrast to its effect on bacterial systems, chloramphenicol has been reported to have little or no action on the protein synthesis by cell-free extracts of mammalian cells (Rendi, 1959; Ehrenstein & Lipmann, 1961). A basis for this resistance has been proposed by Vazquez (1964), who finds that whereas bacterial ribosomes bind chloramphenicol, ribosomes from other organisms do not. Nevertheless, it cannot be stated with any confidence that chloramphenicol has no effect on the protein synthesis of animal cells.

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