SEPARATION OF MITOCHONDRIAL MEMBRANES OF <i>NEUROSPORA CRASSA  </i>

Cassady, W. E.; Leiter, Edward H.; Bergquist, Arloa; Wagner, Robert P.

doi:10.1083/jcb.53.1.66

Cited by 29 publications

(5 citation statements)

References 21 publications

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“…Our results also demonstrate that MoIlv6 is localized to the mitochondria and that the conserved domains are not necessary for this localization. Previous studies of fungal Ilv6 proteins have indicated that the precursor proteins maintain a signal peptide targeting ALS to the mitochondria (Cassady et al, 1972;Lee and Duggleby, 2006). The signal peptide is required for mitochondrial localization, and for conidial morphology and infection-related development.…”

Section: Discussionmentioning

confidence: 99%

“…In fungi, the ALS enzyme contains a catalytic subunit of 60-70 kDa and a regulatory subunit that varies in size depending on the species and isoform Duggleby, 1999, 2001). Both the catalytic and regulatory subunits are nuclear encoded and targeted to the mitochondria as a result of the presence of a mitochondrial signal peptide (Cassady et al, 1972;Lee and Duggleby, 2006;Polaina, 1984;Sherman et al, 1974). As no ALS has been identified in animals, inhibitors of ALS, aiming to inhibit the BCAA biosynthetic pathway, have been explored as potential antifungal drug targets (Chipman et al, 1998;Goldstein and McCusker, 2001;McCourt and Duggleby, 2006).…”

Section: Introductionmentioning

confidence: 99%

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Acetolactate synthases MoIlv2 and MoIlv6 are required for infection‐related morphogenesis in Magnaporthe oryzae

Zhang

et al. 2013

Molecular Plant Pathology

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Amino acids are important components in the metabolism of a variety of pathogens, plants and animals. Acetolactate synthase (ALS) catalyses the first common step in leucine, isoleucine and valine biosynthesis, and is the target of several classes of inhibitors. Here, MoIlv2, an orthologue of the Saccharomyces cerevisiae ALS catalytic subunit Ilv2, and MoIlv6, an orthologue of the S. cerevisiae ALS regulatory subunit Ilv6, were identified. To characterize MoILV2 and MoILV6 functions, we generated the deletion mutants ΔMoilv2 and ΔMoilv6. Phenotypic analysis showed that both mutants were auxotrophic for leucine, isoleucine and valine, and were defective in conidial morphogenesis, appressorial penetration and pathogenicity. Further studies suggested that MoIlv2 and MoIlv6 play a critical role in maintaining the balance of intracellular amino acid levels. MoIlv2 and MoIlv6 are both localized to the mitochondria and the signal peptide of MoIlv6 is critical for its localization. In summary, our evidence indicates that MoIlv2 plays a crucial role in isoleucine and valine biosynthesis, whereas MoIlv6 contributes to isoleucine and leucine biosynthesis; both genes are required for fungal pathogenicity. This study indicates the potential of targeting branched-chain amino acid biosynthesis for anti-rice blast management.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Acetolactate synthases MoIlv2 and MoIlv6 are required for infection‐related morphogenesis in Magnaporthe oryzae

Zhang

et al. 2013

Molecular Plant Pathology

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“…Of these, threonine deaminase is apparently cytosolic while most, if not all, of a cell's acetohydroxy acid synthase, acetohydroxy acid isomeroreductase, and dihydroxy acid dehydratase is localized to the mitochondrial matrix (22,80).…”

Section: Neurospora Crassamentioning

confidence: 99%

Leucine Biosynthesis in Fungi: Entering Metabolism through the Back Door

Kohlhaw

2003

Microbiol Mol Biol Rev

221

249

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After exploring evolutionary aspects of branched-chain amino acid biosynthesis, the review focuses on the extended leucine biosynthetic pathway as it operates in Saccharomyces cerevisiae. First, the genes and enzymes specific for the leucine pathway are considered: LEU4 and LEU9 (encoding the α-isopropylmalate synthase isoenzymes), LEU1 (isopropylmalate isomerase), and LEU2 (β-isopropylmalate dehydrogenase). Emphasis is given to the unusual distribution of the branched-chain amino acid pathway enzymes between mitochondrial matrix and cytosol, on the newly defined role of Leu5p, and on regulatory mechanisms governing gene expression and enzyme activity, including new evidence for the metabolic importance of the regulation of α-isopropylmalate synthase by coenzyme A. Next, structure-function relationships of the transcriptional regulator Leu3p are addressed, defining its dual role as activator and repressor and discussing evidence in support of the self-masking model. Recent data pointing at a more extended Leu3p regulon are discussed. An overview of the layered controls of the extended leucine pathway is provided that includes a description of the newly recognized roles of Ilv5p and Bat1p in maintaining mitochondrial integrity. Finally, branched-chain amino acid biosynthesis and its regulation in other fungi are summarized, the question of leucine as metabolic signal is addressed, and possible directions of future research in this area are outlined

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“…In contrast to the bacterial enzyme, the structure and biochemical properties of AHAS from eucaryotes have not been extensively characterized, and in most cases the enzyme has been studied in crude cell extracts only. Fungal AHAS is localized in the mitochondria ( , ) and plant AHAS in the chloroplast (), although the genes may be found in the nuclear or in the organelle genome ( − ). In those cases where it is nuclear-encoded, the enzyme is transported to the appropriate subcellullar compartment.…”

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confidence: 99%

Expression, Purification, Characterization, and Reconstitution of the Large and Small Subunits of Yeast Acetohydroxyacid Synthase

1999

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Acetohydroxyacid synthase (AHAS, EC 4.1.3.18) catalyzes the first step in the biosynthesis of the branched-chain amino acids. In bacteria, the enzyme has a large subunit containing the catalytic machinery and a small subunit with a regulatory role. In eucaryotes, the evidence for a regulatory subunit is largely indirect and circumstantial. We investigated the possibility that the yeast open reading frame YCL009c is an AHAS small subunit. Analysis of the DNA sequence shows that it contains all the appropriate transcription, translation and regulatory signals. YCL009c was shown to be expressed in yeast and the protein localized in mitochondria where it undergoes removal of a transit peptide targeting sequence. This putative small subunit protein (ilv6) and the catalytic subunit of yeast AHAS (ilv2) were each overexpressed in Escherichia coli and purified to near homogeneity. Reconstitution studies showed that the ilv6 protein stimulates the catalytic activity of the ilv2 protein by up to 7-fold (from 6.8 +/- 0.7 to 49.0 +/- 1.8 U/mg) and confers upon it sensitivity to inhibition by valine (Ki = 0.16 +/- 0.02 mM). Valine inhibition is partially reversed by ATP. The reconstitution is favored by high concentrations of potassium phosphate ( approximately 1 M) and at neutral pH. Under optimal conditions for reconstitution, a dissociation constant for the subunits of 70 +/- 7 nM was determined. Valine inhibition is partial, resulting in a specific activity that is similar to that of the ilv2 protein alone. However, measurements of the Km for substrate rule out the possibility that valine inhibition is accomplished by dissociation of the subunits.

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SEPARATION OF MITOCHONDRIAL MEMBRANES OF NEUROSPORA CRASSA

Cited by 29 publications

References 21 publications

Acetolactate synthases MoIlv2 and MoIlv6 are required for infection‐related morphogenesis in Magnaporthe oryzae

Acetolactate synthases MoIlv2 and MoIlv6 are required for infection‐related morphogenesis in Magnaporthe oryzae

Leucine Biosynthesis in Fungi: Entering Metabolism through the Back Door

Expression, Purification, Characterization, and Reconstitution of the Large and Small Subunits of Yeast Acetohydroxyacid Synthase

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