scholarly journals The absence of Pmp47, a putative yeast peroxisomal transporter, causes a defect in transport and folding of a specific matrix enzyme.

1996 ◽  
Vol 134 (1) ◽  
pp. 37-51 ◽  
Author(s):  
Y Sakai ◽  
A Saiganji ◽  
H Yurimoto ◽  
K Takabe ◽  
H Saiki ◽  
...  
Keyword(s):  
Single Gene ◽  
Alcohol Oxidase ◽  
Candida Boidinii ◽  
High Electron ◽  
Gene Encoding ◽  
Peroxisomal Membrane ◽  
Carboxyl Terminal ◽  
Dihydroxyacetone Synthase ◽  
Physiological And Biochemical ◽  
Solute Transporters

Candida boidinii Pmp47, an integral peroxisomal membrane protein, belongs to a family of mitochondrial solute transporters (e.g., ATP/ADP exchanger), and is the only known peroxisomal member of this family. However, its physiological and biochemical functions have been unrevealed because of the difficulties in the molecular genetics of C. boidinii. In this study, we first isolated the PMP47 gene, which was the single gene encoding for Pmp47 in a gene-engineerable strain S2 of C. boidinii. Sequence analysis revealed that it was very similar to PMP47A and PMP47B genes from a polyploidal C. Boidinii strain (ATCC32195). Next, the PMP47 gene was disrupted and the disruption strain (pmp47delta) was analyzed. Depletion of PMP47 from strain S2 resulted in a retarded growth on oleate and a complete loss of growth on methanol. Both growth substrates require peroxisomal metabolism. EM observations revealed the presence of peroxisomes in methanol- and oleate-induced cells of pmp47delta, but in reduced numbers, and the presence of material of high electron density in the cytoplasm in both cases. Methanol-induced cells of pmp47delta were investigated in detail. The activity of one of the methanol-induced peroxisome matrix enzymes, dihydroxyacetone synthase (DHAS), was not detected in pmp47delta. Further biochemical and immunocytochemical experiments revealed that the DHAS protein aggregated in the cytoplasm as an inclusion body, while two other peroxisome matrix enzymes, alcohol oxidase (AOD) and catalase, were active and found in peroxisomes. Two peroxisome-deficient mutants, strains M6 and M13 (described in previous studies), retained DHAS activity although it was mislocalized to the cytoplasm and the nucleus. We disrupted PMP47 in these peroxisome-deficient mutants. In both strains, M6-pmp47delta and M13-pmp47delta, DHAS was enzymatically active and was located in the cytoplasm and the nucleus. We suggest that an unknown small molecule, which PMP47 transports, is necessary for the folding or the translocation machinery of DHAS within peroxisomes. Pmp47 does not catalyze folding directly because active DHAS is observed in the M6-pmp47delta and M13-pmp47delta strains. Since both AOD and DHAS have the PTS1 motif sequences at their carboxyl terminal, our results first show that depletion of Pmp47 could dissect the peroxisomal import pathway (PTS1 pathway) of these proteins.

scholarly journals Peroxisomal assembly: membrane proliferation precedes the induction of the abundant matrix proteins in the methylotrophic yeast Candida boidinii

1990 ◽  
Vol 96 (4) ◽  
pp. 583-590
Author(s):  
M. Veenhuis ◽  
J.M. Goodman
Keyword(s):  
Early Stage ◽  
Morphological Changes ◽  
Polyclonal Antibodies ◽  
Alcohol Oxidase ◽  
Methylotrophic Yeast ◽  
Candida Boidinii ◽  
Peroxisomal Membrane ◽  
Cell Induction ◽  
The Matrix ◽  
Dihydroxyacetone Synthase

Peroxisomes are massively induced when methylotrophic yeasts are cultured in medium containing methanol. These organelles contain enzymes that catalyze the initial steps of methanol assimilation. In Candida boidinii, a methylotrophic yeast, the peroxisomal matrix (internal compartment) is composed almost exclusively of two proteins, alcohol oxidase and dihydroxyacetone synthase; catalase is present in much lower abundance. Monoclonal and polyclonal antibodies are available against peroxisomal matrix and membrane proteins. These were utilized to correlate the induction of specific proteins with the morphological changes occurring during peroxisomal proliferation. Cells cultured in glucose-containing medium contain two to five small microbodies, which are identifiable by catalase staining and immunoreactivity with a monoclonal antibody against PMP47, an integral peroxisomal membrane protein. Three stages of proliferation can be distinguished when cells are switched to methanol as the carbon source. (1) There is an early stage (within 1 h) in which several peroxisomes develop from a preexisting organelle. This is accompanied by an increase in catalase activity and an induction of PMP47, but no detectable induction of alcohol oxidase or dihydroxyacetone synthase is observed. (2) From 1 to 2.5 h there is further division of these microbodies until up to 30 small peroxisomes generally are present in each of one or two clusters per cell. Induction of alcohol oxidase, dihydroxyacetone synthase and PMP20, a protein that is distributed in the matrix and membrane, is detectable during this time. Serial sections reveal that some peroxisomes remain uninduced while others undergo proliferation. Such sections also show no obvious connections between peroxisomes within clusters.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Regulation and Physiological Role of theDAS1 Gene, Encoding Dihydroxyacetone Synthase, in the Methylotrophic Yeast Candida boidinii

1998 ◽  
Vol 180 (22) ◽  
pp. 5885-5890 ◽  
Author(s):  
Yasuyoshi Sakai ◽  
Tomoyuki Nakagawa ◽  
Masayuki Shimase ◽  
Nobuo Kato
Keyword(s):  
Formate Dehydrogenase ◽  
Physiological Role ◽  
Nitrogen Sources ◽  
Host Genome ◽  
Candida Boidinii ◽  
Carbon And Nitrogen Sources ◽  
Gene Encoding ◽  
Carbon And Nitrogen ◽  
Dihydroxyacetone Synthase

ABSTRACT The physiological role of dihydroxyacetone synthase (DHAS) inCandida boidinii was evaluated at the molecular level. TheDAS1 gene, encoding DHAS, was cloned from the host genome, and regulation of its expression by various carbon and nitrogen sources was analyzed. Western and Northern analyses revealed thatDAS1 expression was regulated mainly at the mRNA level. The regulatory pattern of DHAS was similar to that of alcohol oxidase but distinct from that of two other enzymes in the formaldehyde dissimilation pathway, glutathione-dependent formaldehyde dehydrogenase and formate dehydrogenase. The DAS1 gene was disrupted in one step in the host genome (das1Δ strain), and the growth of the das1Δ strain in various carbon and nitrogen sources was compared with that of the wild-type strain. Thedas1Δ strain had completely lost the ability to grow on methanol, while the strain with a disruption of the formate dehydrogenase gene could survive (Y. Sakai et al., J. Bacteriol. 179:4480–4485, 1997). These and other experiments (e.g., those to determine the expression of the gene and the growth ability of thedas1Δ strain on media containing methylamine or choline as a nitrogen source) suggested that DAS1 is involved in assimilation rather than dissimilation or detoxification of formaldehyde in the cells.

Alcohol oxidase and dihydroxyacetone synthase, the abundant peroxisomal proteins of methylotrophic yeasts, assemble in different cellular compartments

2001 ◽  
Vol 114 (15) ◽  
pp. 2863-2868
Author(s):  
Mary Q. Stewart ◽  
Renee D. Esposito ◽  
Jehangir Gowani ◽  
Joel M. Goodman
Keyword(s):  
Alcohol Oxidase ◽  
Protein Assembly ◽  
Matrix Proteins ◽  
Thiamine Pyrophosphate ◽  
Model Organisms ◽  
Candida Boidinii ◽  
Methylotrophic Yeasts ◽  
The Matrix ◽  
Cellular Compartments ◽  
Dihydroxyacetone Synthase

Alcohol oxidase (AO) and dihydroxyacetone synthase (DHAS) constitute the bulk of matrix proteins in methylotrophic yeasts, model organisms for the study of peroxisomal assembly. Both are homooligomers; AO is a flavin-containing octamer, whereas DHAS is a thiamine pyrophosphate-containing dimer. Experiments in recent years have demonstrated that assembly of peroxisomal oligomers can occur before import; indeed the absence of chaperones within the peroxisomal matrix calls into question the ability of this compartment to assemble proteins at all. We have taken a direct pulse-chase approach to monitor import and assembly of the two major proteins of peroxisomes in Candida boidinii. Oligomers of AO are not observed in the cytosol, consistent with the proteins inability to undergo piggyback import. Indeed, oligomerization of AO can be followed within the peroxisomal matrix, directly demonstrating the capacity of this compartment for protein assembly. By contrast, DHAS quickly dimerizes in the cytosol before import. Binding and import was slowed at 15°C; the effect on AO was more dramatic. In conclusion, our data indicate that peroxisomes assemble AO in the matrix, while DHAS undergoes dimerization prior to import.

scholarly journals A Methylotrophic Pathway Participates in Pectin Utilization by Candida boidinii

2000 ◽  
Vol 66 (10) ◽  
pp. 4253-4257 ◽  
Author(s):  
Tomoyuki Nakagawa ◽  
Tatsuro Miyaji ◽  
Hiroya Yurimoto ◽  
Yasuyoshi Sakai ◽  
Nobuo Kato ◽  
...  
Keyword(s):  
Methyl Ester ◽  
Alcohol Oxidase ◽  
Pectate Lyase ◽  
Methylotrophic Yeast ◽  
Pectin Methylesterase ◽  
Candida Boidinii ◽  
Methanol Metabolism ◽  
Genes Encoding ◽  
Dihydroxyacetone Synthase ◽  
In Cells

ABSTRACT The methylotrophic yeast Candida boidinii S2 was found to be able to grow on pectin or polygalacturonate as a carbon source. When cells were grown on 1% (wt/vol) pectin, C. boidinii exhibited induced levels of the pectin-depolymerizing enzymes pectin methylesterase (208 mU/mg of protein), pectin lyase (673 mU/mg), pectate lyase (673 mU/mg), and polygalacturonase (3.45 U/mg) and two methanol-metabolizing peroxisomal enzymes, alcohol oxidase (0.26 U/mg) and dihydroxyacetone synthase (94 mU/mg). The numbers of peroxisomes also increased ca. two- to threefold in cells grown on these pectic compounds (3.34 and 2.76 peroxisomes/cell for cells grown on pectin and polygalacturonate, respectively) compared to the numbers in cells grown on glucose (1.29 peroxisomes/cell). The cell density obtained with pectin increased as the degree of methyl esterification of pectic compounds increased, and it decreased in strains from which genes encoding alcohol oxidase and dihydroxyacetone synthase were deleted and in a peroxisome assembly mutant. Our study showed that methanol metabolism and peroxisome assembly play important roles in the degradation of pectin, especially in the utilization of its methyl ester moieties.

Subfunctionalization of Duplicate mitf Genes Associated With Differential Degeneration of Alternative Exons in Fish

Genetics ◽  
2002 ◽  
Vol 161 (1) ◽  
pp. 259-267 ◽  
Author(s):  
Joachim Altschmied ◽  
Jacqueline Delfgaauw ◽  
Brigitta Wilde ◽  
Jutta Duschl ◽  
Laurence Bouneau ◽  
...  
Keyword(s):  
Danio Rerio ◽  
Evolutionary History ◽  
Single Gene ◽  
Regulatory Elements ◽  
Fugu Rubripes ◽  
Gene Encoding ◽  
Tetraodon Nigroviridis ◽  
Mammalian Lineage ◽  
History Of ◽  
Fish Proteins

Abstract The microphthalmia-associated transcription factor (MITF) exists in at least four isoforms. These are generated in higher vertebrates using alternative 5′ exons and promoters from a single gene. Two separate genes (mitf-m and mitf-b), however, are present in different teleost fish species including the poeciliid Xiphophorus, the pufferfishes Fugu rubripes and Tetraodon nigroviridis, and the zebrafish Danio rerio. Fish proteins MITF-m and MITF-b correspond at both the structural and the expression levels to one particular bird/mammalian MITF isoform. In the teleost lineage subfunctionalization of mitf genes after duplication at least 100 million years ago is associated with the degeneration of alternative exons and, probably, regulatory elements and promoters. For example, a remnant of the first exon specific for MITF-m is detected within the pufferfish gene encoding MITF-b. Retracing the evolutionary history of mitf genes in vertebrates uncovered the differential recruitment of new introns specific for either the teleost or the bird/mammalian lineage.

scholarly journals Enlarged Peroxisomes Are Present in Oleic Acid–grown Yarrowia lipolytica Overexpressing the PEX16 Gene Encoding an Intraperoxisomal Peripheral Membrane Peroxin

1997 ◽  
Vol 137 (6) ◽  
pp. 1265-1278 ◽  
Author(s):  
Gary A. Eitzen ◽  
Rachel K. Szilard ◽  
Richard A. Rachubinski
Keyword(s):  
Oleic Acid ◽  
Yarrowia Lipolytica ◽  
Consensus Sequence ◽  
Wild Type ◽  
Gene Encoding ◽  
Peroxisomal Membrane ◽  
Peripheral Protein ◽  
The Matrix ◽  
Peroxisomal Targeting ◽  
Targeting Signal

Pex mutants of the yeast Yarrowia lipolytica are defective in peroxisome assembly. The mutant strain pex16-1 lacks morphologically recognizable peroxisomes. Most peroxisomal proteins are mislocalized to a subcellular fraction enriched for cytosol in pex16 strains, but a subset of peroxisomal proteins is localized at, or near, wild-type levels to a fraction typically enriched for peroxisomes. The PEX16 gene was isolated by functional complementation of the pex16-1 strain and encodes a protein, Pex16p, of 391 amino acids (44,479 D). Pex16p has no known homologues. Pex16p is a peripheral protein located at the matrix face of the peroxisomal membrane. Substitution of the carboxylterminal tripeptide Ser-Thr-Leu, which is similar to the consensus sequence of peroxisomal targeting signal 1, does not affect targeting of Pex16p to peroxisomes. Pex16p is synthesized in wild-type cells grown in glucose-containing media, and its levels are modestly increased by growth of cells in oleic acid–containing medium. Overexpression of the PEX16 gene in oleic acid– grown Y. lipolytica leads to the appearance of a small number of enlarged peroxisomes, which contain the normal complement of peroxisomal proteins at levels approaching those of wild-type peroxisomes.

Structural diseases of the heart: syndromes affecting the cardiovascular system

ESC CardioMed ◽  
2018 ◽  
pp. 719-722
Author(s):  
Sabine Klaassen
Keyword(s):  
Single Gene ◽  
Significant Proportion ◽  
Signal Transduction Pathway ◽  
Alagille Syndrome ◽  
Mitogen Activated Protein Kinase ◽  
Neurological Deficits ◽  
Deletion Syndrome ◽  
Gene Encoding ◽  
Mitogen Activated Protein ◽  
Craniofacial Features

Congenital heart disease (CHD) occurs in association with extracardiac anomalies or as part of an identified syndrome in 25–40% of cases. Approximately 30% of children with a chromosomal abnormality have CHD. Aneuploidy, or abnormal chromosomal number, accounts for a significant proportion of CHD. Of individuals born with trisomy 21, 50% have CHD, the most common being an atrioventricular septal defect (45%). In segmental aneuploidies, the so-called microdeletion syndromes, small submicroscopic chromosomal deletions can lead to CHD. The 22q11 deletion syndrome causes CHD with thymic and parathyroid hypoplasia (DiGeorge syndrome) and characteristic dysmorphic craniofacial features due to abnormal pharyngeal arch development. Williams–Beuren syndrome with renovascular anomalies, typical elfin facies, and neurological deficits, is characterized by cardiac involvement in the form of supravalvar aortic and peripheral pulmonic stenosis. Chromosome 1p36 deletion syndrome is the most common subterminal deletion syndrome. A substantial proportion of individuals with 1p36 deletion syndrome have CHD which may occur in the presence or absence of cardiomyopathy, most commonly left ventricular non-compaction cardiomyopathy. Single gene mutations may also cause syndromic CHD. Noonan syndrome and related disorders (‘RASopathies’) are caused by dominant gain-of-function mutations in one of the genes which encode proteins that function in the Ras/mitogen-activated protein kinase (RAS-MAPK) signal transduction pathway. Holt–Oram syndrome is associated with mutations in the transcription factor TBX5. Alagille syndrome is caused by mutations in JAG1, a gene encoding a ligand in the Notch signaling pathway. Heterotaxy syndrome, which means randomization of cardiac, pulmonary, or gastrointestinal situs, is frequently associated with CHD.

scholarly journals Mls is not a single gene, allelic system. Different stimulatory Mls determinants are the products of at least two nonallelic, unlinked genes.

1987 ◽  
Vol 166 (4) ◽  
pp. 1150-1155 ◽  
Author(s):  
R Abe ◽  
J J Ryan ◽  
R J Hodes
Keyword(s):  
T Cells ◽  
Single Gene ◽  
Surface Structures ◽  
Chromosome 1 ◽  
Gene Products ◽  
Spleen Cells ◽  
Gene Encoding ◽  
Cell Surface Structures ◽  
Genes Encoding ◽  
Polymorphic Cell

Mls determinants share with MHC products the unique property of stimulating T cells at extraordinarily high precursor frequencies. The Mls system was originally described as a single locus on chromosome 1, with four alleles, Mlsa, Mlsb, Mlsc, and Mlsd, that encode polymorphic cell surface structures. However, the fundamental issues of polymorphism and allelism in the Mls system remain controversial. To clarify these questions, a formal segregation analysis of the genes encoding Mlsa and Mlsc determinants was carried out by testing the capacity of spleen cells from progeny of (Mlsa X Mlsc)F1 X Mlsb breedings to stimulate responses by unprimed T cells and by Mlsa- and Mlsc-specific cloned T cells. The results of this analysis indicated that the gene encoding Mlsa determinants is neither allelic to nor linked to the gene encoding Mlsc determinants. Together with previous findings, these results also suggest that another strongly stimulatory type, Mlsd, in fact results from the independent expression of unlinked Mlsa and Mlsc gene products. Based on these observations, it is concluded that, contrary to conventional concepts, the stimulatory phenotypes designated as Mlsa, Mlsc, and Mlsd can be accounted for by the independent expression of the products of at least two unlinked gene loci.

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