VII. Yeast sequencing reports. The sequence of a 27 kb segment on the right arm of chromosome VII from Saccharomyces cerevisiae reveals MOL1, NAT2, RPL30B,RSR1, CYS4, PEM1/CHO2, NSR1 genes and ten new open reading frames

The DNA sequence of a 26 677 bp fragment from the right arm of chromosome VII from Saccharomyces cerevisiae reveals 18 open reading frames (ORFs) longer than 300 bp. Eight ORFs correspond to previously characterized genes. G6620 is the 3′ end of the MOL1 gene coding for a polypeptide similar to stress-inducible proteins from Fusarium; G6630 is the NAT2 gene which encodes a methionine N-acetyltransferase; G6635 is the RPL30B gene coding for the ribosomal protein L30; G6658 is RSR1 encoding a ras-related protein; G6667 is CYS4, the gene for cystathionine β-synthase; G6670 is identical to ORF2 located close to CYS4; G6673 is PEM1/CHO2 encoding a phosphatidylethanolamine methyltransferase; G7001 is the NSR1 gene coding for a nuclear signal recognition protein. G6664 shares significant homology with the ORF YKR076w from chromosome XI. The other nine ORFs show no significant homology to any protein sequence presently available in the public data bases. The sequence has been deposited in the EMBL data library under Accession Number X85807.     

XIV. Yeast sequencing reports. A 43·5 kb segment of yeast chromosome XIV,which contains MFA2, MEP2, CAP/SRV2, NAM9, FKB1/FPR1/RBP1, MOM22 and CPT1, predicts an adenosine deaminase gene and 14 new open reading frames

A 43,481 bp fragment from the left arm of chromosome XIV of Saccharomyces cerevisiae was sequenced. A gene for tRNA^phe and 23 non-overlapping open reading frames (ORFs) were identified, seven of which correspond to known yeast genes: MFA2, MEP2, CAP/SRV2, NAM9, FKB1/FPR1/RBP1, MOM22 and CPT1. One ORF may correspond to the yet unindentified yeast adenosine deaminase gene. Among the 15 other ORFs, four exhibit known signatures, which include a protein tyrosine phosphatase, a cytoskeleton-associated protein and two ATP-binding proteins, four have similarities with putative proteins of yeast or proteins from other organisms and seven exibit no significant similarity with amino acid sequences described in data banks. One ORF is identical to yeast expressed sequence tags (EST) and therefore corresponds to an expressed gene. Six ORFs present similarities to human dbESTs, thus identifying motifs conserved during evolution. Nine ORFs are putative transmembrane proteins. In addition, one overlapping and three antisense ORFs, which are not likely to be functional, were detected. The sequence has been deposited in the EMBL data bank under Accession Number Z46843.     

Molecular cloning of CaYRB1, the Candida albicans RanBP1/YRB1 homologue.

The yeast Ran binding protein 1 (Yrb1p) is a small protein of 23 kDa that is highly conserved among eukaryotes. It stimulates the GTPase activity of Gsp1p in the presence of the GTPase activating protein Rna1p. In addition to its role in nucleocytoplasmic transport of macromolecules, YRB1/RanBP1 could be involved in the regulation of microtubules structure and dynamics. Since microtubules are tightly associated with morphological changes, we have been interested to study the role and function of YRB1 in the pathogenic fungus Candida albicans, where there is regulated change in cellular morphology. The gene product of CaYRB1 encodes a 212 amino acid protein displaying 73% homology to the S. cerevisiae homologue. The bacterially expressed gene product has an apparent molecular weight of 35.7 kDa. We show that it can complement a S. cerevisiae yrb1 null mutant and that its mRNA does not appear to be regulated in response to conditions inducing morphological changes in C. albicans.     

VII. Yeast sequencing reports. The sequence of an 11·1 kb fragment on the left arm of Saccharomyces cerevisiae chromosome VII reveals six open reading frames including NSP49, KEM1 and four putative new genes

We report the sequence of an 11·1 kb fragment located on the left arm of chromosome VII of Saccharomyces cerevisiae. By sequence analysis we have detected six open reading frames (ORFs) longer than 300 bp, which cover 87% of the entire sequence. ORF G1645 is 100% identical to the KEM1 gene, also identified as DST2, XRN1, SEP1 and RAR5, while G1648 is 100% identical to the NSP49 or NUP49 gene. ORF G1642 shares some identity with a hypothetical protein of Caenorhabditis elegans, while the other four ORFs show no significant homology to known proteins. The sequence has been submitted to the EMBL data library under Accession Number X84705.     

ERG1, encoding squalene epoxidase,is located on the right arm of chromosome VII of Saccharomyces cerevisiae

The ERG1 gene of Saccharomyces cerevisiae encodes squalene epoxidase, a key enzyme in the ergosterol pathway. ERG1 is an essential gene. Disruption of the gene with URA3 results in a lethal phenotype when cells are grown under aerobic conditions, even in the presence of ergosterol. However, cells are viable in the presence of ergosterol under anaerobic growth conditions during which ergosterol is taken up by cells. Physical and genetic mapping data reveal that ERG1 is located on the right arm of chromosome VII proximal to QCR9 at a distance of 14·6 cM from ADE3.     

Sequencing Analysis of a 36·8 kb Fragment of Yeast Chromosome XV Reveals 26 Open Reading Frames Including SEC63, CDC31, SUG2, GCD1, RBL2, PNT1, PAC1 and VPH1

The complete sequence of a 36 775 bp DNA segment located on the right arm of chromosome XV of Saccharomyces cerevisiae has been determined and analysed. The sequence encodes 26 open reading frames of at least 100 amino acids. Eight of these correspond to known genes, whereas 18 correspond to new genes. The sequence has been deposited in the EMBL databank under Accession Numbers: Z75154, Z75155, Z75156, Z75157, Z75158, Z75159, Z75160, Z75161, Z75162, Z75163, Z75164, Z75165, Z75166, Z75167, Z75168, Z75169, Z75170, Z75171, Z75172, Z75173, Z75174, Z75175, Z75176, Z75177, Z75178, Z75179. © 1997 John Wiley & Sons, Ltd.     

RHO gene products, putative small GTP-binding proteins, are important for activation of the CAL1/CDC43 gene product, a protein geranylgeranyltransferase in Saccharomyces cerevisiae.

Two multicopy suppressors of the cal1-1 mutation in the yeast Saccharomyces cerevisiae have been isolated and characterized. They are identical to the yeast RHO1 and RHO2 genes, which encode putative small GTP-binding proteins. Multiple copies of either RHO gene suppressed temperature-sensitive growth of the cal1-1 mutant but did not suppress the cal1 null mutant. Genetic analysis suggests that overproduction of either RHO gene product acts for activation of the CAL1 gene product.     

IIX. Yeast mapping reports. ATP1 and ATP2, the F1F0-ATPase α and β subunit genes of Saccharomyces cerevisiae,are respectively located on chromosomes II and X

Southern blot analysis showed that ATP1 and ATP2 map on chromosomes II and X, respectively. Physical mapping of ATP1 and ATP2 by chromosome fragmentation showed that ATP1 is at the left end of chromosome II and ATP2 is at the right end of chromosome X. Both are located close to telomere sequences of each chromosome; ATP1 and ATP2 being approximately 30 kb and 85 kb from the respective telomeres.     

The Sequence of a 36·7 kb Segment on the Left Arm of Chromosome IV from Saccharomyces cerevisiae Reveals 20 Non-overlapping Open Reading Frames (ORFs) Including SIT4, FAD1, NAM1, RNA11, SIR2, NAT1, PRP9, ACT2 and MPS1 and 11 New ORFs

A 36 688 bp fragment from the left arm of chromosome IV of Saccharomyces cerevisiae was sequenced. Sequence analysis identified 20 complete non-overlapping open reading frames (ORFs) of at least 100 amino acids. Nine of these correspond to previously identified and sequenced genes: SIT4/PPH1, FAD1, NAM1/MTF2, RNA11, SIR2/MAR1, NAT1/AAA1, PRP9, ACT2 and MPS1/RPK1. Three ORFs show homology to previously sequenced genes. One ORF exhibits a hypothetical yabO/yceC/yfiI family signature and one has the ATP-dependent helicase signature of the DEAD and DEAH box families. Six ORFs show no appreciable homology to any proteins in the database. One of these is identical to yeast expressed sequence tags and therefore corresponds to an expressed gene. In addition, two partial ORFs and 11 ORFs that are totally internal and are not likely to be functional were detected. The sequence has been submitted to the EMBL data library under Accession Number Z71781. © 1997 by John Wiley & Sons, Ltd.     

Modulation of Th1/Th2 balance by Lactobacillus strains isolated from Kimchi via stimulation of macrophage cell line J774A.1 in vitro

Lactobacilli isolated from Kimchi, a Korean traditional food, were tested for their capacity to modulate the T helper (Th) 1/Th2 balance. Ovalbumin (OVA)-sensitized mouse splenocytes were cultured with 26 strains of lactobacilli; the highest IL-12 induction and lowest IL-4 production were then observed in 4 strains, including Lactobacillus plantarum CJLP55, CJLP56, CJLP133, and CJLP136. These strains produced a larger amount of IL-12, which enhances differentiation and activation of Th1 cells, in macrophage cell-lines more than positive control strains L. casei KCTC 3109(T) and L. rhamnosus GG, although they also induced production of IL-10, which is a suppressor of IL-12. Indeed, CJLP133-stimulated macrophages induced production of more Th1 cytokine IFN-γ and less Th2 cytokine IL-4 than KCTC 3109(T) and GG in co-cultivation with T cells. These findings suggest that lactobacilli from Kimchi may modulate the Th1/Th2 balance via macrophage activation in the hypersensitive reaction caused by Th2 cells. PRACTICAL APPLICATION: Allergic reactions including asthma and atopy are caused by predominance of Th2 response over Th1 response. Lactobacilli isolated from fermented foods such as yogurt, cheese, and Kimchi showed health-promoting activities. The present study indicated that several lactobacilli strains from Kimchi may reduce allergic reactions through macrophage-mediated induction of Th1 response.     

A 37·5 kb region of yeast chromosome X includes the SME1, MEF2, GSH1 and CSD3 genes,a TCP-1-related gene,an open reading frame similar to the DAL80 gene,and a tRNAArg

The complete DNA sequence of cosmid clone p59 comprising 37,549 bp derived from chromsome X was determined from an ordered set of subclones. The sequence contains 14 open reading frames (ORFs) containing at least 100 consecutive sense codons. Four of the ORFs represent already known and sequenced yeast genes: B645 is identical to the SME1 gene encoding a protein kinase, required for induction of meiosis in yeast, D819 represents the MEF2 gene probably encoding a second mitochondrial elongation factor-like protein, D678 is identical to the yeast GSH1 gene encoding γ-glutamylcysteine synthetase and B746 is identical to the CSD3 gene, which plays an as yet unidentified role in chitin biosynthesis and/or its regulation. The deduced amino acid sequence of A550 is 63% identical to the Ccη subunit of a murine TCP-1-containing chaperonin and more than 35% identical to thermophilic factor 55 from Sulfolobus shibatae, as well as to a number of proteins belonging to the chaperonin TCP-1 family. Open reading frame F551 exhibits homology to two regions of the DAL80 gene located on yeast chromosome XI encoding a pleiotropic negative regulatory protein. In addition, extensive homology was detected in three regions including parts of ORFs A560, B746/CSD3 and the incomplete ORF C852 to three consecutive ORFs of unknown function in the middle of the right arm of chromosome XI. Finally, the sequence contained a tRNA^Arg3 (AGC) gene. The nucleotide sequence data reported in this paper have been deposited in the EMBL and GenBank databases under the accession number X85021.     

Syringetin,a flavonoid derivative in grape and wine,induces human osteoblast differentiation through bone morphogenetic protein‐2/extracellular signal‐regulated kinase 1/2 pathway

Syringetin (3,5,7,4′‐tetrahydroxy‐3′,5′dimethoxyflavone), a flavonoid derivative, is present in grape and wine. By means of alkaline phosphatase (ALP) activity, osteocalcin, and type I collagen ELISA, we have shown that syringetin exhibits a significant induction of differentiation in MC3T3‐E1 mouse calvaria osteoblasts and human fetal osteoblastic 1.19 cell line human osteoblasts. ALP and osteocalcin are phenotypic markers for early‐stage differentiated osteoblasts and terminally differentiated osteoblasts, respectively. Our results indicate that syringetin stimulates osteoblast differentiation at various stages, from maturation to terminally differentiated osteoblasts. Induction of differentiation by syringetin is associated with increased bone morphogenetic protein‐2 (BMP‐2) production. The BMP‐2 antagonist noggin blocked syringetin‐mediated ALP activity and osteocalcin secretion enhancement, indicating that BMP‐2 production is required in syringetin‐mediated osteoblast maturation and differentiation. Induction of differentiation by syringetin is associated with increased activation of SMAD1/5/8 and extracellular signal‐regulated kinase 1/2 (ERK1/2). Cotreatment of ERK1/2 inhibitor 2′‐amino‐3′‐methoxyflavone inhibited syringetin‐mediated ALP upregulation and osteocalcin production. In conclusion, syringetin increased BMP‐2 synthesis, and subsequently activated SMAD1/5/8 and ERK1/2, and this effect may contribute to its action on the induction of osteoblast maturation and differentiation, followed by an increase of bone mass.     

Saccharomyces cerevisiae Gle2/Rae1 is involved in septin organization,essential for cell cycle progression

Gle2/Rae1 is highly conserved from yeast to humans and has been described as an mRNA export factor. Additionally, it is implicated in the anaphase‐promoting complex‐mediated cell cycle regulation in higher eukaryotes. Here we identify an involvement for Saccharomyces cerevisiae Gle2 in septin organization, which is crucial for cell cycle progression and cell division. Gle2 genetically and physically interacts with components of the septin ring. Importantly, deletion of GLE2 leads to elongated buds, severe defects in septin‐assembly and their cellular mislocalization. Septin‐ring formation is triggered by the septin‐regulating GTPase Cdc42, which establishes and maintains cell polarity. Additionally, activity of the master cell cycle regulator Cdc28 (Cdk1) is needed, which is, besides other functions, also required for G₂/M‐transition, and in yeast particularly responsible for initiating the apical–isotropic switch. We show genetic and physical interactions of Gle2 with both Cdc42 and Cdc28. Most importantly, we find that gle2? severely mislocalizes Cdc42, leading to defects in septin‐complex formation and cell division. Thus, our findings suggest that Gle2 participates in the efficient organization of the septin assembly network, where it might act as a scaffold protein. © 2017 The Authors. Yeast published by John Wiley & Sons, Ltd.