Slow growth phenotype - A possible approach to improved plasmid maintenance in Saccharomyces cerevisiae

1996 ◽  
Vol 18 (6) ◽  
pp. 713-718 ◽  
Author(s):  
Ronan O'Kennedy ◽  
J. W. Patching
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Slow Growth ◽  
Plasmid Maintenance ◽  
Growth Phenotype ◽  
Slow Growth Phenotype

scholarly journals CNS1 Encodes an Essential p60/Sti1 Homolog in Saccharomyces cerevisiae That Suppresses Cyclophilin 40 Mutations and Interacts with Hsp90

1998 ◽  
Vol 18 (12) ◽  
pp. 7344-7352 ◽  
Author(s):  
Kara J. Dolinski ◽  
Maria E. Cardenas ◽  
Joseph Heitman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cyclosporin A ◽  
Slow Growth ◽  
Protein A ◽  
Amino Terminal ◽  
Receptor Complexes ◽  
Growth Phenotype ◽  
Cyclophilin 40 ◽  
Tpr Domain ◽  
Slow Growth Phenotype

ABSTRACT Cyclophilins are cis-trans-peptidyl-prolyl isomerases that bind to and are inhibited by the immunosuppressant cyclosporin A (CsA). The toxic effects of CsA are mediated by the 18-kDa cyclophilin A protein. A larger cyclophilin of 40 kDa, cyclophilin 40, is a component of Hsp90-steroid receptor complexes and contains two domains, an amino-terminal prolyl isomerase domain and a carboxy-terminal tetratricopeptide repeat (TPR) domain. There are two cyclophilin 40 homologs in the yeast Saccharomyces cerevisiae, encoded by the CPR6 and CPR7 genes. Yeast strains lacking the Cpr7 enzyme are viable but exhibit a slow-growth phenotype. In addition, we show here that cpr7 mutant strains are hypersensitive to the Hsp90 inhibitor geldanamycin. When overexpressed, the TPR domain of Cpr7 alone complements both cpr7 mutant phenotypes, while overexpression of the cyclophilin domain of Cpr7, full-length Cpr6, or human cyclophilin 40 does not. The open reading frame YBR155w, which has moderate identity to the yeast p60 homologSTI1, was isolated as a high-copy-number suppressor of thecpr7 slow-growth phenotype. We show that this Sti1 homolog Cns1 (cyclophilin seven suppressor) is constitutively expressed, essential, and found in protein complexes with both yeast Hsp90 and Cpr7 but not with Cpr6. Cyclosporin A inhibited Cpr7 interactions with Cns1 but not with Hsp90. In summary, our findings identify a novel component of the Hsp90 chaperone complex that shares function with cyclophilin 40 and provide evidence that there are functional differences between two conserved sets of Hsp90 binding proteins in yeast.

scholarly journals Expression of Escherichia coli Methionyl-tRNA Formyltransferase in Saccharomyces cerevisiae Leads to Formylation of the Cytoplasmic Initiator tRNA and Possibly to Initiation of Protein Synthesis with Formylmethionine

2002 ◽  
Vol 22 (15) ◽  
pp. 5434-5442 ◽  
Author(s):  
Vaidyanathan Ramesh ◽  
Caroline Köhrer ◽  
Uttam L. RajBhandary
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Protein Synthesis ◽  
Slow Growth ◽  
Cytoplasmic Protein ◽  
Formyl Group ◽  
Initiator Trna ◽  
E Coli ◽  
Growth Phenotype ◽  
Slow Growth Phenotype

ABSTRACT Protein synthesis in eukaryotic cytoplasm and in archaebacteria is initiated with methionine, whereas, that in eubacteria and in eukaryotic organelles, such as mitochondria and chloroplasts, is initiated with formylmethionine. In view of this clear distinction, we have investigated whether protein synthesis in the eukaryotic cytoplasm can be initiated with formylmethionine, and, if so, what the consequences are to the cell. For this purpose, we have expressed in an inducible manner the Escherichia coli methionyl-tRNA formyltransferase (MTF) in the cytoplasm of the yeast Saccharomyces cerevisiae. Expression of active MTF, but not of an inactive mutant, leads to formylation of methionine attached to the yeast cytoplasmic initiator tRNA to the extent of about 70%. As a consequence, the yeast strain grows slowly. Coexpression of the E. coli polypeptide deformylase (DEF), which removes the formyl group from the N-terminal formylmethionine in a polypeptide, rescues the slow-growth phenotype, whereas, coexpression of an inactive mutant of DEF does not. These results suggest that the cytoplasmic protein-synthesizing system of yeast, like that of eubacteria, can at least to some extent utilize formylated initiator Met-tRNA to initiate protein synthesis and that initiation of proteins with formylmethionine leads to the slow-growth phenotype. Removal of the formyl group in these proteins by DEF would explain the rescue of the slow-growth phenotype.

Mutations in GSF1 and GSF2 Alter Glucose Signaling in Saccharomyces cerevisiae

Genetics ◽  
1997 ◽  
Vol 147 (2) ◽  
pp. 557-566 ◽  
Author(s):  
Peter W Sherwood ◽  
Marian Carlson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Slow Growth ◽  
Glucose Repression ◽  
Snf1 Kinase ◽  
Glucose Signaling ◽  
Glucose Depletion ◽  
Complementation Groups ◽  
Growth Phenotype ◽  
His3 Gene ◽  
Slow Growth Phenotype

One function of the Saccharomyces cerevisiae Snf1 protein kinase is to relieve glucose repression of SUC, GAL, and other genes in response to glucose depletion. To identify genes that regulate Snf1 kinase activity, we have selected mutants that inappropriately express a SUC2promoter::HIS3 gene fusion when grown in glucose and that require Snf1 function for this phenotype. Mutations representing two new complementation groups (gsf1 and gsf2) were isolated. gsf1 mutations affect two distinct responses to glucose: the Snf1-regulated glucose repression of SUC2 and GAL10 transcription and the Snf1-independent induction by glucose of HXT1 transcription. gsf2 mutations relieve glucose repression of SUC2 and GAL10 transcription and, in combination with snf1Δ, cause an extreme slow growth phenotype. The GSF2 gene was cloned by complementation of the gsf2-1 snf1Δ slow growth phenotype and encodes a previously uncharacterized 46 kD protein.

scholarly journals Responsiveness to exogenous cAMP of a Saccharomyces cerevisiae strain conferred by naturally occurring alleles of PDE1 and PDE2.

Genetics ◽  
1993 ◽  
Vol 135 (2) ◽  
pp. 321-326 ◽  
Author(s):  
H Mitsuzawa
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Slow Growth ◽  
Loss Of Function ◽  
Wild Type ◽  
Triple Mutant ◽  
Apparent Absence ◽  
Camp Pathway ◽  
Naturally Occurring ◽  
Elevated Activity ◽  
Growth Phenotype

Abstract The Saccharomyces cerevisiae strain P-28-24C, from which cAMP requiring mutants derived, responded to exogenously added cAMP. Upon the addition of cAMP, this strain showed phenotypes shared by mutants with elevated activity of the cAMP pathway. Genetic analysis involving serial crosses of this strain to a strain with another genetic background revealed that the responsiveness to cAMP results from naturally occurring loss-of-function alleles of PDE1 and PDE2, which encode low and high affinity cAMP phosphodiesterases, respectively. In addition, P-28-24C was found to carry a mutation conferring slow growth that lies in CYR1, which encodes adenylate cyclase, and the slow growth phenotype caused by the cyr1 mutation was suppressed by the pde2 mutation. Therefore P-28-24C is fortuitously a pde1 pde2 cyr1 triple mutant. Responsiveness to cAMP conferred by pde mutations suggests that S. cerevisiae cells are permeable to cAMP to some extent and that the apparent absence of effect of exogenously added cAMP on wild-type cells is due to immediate degradation by cAMP phosphodiesterases.

scholarly journals Transcriptional pathways associated with the slow growth phenotype of transformed Anaplasma marginale

BMC Genomics ◽  
2013 ◽  
Vol 14 (1) ◽  
pp. 272 ◽  
Author(s):  
Sebastián Aguilar Pierlé ◽  
Gena Kenitra Hammac ◽  
Guy H Palmer ◽  
Kelly A Brayton
Keyword(s):  
Slow Growth ◽  
Anaplasma Marginale ◽  
Growth Phenotype ◽  
Slow Growth Phenotype

scholarly journals Identification of Genes Required for Glucan Exopolysaccharide Production in Lactobacillus johnsonii Suggests a Novel Biosynthesis Mechanism

2020 ◽  
Vol 86 (8) ◽  
Author(s):  
Melinda J. Mayer ◽  
Alfonsina D’Amato ◽  
Ian J. Colquhoun ◽  
Gwénaëlle Le Gall ◽  
Arjan Narbad
Keyword(s):  
Slow Growth ◽  
Alternative Mechanism ◽  
Wild Type ◽  
Lactobacillus Johnsonii ◽  
Neighboring Gene ◽  
Amino Acid Homology ◽  
Content Type ◽  
Exopolysaccharide Production ◽  
Growth Phenotype ◽  
Slow Growth Phenotype

ABSTRACT Lactobacillus johnsonii FI9785 makes two capsular exopolysaccharides—a heteropolysaccharide (EPS2) encoded by the eps operon and a branched glucan homopolysaccharide (EPS1). The homopolysaccharide is synthesized in the absence of sucrose, and there are no typical glucansucrase genes in the genome. Quantitative proteomics was used to compare the wild type to a mutant where EPS production was reduced to attempt to identify proteins associated with EPS1 biosynthesis. A putative bactoprenol glycosyltransferase, FI9785_242 (242), was less abundant in the Δeps_cluster mutant strain than in the wild type. Nuclear magnetic resonance (NMR) analysis of isolated EPS showed that deletion of the FI9785_242 gene (242) prevented the accumulation of EPS1, without affecting EPS2 synthesis, while plasmid complementation restored EPS1 production. The deletion of 242 also produced a slow-growth phenotype, which could be rescued by complementation. 242 shows amino acid homology to bactoprenol glycosyltransferase GtrB, involved in O-antigen glycosylation, while in silico analysis of the neighboring gene 241 suggested that it encodes a putative flippase with homology to the GtrA superfamily. Deletion of 241 also prevented production of EPS1 and again caused a slow-growth phenotype, while plasmid complementation reinstated EPS1 synthesis. Both genes are highly conserved in L. johnsonii strains isolated from different environments. These results suggest that there may be a novel mechanism for homopolysaccharide synthesis in the Gram-positive L. johnsonii. IMPORTANCE Exopolysaccharides are key components of the surfaces of their bacterial producers, contributing to protection, microbial and host interactions, and even virulence. They also have significant applications in industry, and understanding their biosynthetic mechanisms may allow improved production of novel and valuable polymers. Four categories of bacterial exopolysaccharide biosynthesis have been described in detail, but novel enzymes and glycosylation mechanisms are still being described. Our findings that a putative bactoprenol glycosyltransferase and flippase are essential to homopolysaccharide biosynthesis in Lactobacillus johnsonii FI9785 indicate that there may be an alternative mechanism of glucan biosynthesis to the glucansucrase pathway. Disturbance of this synthesis leads to a slow-growth phenotype. Further elucidation of this biosynthesis may give insight into exopolysaccharide production and its impact on the bacterial cell.

NUCLEAR SUPPRESSORS OF THE [POKY] CYTOPLASMIC MUTANT IN NEUROSPORA CRASSA. I. GENETICS AND RESPIRATORY PROPERTIES

1976 ◽  
Vol 18 (2) ◽  
pp. 311-324 ◽  
Author(s):  
J. Kohout ◽  
H. Bertrand
Keyword(s):  
Neurospora Crassa ◽  
Ribosomal Proteins ◽  
Respiratory Activity ◽  
Slow Growth ◽  
Phenotypic Expression ◽  
Structure And Properties ◽  
Molecular Defects ◽  
Growth Phenotype ◽  
Slow Growth Phenotype

Six nuclear suppressors of the [poky] cytoplasmic mutant (sup-1, sup-3, sup-4, sup-5, sup-10, sup-14) have been obtained in Neurospora crassa. The sup genes suppress the slow growth phenotype of [poky], and alleviate, at least partially, the deficiency of cyanide-sensitive respiratory activity in the mycelium of this cytoplasmic mutant. The six suppressors are nonallelic, suppress the phenotypic effects of [stp-Bl] in addition to [poky], but have no effect on the phenotypic expression of the [mi-3] cytoplasmic mutant. On the basis of experimentally established molecular defects in [poky] and on the basis of hypothetical considerations, it is proposed that the sup mutations affect the structure and properties of mitochondrial ribosomal proteins.

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