Loss of mpv17 affected early embryonic development via mitochondria dysfunction in zebrafish

AbstractMVP17 encodes a mitochondrial inner-membrane protein, and mutation of human MVP17 can cause mitochondria DNA depletion syndrome (MDDS). However, the underlying function of mpv17 is still elusive. Here, we developed a new mutant with mpv17 knockout by using the CRISPR/Cas9 system. The mpv17−/− zebrafish showed developmental defects in muscles, liver, and energy supply. The mpv17−/− larvae hardly survived beyond a month, and they showed abnormal growth during the development stage. Abnormal swimming ability was also found in the mpv17−/− zebrafish. The transmission electron microscope (TEM) observation indicated that the mpv17−/− zebrafish underwent severe mitochondria dysfunction and the disorder of mitochondrial cristae. As an energy producer, the defects of mitochondria significantly reduced ATP content in mpv17−/− zebrafish, compared to wild-type zebrafish. We hypothesized that the disorder of mitochondria cristae was contributed to the dysfunction of muscle and liver in the mpv17−/− zebrafish. Moreover, the content of major energy depot triglycerides (TAG) was decreased dramatically. Interestingly, after rescued with normal exogenous mitochondria by microinjection, the genes involved in the TAG metabolism pathway were recovered to a normal level. Taken together, this is the first report of developmental defects in muscles, liver, and energy supply via mitochondria dysfunction, and reveals the functional mechanism of mpv17 in zebrafish.

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Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix

Molecular and Cellular Biology ◽

10.1128/mcb.10.5.1873-1881.1990 ◽

1990 ◽

Vol 10 (5) ◽

pp. 1873-1881

Author(s):

S M Glaser ◽

B R Miller ◽

M G Cumsky

Keyword(s):

Amino Acids ◽

Mitochondrial Matrix ◽

Mature Protein ◽

Wild Type ◽

Inner Membrane ◽

Hydrophobic Domain ◽

Mitochondrial Inner Membrane ◽

The Matrix ◽

Inner Membrane Protein ◽

Cognate Protein

We have examined the import and intramitochondrial localization of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. The results of studies on the import of subunit Va derivatives carrying altered presequences suggest that the uptake of this protein is highly efficient. We found that a presequence of only 5 amino acids (Met-Leu-Ser-Leu-Arg) could direct the import and localization of subunit Va with wild-type efficiency, as judged by several different assays. We also found that subunit Va could be effectively targeted to the mitochondrial inner membrane with a heterologous presequence that failed to direct import of its cognate protein. The results presented here confirmed those of an earlier study and showed clearly that the information required to "sort" subunit Va to the inner membrane resides in the mature protein sequence, not within the presequence per se. We present additional evidence that the aforementioned sorting information is contained, at least in part, in a hydrophobic stretch of 22 amino acids residing within the C-terminal third of the protein. Removal of this domain caused subunit Va to be mislocalized to the mitochondrial matrix.

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Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix.

Molecular and Cellular Biology ◽

10.1128/mcb.10.5.1873 ◽

1990 ◽

Vol 10 (5) ◽

pp. 1873-1881 ◽

Cited By ~ 21

Author(s):

S M Glaser ◽

B R Miller ◽

M G Cumsky

Keyword(s):

Amino Acids ◽

Mitochondrial Matrix ◽

Mature Protein ◽

Wild Type ◽

Inner Membrane ◽

Hydrophobic Domain ◽

Mitochondrial Inner Membrane ◽

The Matrix ◽

Inner Membrane Protein ◽

Cognate Protein

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Mutations Affecting a Yeast Mitochondrial Inner Membrane Protein, Pnt1p, Block Export of a Mitochondrially Synthesized Fusion Protein from the Matrix

Molecular and Cellular Biology ◽

10.1128/mcb.19.10.6598 ◽

1999 ◽

Vol 19 (10) ◽

pp. 6598-6607 ◽

Cited By ~ 46

Author(s):

Shichuan He ◽

Thomas D. Fox

Keyword(s):

Membrane Protein ◽

Fusion Protein ◽

Kluyveromyces Lactis ◽

Nuclear Gene ◽

Wild Type ◽

Inner Membrane ◽

Mitochondrial Inner Membrane ◽

Mitochondrial Target ◽

The Matrix ◽

Inner Membrane Protein

ABSTRACT The machinery that inserts mitochondrially encoded proteins into the inner membrane and translocates their hydrophilic domains through the membrane is poorly understood. We have developed a genetic screen for Saccharomyces cerevisiae mutants defective in this export process. The screen is based on the fact that the hydrophilic polypeptide Arg8mp is exported from the matrix if it is synthesized within mitochondria as a bifunctional Cox2p-Arg8mp fusion protein. Since export of Arg8mp causes an Arg− phenotype, defective mutants can be selected as Arg+. Here we show that mutations in the nuclear gene PNT1 block the translocation of mitochondrially encoded fusion proteins across the inner membrane. Pnt1p is a mitochondrial integral inner membrane protein that appears to have two hydrophilic domains in the matrix, flanking a central hydrophobic hairpin-like anchor. While an S. cerevisiae pnt1 deletion mutant was more sensitive to H2O2 than the wild type was, it was respiration competent and able to export wild-type Cox2p. However, deletion of thePNT1 orthologue from Kluyveromyces lactis,KlPNT1, caused a clear nonrespiratory phenotype, absence of cytochrome oxidase activity, and a defect in the assembly of KlCox2p that appears to be due to a block of C-tail export. SincePNT1 was previously described as a gene affecting resistance to the antibiotic pentamidine, our data support a mitochondrial target for this drug.

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Long-term high-fat feeding induces greater fat storage in mice lacking UCP3

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00779.2007 ◽

2008 ◽

Vol 295 (5) ◽

pp. E1018-E1024 ◽

Cited By ~ 46

Author(s):

Sheila R. Costford ◽

Shehla N. Chaudhry ◽

Sean A. Crawford ◽

Mahmoud Salkhordeh ◽

Mary-Ellen Harper

Keyword(s):

Uncoupling Protein ◽

Muscle Metabolism ◽

Whole Body ◽

Wild Type ◽

High Fat ◽

Mitochondrial Inner Membrane ◽

Insulin Tolerance ◽

Glucose Tolerant ◽

Inner Membrane Protein

Uncoupling protein-3 (UCP3) is a mitochondrial inner-membrane protein highly expressed in skeletal muscle. While UCP3's function is still unknown, it has been hypothesized to act as a fatty acid (FA) anion exporter, protecting mitochondria against lipid peroxidation and/or facilitating FA oxidation. The aim of this study was to determine the effects of long-term feeding of a 45% fat diet on whole body indicators of muscle metabolism in congenic C57BL/6 mice that were either lacking UCP3 ( Ucp3−/−) or had a transgenically induced approximately twofold increase in UCP3 levels ( UCP3tg). Mice were fed the high-fat (HF) diet for a period of either 4 or 8 mo immediately following weaning. After long-term HF feeding, UCP3tg mice weighed an average of 15% less than wild-type mice ( P < 0.05) and were 20% less metabolically efficient than both wild-type and Ucp3−/− mice ( P < 0.01). Additionally, wild-type mice had 21% lower, whereas UCP3tg mice had 36% lower, levels of adiposity compared with Ucp3−/− mice ( P < 0.05 and P < 0.001, respectively), indicating a protective effect of UCP3 against fat gain. No differences in whole body oxygen consumption were detected following long-term HF feeding. Glucose and insulin tolerance tests revealed that both the UCP3tg and Ucp3−/− mice were more glucose tolerant and insulin sensitive compared with wild-type mice after short-term HF feeding, but this protection was not maintained in the long term. Findings indicate that UCP3 is involved in protection from fat gain induced by long-term HF feeding, but not in protection from insulin resistance.

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SEC3 Mutations Are Synthetically Lethal With Profilin Mutations and Cause Defects in Diploid-Specific Bud-Site Selection

Genetics ◽

10.1093/genetics/144.2.495 ◽

1996 ◽

Vol 144 (2) ◽

pp. 495-510 ◽

Cited By ~ 3

Author(s):

B K Haarer ◽

A Corbett ◽

Y Kweon ◽

A S Petzold ◽

P Silver ◽

...

Keyword(s):

Site Selection ◽

Secretory Pathway ◽

Wild Type ◽

Synthetic Lethal ◽

Abnormal Growth ◽

Colony Color ◽

Synthetically Lethal ◽

Homozygous Diploid ◽

Bud Site ◽

Wild Type Yeast

Abstract Replacement of the wild-type yeast profilin gene (PFY1) with a mutated form (pfy1-111) that has codon 72 changed to encode glutamate rather than arginine results in defects similar to, but less severe than, those that result from complete deletion of the profilin gene. We have used a colony color-sectoring assay to identify mutations that cause pfy1-111, but not wild-type, cells to be inviable. These profilin synthetic lethal (psl) mutations result in various degrees of abnormal growth, morphology, and temperature sensitivity in PFY1 cells. We have examined psl1 strains in the most detail. Interestingly, these strains display a diploid-specific defect in bud-site selection; haploid strains bud normally, while homozygous diploid strains show a dramatic increase in random budding. We discovered that PSL1 is the late secretory gene, SEC3, and have found that mutations in several other late secretory genes are also synthetically lethal with pfy1-111. Our results are likely to reflect an interdependence between the actin cytoskeleton and secretory processes in directing cell polarity and growth. Moreover, they indicate that the secretory pathway is especially crucial for maintaining budding polarity in diploids.

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Wild-Type Endoderm Abrogates the Ventral Developmental Defects Associated with GATA-4 Deficiency in the Mouse

Developmental Biology ◽

10.1006/dbio.1997.8684 ◽

1997 ◽

Vol 189 (2) ◽

pp. 270-274 ◽

Cited By ~ 137

Author(s):

Naoko Narita ◽

Malgorzata Bielinska ◽

David B. Wilson

Keyword(s):

Wild Type ◽

Developmental Defects

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The integrin coactivator Kindlin-2 plays a critical role in angiogenesis in mice and zebrafish

Blood ◽

10.1182/blood-2010-11-321182 ◽

2011 ◽

Vol 117 (18) ◽

pp. 4978-4987 ◽

Cited By ~ 40

Author(s):

Elzbieta Pluskota ◽

James J. Dowling ◽

Natalie Gordon ◽

Jeffrey A. Golden ◽

Dorota Szpak ◽

...

Keyword(s):

Endothelial Cells ◽

Critical Role ◽

Integrin Αvβ3 ◽

Tube Formation ◽

Basement Membranes ◽

Cytoskeletal Protein ◽

Integrin Expression ◽

Partial Reduction ◽

Wild Type ◽

Developmental Defects

Abstract Kindlin-2, a widely distributed cytoskeletal protein, has been implicated in integrin activation, and its absence is embryonically lethal in mice and causes severe developmental defects in zebrafish. Knockdown of kindlin-2 levels in endothelial cells resulted in defective adhesive and migratory responses, suggesting that angiogenesis might be aberrant even with partial reduction of kindlin-2. This hypothesis has now been tested in the kindlin-2+/− mice. RM1 prostate tumors grown in kindlin-2+/− mice had fewer blood vessels, which were thinner and shorter and supported less tumor growth compared with wild-type littermates. The vessels that did form in the kindlin-2+/− mice lacked smooth muscle cells and pericytes and had thinner basement membranes, indicative of immature vessels. VEGF-induced angiogenesis in matrigel implants was also abnormal in the kindlin-2+/− mice. Vessels in the kindlin-2+/− mice were leaky, and BM transplantation from kindlin-2+/− to WT mice did not correct this defect. Endothelial cells derived from kindlin-2+/− mice had integrin expression levels similar to WT mice but reduced αVβ3-dependent signaling, migration, adhesion, spreading, and tube formation. Developmental angiogenesis was markedly impaired by kindlin-2 morpholinos in zebrafish. Taken together, kindlin-2 plays an important role in pathologic and developmental angiogenesis, which arises from defective activation of integrin αVβ3.

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Characterization of the Role of the FluG Protein in Asexual Development of Aspergillus nidulans

Genetics ◽

10.1093/genetics/158.3.1027 ◽

2001 ◽

Vol 158 (3) ◽

pp. 1027-1036 ◽

Cited By ~ 2

Author(s):

Cletus A D'Souza ◽

Bee Na Lee ◽

Thomas H Adams

Keyword(s):

Aspergillus Nidulans ◽

Negative Influence ◽

Asexual Development ◽

Wild Type ◽

Significant Similarity ◽

Developmental Defects ◽

Asexual Sporulation ◽

Type Strains

Abstract We showed previously that a ΔfluG mutation results in a block in Aspergillus nidulans asexual sporulation and that overexpression of fluG activates sporulation in liquid-submerged culture, a condition that does not normally support sporulation of wild-type strains. Here we demonstrate that the entire N-terminal region of FluG (∼400 amino acids) can be deleted without affecting sporulation, indicating that FluG activity resides in the C-terminal half of the protein, which bears significant similarity with GSI-type glutamine synthetases. While FluG has no apparent role in glutamine biosynthesis, we propose that it has an enzymatic role in sporulation factor production. We also describe the isolation of dominant suppressors of ΔfluG(dsg) that should identify components acting downstream of FluG and thereby define the function of FluG in sporulation. The dsgA1 mutation also suppresses the developmental defects resulting from ΔflbA and dominant activating fadA mutations, which both cause constitutive induction of the mycelial proliferation pathway. However, dsgA1 does not suppress the negative influence of these mutations on production of the aflatoxin precursor, sterigmatocystin, indicating that dsgA1 is specific for asexual development. Taken together, our studies define dsgA as a novel component of the asexual sporulation pathway.

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Differential expression of five tRNA(UAGTrp) amber suppressors in Caenorhabditis elegans.

Molecular and Cellular Biology ◽

10.1128/mcb.8.9.3627 ◽

1988 ◽

Vol 8 (9) ◽

pp. 3627-3635 ◽

Cited By ~ 13

Author(s):

K Kondo ◽

J Hodgkin ◽

R H Waterston

Keyword(s):

Nervous System ◽

Caenorhabditis Elegans ◽

Gene Family ◽

Development Stage ◽

Base Change ◽

Wild Type ◽

Specific Manner ◽

Flanking Sequences ◽

Single Base Change ◽

The Individual

Caenorhabditis elegans has 12 tRNA(UGGTrp) genes as defined by Southern analysis. In order to evaluate the function of the individual members of this multigene family, we sought to recover amber (UAG)-suppressing mutations from reversion experiments with animals carrying amber mutations in a nervous system-affecting gene (unc-13) or a sex-determining gene (tra-3). Revertants were analyzed by Southern blot, exploiting the fact that the CCA to CTA change at the anticodon creates a new XbaI site. Five different members of the tRNATrp gene family were identified as suppressors: sup-7 X, sup-5 III, sup-24 IV, sup-28 X, and sup-29 IV. All five suppressor genes were sequenced and found to encode identical tRNA(UAGTrp) molecules with a single base change (CCA to CTA) at the anticodon compared with their wild-type counterparts. The flanking sequences had only limited homology. The relative expression of these five genes was determined by measuring the efficiencies of suppressers against amber mutations in genes affecting the nervous system, hypodermis, muscle, and sex determination. The results of these cross-suppression tests showed that the five members of the tRNA(Trp) gene family were differentially regulated in a tissue- or development stage-specific manner.

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YME1L controls the accumulation of respiratory chain subunits and is required for apoptotic resistance, cristae morphogenesis, and cell proliferation

Molecular Biology of the Cell ◽

10.1091/mbc.e11-08-0674 ◽

2012 ◽

Vol 23 (6) ◽

pp. 1010-1023 ◽

Cited By ~ 89

Author(s):

Lukas Stiburek ◽

Jana Cesnekova ◽

Olga Kostkova ◽

Daniela Fornuskova ◽

Kamila Vinsova ◽

...

Keyword(s):

Cell Proliferation ◽

Membrane Protein ◽

Respiratory Chain ◽

Integral Membrane Protein ◽

Intermembrane Space ◽

Wild Type ◽

Inner Membrane ◽

Mitochondrial Inner Membrane ◽

Human Orthologue ◽

Excessive Accumulation

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.

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