scholarly journals Interactions among three proteins that specifically activate translation of the mitochondrial COX3 mRNA in Saccharomyces cerevisiae.

1994 ◽  
Vol 14 (2) ◽  
pp. 1045-1053 ◽  
Author(s):  
N G Brown ◽  
M C Costanzo ◽  
T D Fox
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Membrane ◽  
Wild Type ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Mutation ◽  
Mitochondrial Ribosomes ◽  
Functional Interactions ◽  
Cold Sensitive ◽  
Two Hybrid ◽  
Missense Substitution

The PET54, PET122, and PET494 proteins, which are associated with the yeast inner mitochondrial membrane, specifically activate translation of the mitochondrially encoded COX3 mRNA. We used the two-hybrid system to test whether pairs of these proteins, when fused to either the GAL4 DNA-binding or transcriptional activating domain, can physically associate as measured by the expression of the GAL4-dependent reporter, lacZ. PET54 and PET122 interacted in this system, and an amino-terminally truncated PET494 fragment showed an interaction with PET54. We also detected functional interactions between PET54 and PET122 genetically: a pet54 missense substitution (Phe to Gly at position 244) that caused a severe respiratory defect was suppressed both by a missense substitution affecting PET122 (Gly to Val at position 211) and by overproduction of wild-type PET122. Both Gly and Ala, substituted at PET54 position 244, disrupted the two-hybrid interactions with PET122 and PET494. While Ala at PET54 position 244 caused only a modest respiratory phenotype alone, it caused a severe respiratory defect when combined with a cold-sensitive mitochondrial mutation affecting the COX3 mRNA 5' leader. This synthetic defect was suppressed by a missense substitution in PET122 and by overproduction of wild-type PET122, indicating functional interactions among PET54, PET122, and the mRNA. Taken together with previous work, these data suggest that a complex containing PET54, PET122, and PET494 mediates the interaction of the COX3 mRNA with mitochondrial ribosomes at the surface of the inner membrane.

scholarly journals The ankyrin repeat-containing protein Akr1p is required for the endocytosis of yeast pheromone receptors.

1997 ◽  
Vol 8 (7) ◽  
pp. 1317-1327 ◽  
Author(s):  
S A Givan ◽  
G F Sprague
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Surface ◽  
Hybrid System ◽  
Secretory Pathway ◽  
Cytoplasmic Tail ◽  
Ankyrin Repeat ◽  
Wild Type ◽  
Delta Cells ◽  
Type Receptor ◽  
Two Hybrid

The Saccharomyces cerevisiae a-factor receptor (Ste3p) requires its C-terminal cytoplasmic tail for endocytosis. Wild-type receptor is delivered to the cell surface via the secretory pathway but remains there only briefly before being internalized and delivered to the vacuole for degradation. Receptors lacking all or part of the cytoplasmic tail are not subject to this constitutive endocytosis. We used the cytoplasmic tail of Ste3p as bait in the two-hybrid system in an effort to identify other proteins involved in endocytosis. One protein identified was Akr1p, an ankyrin repeat-containing protein. We applied three criteria to demonstrate that Akr1p is involved in the constitutive endocytosis of Ste3p. First, when receptor synthesis is shut off, akr1 delta cells retain the ability to mate longer than do AKR1 cells. Second, Ste3p half-life is increased by greater than 5-fold in akr1 delta cells compared with AKR1 cells. Third, after a pulse of synthesis, newly synthesized receptor remains at the cell surface in akr1 delta mutants, whereas it is rapidly internalized in AKR1 cells. Specifically, in akr1 delta mutants, newly synthesized receptor is accessible to exogenous protease, and by indirect immunofluorescence, the receptor is located at the cell surface. akr1 delta cells are also defective for endocytosis of the alpha-factor receptor (Ste2p). Despite the block to constitutive endocytosis exhibited by akr1 delta cells, they are competent to carry out ligand-mediated endocytosis of Ste3p. In contrast, akr1 delta cells cannot carry out ligand-mediated endocytosis of Ste2p. We discuss the implications for Akr1p function in endocytosis and suggest a link to the regulation of ADP-ribosylation proteins (Arf proteins).

Altered mitochondrial ribosomes in a cold-sensitive mutant of Saccharomyces cerevisiae

1978 ◽  
Vol 4 (2) ◽  
pp. 83-86 ◽  
Author(s):  
Terence W. Spithill ◽  
K. J. English ◽  
Phillip Nagley ◽  
Anthony W. Linnane
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Sensitive Mutant ◽  
Mitochondrial Ribosomes ◽  
Cold Sensitive

scholarly journals Suspended Animation Extends Survival Limits of Caenorhabditis elegans and Saccharomyces cerevisiae at Low Temperature

2010 ◽  
Vol 21 (13) ◽  
pp. 2161-2171 ◽  
Author(s):  
Kin Chan ◽  
Jesse P. Goldmark ◽  
Mark B. Roth
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Division ◽  
Caenorhabditis Elegans ◽  
Cell Division Cycle ◽  
Wild Type ◽  
Suspended Animation ◽  
C Elegans ◽  
High Viability ◽  
Cold Sensitive

The orderly progression through the cell division cycle is of paramount importance to all organisms, as improper progression through the cycle could result in defects with grave consequences. Previously, our lab has shown that model eukaryotes such as Saccharomyces cerevisiae, Caenorhabditis elegans, and Danio rerio all retain high viability after prolonged arrest in a state of anoxia-induced suspended animation, implying that in such a state, progression through the cell division cycle is reversibly arrested in an orderly manner. Here, we show that S. cerevisiae (both wild-type and several cold-sensitive strains) and C. elegans embryos exhibit a dramatic decrease in viability that is associated with dysregulation of the cell cycle when exposed to low temperatures. Further, we find that when the yeast or worms are first transitioned into a state of anoxia-induced suspended animation before cold exposure, the associated cold-induced viability defects are largely abrogated. We present evidence that by imposing an anoxia-induced reversible arrest of the cell cycle, the cells are prevented from engaging in aberrant cell cycle events in the cold, thus allowing the organisms to avoid the lethality that would have occurred in a cold, oxygenated environment.

scholarly journals Increased expression of Saccharomyces cerevisiae translation elongation factor 1 alpha bypasses the lethality of a TEF5 null allele encoding elongation factor 1 beta.

Genetics ◽  
1995 ◽  
Vol 141 (2) ◽  
pp. 481-489 ◽  
Author(s):  
T G Kinzy ◽  
J L Woolford
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Null Allele ◽  
Elongation Factor ◽  
Wild Type ◽  
Translation Elongation Factor ◽  
Gene Encoding ◽  
Translation Elongation ◽  
Dominant Mutant ◽  
Cold Sensitive ◽  
Elongation Factor 1

Abstract Translation elongation factor 1beta (EF-1beta) catalyzes the exchange of bound GDP for GTP on EF-1alpha. The lethality of a null allele of the TEF5 gene encoding EF-1beta in Saccharomyces cerevisiae was suppressed by extra copies of the TEF2 gene encoding EF-1alpha. The strains with tef5::TRP1 suppressed by extra copies of TEF were slow growing, cold sensitive, hypersensitive to inhibitors of translation elongation and showed increased phenotypic suppression of +1 frameshift and UAG nonsense mutations. Nine dominant mutant alleles of TEF2 that cause increased suppression of frameshift mutations also suppressed the lethality of tef5::TRP1. Most of the strains in which tef5::TRP1 is suppressed by dominant mutant alleles of TEF2 grew more slowly and were more antibiotic sensitive than strains with tef5::TRP1 is suppressed by wild-type TEF2. Two alleles, TEF2-4 and TEF2-10, interact with tef5::TRP1 to produce strains that showed doubling times similar to tef5::TRP1 strains containing extra copies of wild-type TEF2. These strains were less cold sensitive, drug sensitive and correspondingly less efficient suppressor of +1 frameshift mutations. These phenotypes indicate that translation and cell growth are highly sensitive to changes in EF-1alpha and EF-1beta activity.

scholarly journals Role of cytochrome c heme lyase in mitochondrial import and accumulation of cytochrome c in Saccharomyces cerevisiae.

1991 ◽  
Vol 11 (11) ◽  
pp. 5487-5496 ◽  
Author(s):  
M E Dumont ◽  
T S Cardillo ◽  
M K Hayes ◽  
F Sherman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cytochrome C ◽  
Mitochondrial Membrane ◽  
Intermembrane Space ◽  
Inner Mitochondrial Membrane ◽  
Yeast Saccharomyces Cerevisiae ◽  
Apocytochrome C ◽  
Cytochrome C Heme Lyase

Heme is covalently attached to cytochrome c by the enzyme cytochrome c heme lyase. To test whether heme attachment is required for import of cytochrome c into mitochondria in vivo, antibodies to cytochrome c have been used to assay the distributions of apo- and holocytochromes c in the cytoplasm and mitochondria from various strains of the yeast Saccharomyces cerevisiae. Strains lacking heme lyase accumulate apocytochrome c in the cytoplasm. Similar cytoplasmic accumulation is observed for an altered apocytochrome c in which serine residues were substituted for the two cysteine residues that normally serve as sites of heme attachment, even in the presence of normal levels of heme lyase. However, detectable amounts of this altered apocytochrome c are also found inside mitochondria. The level of internalized altered apocytochrome c is decreased in a strain that completely lacks heme lyase and is greatly increased in a strain that overexpresses heme lyase. Antibodies recognizing heme lyase were used to demonstrate that the enzyme is found on the outer surface of the inner mitochondrial membrane and is not enriched at sites of contact between the inner and outer mitochondrial membranes. These results suggest that apocytochrome c is transported across the outer mitochondrial membrane by a freely reversible process, binds to heme lyase in the intermembrane space, and is then trapped inside mitochondria by an irreversible conversion to holocytochrome c accompanied by folding to the native conformation. Altered apocytochrome c lacking the ability to have heme covalently attached accumulates in mitochondria only to the extent that it remains bound to heme lyase.

scholarly journals The cold sensitivity of a mutant of Saccharomyces cerevisiae lacking a mitochondrial heat shock protein 70 is suppressed by loss of mitochondrial DNA.

1996 ◽  
Vol 134 (3) ◽  
pp. 603-613 ◽  
Author(s):  
B Schilke ◽  
J Forster ◽  
J Davis ◽  
P James ◽  
W Walter ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Cold Sensitivity ◽  
Growth Defect ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
The Matrix ◽  
Cold Sensitive

SSH1, a newly identified member of the heat shock protein (hsp70) multigene family of the budding yeast Saccharomyces cerevisiae, encodes a protein localized to the mitochondrial matrix. Deletion of the SSH1 gene results in extremely slow growth at 23 degrees C or 30 degrees C, but nearly wild-type growth at 37 degrees C. The matrix of the mitochondria contains another hsp70, Ssc1, which is essential for growth and required for translocation of proteins into mitochondria. Unlike SSC1 mutants, an SSH1 mutant showed no detectable defects in import of several proteins from the cytosol to the matrix compared to wild type. Increased expression of Ssc1 partially suppressed the cold-sensitive growth defect of the SSH1 mutant, suggesting that when present in increased amounts, Ssc1 can at least partially carry out the normal functions of Ssh1. Spontaneous suppressors of the cold-sensitive phenotype of an SSH1 null mutant were obtained at a high frequency at 23 degrees C, and were all found to be respiration deficient. 15 of 16 suppressors that were analyzed lacked mitochondrial DNA, while the 16th had reduced amounts. We suggest that Ssh1 is required for normal mitochondrial DNA replication, and that disruption of this process in ssh1 cells results in a defect in mitochondrial function at low temperatures.

scholarly journals Genetic Interactions Between HOP1, RED1 and MEK1 Suggest That MEK1 Regulates Assembly of Axial Element Components During Meiosis in the Yeast Saccharomyces cerevisiae

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 33-42 ◽  
Author(s):  
Nancy M Hollingsworth ◽  
Lisa Ponte
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genetic Interactions ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sister Chromatids ◽  
Cytological Data ◽  
Negative Effect ◽  
Two Hybrid ◽  
Axial Element ◽  
Putative Protein Kinase

Abstract During meiosis, axial elements are generated by the condensation of sister chromatids along a protein core as precursors to the formation of the synaptonemal complex (X). Functional axial elements are essential for wild-type levels of recombination and proper reductional segregation at meiosis I. Genetic and cytological data suggest that three meiosis-specific genes, HOP1, RED1 and MEK1, are involved in axial element formation in the yeast Saccharomyces cerevisiae. HOP1 and RED1 encode structural components of axial elements while MEK1 encodes a putative protein kinase. Using a partially functional allele of MEK1, new genetic interactions have been found between HOP1, RED1 and MEK1. Overexpression of HOP1 partially suppresses the spore inviability and recombination defects of mekl-974; in contrast, overexpression of RED1 exacerbates the mek1-974 spore inviability. Co-overexpression of HOP1 and RED1 in mek1-974 diploids alleviates the negative effect of overexpressing RED1 alone. Red1p/Red1p as well as Hop1p/Red1p interactions have been reconstituted in two hybrid experiments. Our results suggest a model whereby Mekl kinase activity controls axial element assembly by regulating the affinity with which Hoplp and Redlp interact with each other.

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