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C Jacq

Publications and source records attributed to C Jacq.

At least 37 records · Page 2Linked to original sources
PubMed1996

Complete nucleotide sequence of Saccharomyces cerevisiae chromosome X.

The complete nucleotide sequence of Saccharomyces cerevisiae chromosome X (745 442 bp) reveals a total of 379 open reading frames (ORFs), the coding region covering approximately 75% of the entire sequence. One hundred and eighteen ORFs (31%) correspond to genes previously identified in S. cerevisiae. All other ORFs represent novel putative yeast genes, whose function will have to be determined experimentally. However, 57 of the latter subset (another 15% of the total) encode proteins that show significant analogy to proteins of known function from yeast or other organisms. The remaining ORFs, exhibiting no significant similarity to any known sequence, amount to 54% of the total. General features of chromosome X are also reported, with emphasis on the nucleotide frequency distribution in the environment of the ATG and stop codons, the possible coding capacity of at least some of the small ORFs (<100 codons) and the significance of 46 non-canonical or unpaired nucleotides in the stems of some of the 24 tRNA genes recognized on this chromosome.

Amino Acid Sequence↗
PubMed1996

Analysis of a 23 kb region on the left arm of yeast chromosome IV.

We have determined the complete nucleotide sequence of a 23 kb segment from the left arm of chromosome IV, which is carried by the cosmid 1L10. This sequence contains the 3' coding region of the STE7 and RET1 (COP1) genes, and 13 complete open reading frames longer than 300 bp, of which ten correspond to putative new genes and three (CLB3, MSH5 and RPC53) have been sequenced previously. The sequence from cosmid IL10 was obtained entirely by a combined subcloning and walking primer strategy.

Amino Acid Sequence↗
PubMed1995

Limitations to in vivo import of hydrophobic proteins into yeast mitochondria. The case of a cytoplasmically synthesized apocytochrome b.

The apocytochrome b gene, exclusively encoded by the mitochondrial genome, was engineered so that it could be expressed in the yeast cytoplasm. Different combinations of the apocytochrome b transmembrane domains were produced in the form of hybrid proteins fused to both the N-terminal mitochondrial targeting sequence of the ATPase subunit 9 from Neurospora crassa and to a cytoplasmic version of the bI4 RNA maturase, localised on the N-terminal and C-terminal sides, respectively, of the hydrophobic stretches. The bI4 RNA maturase, which can complement mitochondrial mutations, was used as an in vivo reporter to assess the mitochondrial import of the different groups of transmembrane helices. This new, reliable and sensitive reporter activity allowed us to experimentally determine the limitations to the mitochondrial import of hydrophobic proteins. All eight transmembrane helices of apocytochrome b could be imported into mitochondria, either alone or in combination, but no more than three to four transmembrane helices could be imported together at one time. This limit is close to that observed in the population of nuclear-encoded mitochondrial proteins. The hydrophobic characteristics of engineered and natural proteins targeted to the mitochondrial inner membrane revealed two factors important in the import process. These were (a) the local hydrophobicity of a transmembrane segment, and (b) the average regional hydrophobicity of the protein over an extended length of 60-80 residues. Such features may have played a major role in the evolution of mitochondrial genomes.

Amino Acid Sequence↗
PubMed1995

Positive autoregulation of the yeast transcription factor Pdr3p, which is involved in control of drug resistance.

Simultaneous resistance to an array of drugs with different cytotoxic activities is a property of Saccharomyces cerevisiae, in which the protein Pdr3p has recently been shown to play a role as a transcriptional regulator. We provide evidence that the yeast PDR3 gene, which encodes a zinc finger transcription factor implicated in certain drug resistance phenomena, is under positive autoregulation by Pdr3p. DNase I footprinting analyses using bacterially expressed Pdr3p showed specific recognition by this protein of at least two upstream activating sequences in the PDR3 promoter. The use of lacZ reporter constructs, a mutational analysis of the upstream activating sequences, as well as band shift experiments enabled the identification of two 5'TC CGCGGA3' sequence motifs in the PDR3 gene as consensus elements for the binding of Pdr3p. Several similar sequence motifs can be found in the promoter of PDR5, a gene encoding an ATP-dependent drug pump whose Pdr3p-induced overexpression is responsible for drug resistance phenomena. Recently one of these sequence elements was shown to be the target of Pdr3p to elevate the level of PDR5 transcription. Finally, we provide evidence in the absence of PDR1 for a PDR3-controlled transcriptional induction of the drug pump by cycloheximide and propose a model for the mechanism governing the transcriptional autoregulation of Pdr3p.

ATP-Binding Cassette Transporters↗
PubMed1994

Complete DNA sequence of yeast chromosome II.

In the framework of the EU genome-sequencing programmes, the complete DNA sequence of the yeast Saccharomyces cerevisiae chromosome II (807 188 bp) has been determined. At present, this is the largest eukaryotic chromosome entirely sequenced. A total of 410 open reading frames (ORFs) were identified, covering 72% of the sequence. Similarity searches revealed that 124 ORFs (30%) correspond to genes of known function, 51 ORFs (12.5%) appear to be homologues of genes whose functions are known, 52 others (12.5%) have homologues the functions of which are not well defined and another 33 of the novel putative genes (8%) exhibit a degree of similarity which is insufficient to confidently assign function. Of the genes on chromosome II, 37-45% are thus of unpredicted function. Among the novel putative genes, we found several that are related to genes that perform differentiated functions in multicellular organisms of are involved in malignancy. In addition to a compact arrangement of potential protein coding sequences, the analysis of this chromosome confirmed general chromosome patterns but also revealed particular novel features of chromosomal organization. Alternating regional variations in average base composition correlate with variations in local gene density along chromosome II, as observed in chromosomes XI and III. We propose that functional ARS elements are preferably located in the AT-rich regions that have a spacing of approximately 110 kb. Similarly, the 13 tRNA genes and the three Ty elements of chromosome II are found in AT-rich regions. In chromosome II, the distribution of coding sequences between the two strands is biased, with a ratio of 1.3:1. An interesting aspect regarding the evolution of the eukaryotic genome is the finding that chromosome II has a high degree of internal genetic redundancy, amounting to 16% of the coding capacity.

Base Composition↗
PubMed1994

PDR3, a new yeast regulatory gene, is homologous to PDR1 and controls the multidrug resistance phenomenon.

The Saccharomyces cerevisiae PDR3 gene, located near the centromere of chromosome II, has been completely sequenced and characterised. Mutations pdr3-1 and pdr3-2, which confer resistance to several antibiotics can be complemented by a wild-type allele of the PDR3 gene. The sequence of the wild-type PDR3 gene revealed the presence of a long open reading frame capable of encoding a 976-amino acid protein. The protein contains a single Zn(II)2Cys6 binuclear-type zinc finger homologous to the DNA-binding motifs of other transcriptional activators from lower eukaryotes. Evidence that the PDR3 protein is a transcriptional activator was provided by demonstrating that DNA-bound LexA-PDR3 fusion proteins stimulate expression of a nearby promoter containing LexA binding sites. The use of LexA-PDR3 fusions revealed that the protein contains two activation domains, one localised near the N-terminal, cysteine-rich domain and the other localised at the C-terminus. The salient feature of the PDR3 protein is its similarity to the protein coded by PDR1, a gene responsible for pleiotropic drug resistance. The two proteins show 36% amino acid identity over their entire length and their zinc finger DNA-binding domains are highly conserved. The fact that the absence of both PDR1 and PDR3 (simultaneous disruption of the two genes) enhances multidrug sensitivity strongly suggests that the two transcriptional factors have closely related functions.

Amino Acid Sequence↗
PubMed1993

I-Sce III an intron-encoded DNA endonuclease from yeast mitochondria. Asymmetrical DNA binding properties and cleavage reaction.

We have previously discovered the new intron-encoded endonuclease I-Sce III by expressing, in E. coli, the ORF contained in the third intron of the yeast mitochondrial COX I gene. In this work, we analyzed the in vitro properties of partially purified I-Sce III and found that it is a very specific DNA endonuclease, tolerating relatively few base changes in its 20 base pair long target site. I-Sce III should be a useful molecular tool to analyze the structure of large genomes. Interestingly, I-Sce III is the first P1-P2 DNA endonuclease for which DNA binding properties could be analyzed by band-shift experiments. Clearly, the cleavage products corresponding to the upstream A3 exon and to the downstream A4 exon could compete with the substrate A3-A4 in forming a DNA-protein complex. However, the A3 exon competes more efficiently than the downstream A4 product. The cleavage of the two DNA strands is also asymmetric the top strand (non-transcribed strand) is cleaved faster than the bottom strand, a property found under various experimental conditions. These findings suggest that this intron-encoded DNA endonuclease may have role in the RNA splicing process of the intron.

Amino Acid Sequence↗
PubMed1992

The yeast mitochondrial intron aI5 alpha: associated endonuclease activity and in vivo mobility.

By analyzing crosses between yeast strains carrying different combinations of mitochondrial (mt) introns, we have shown that the aI5 alpha intron is mobile in vivo. Furthermore, we have observed that the mobility of intron aI5 alpha is affected by both the nuclear and mt genotypes. We have also detected a restriction endonuclease (ENase) activity that cleaves intronless mt genomes close to the aI5 alpha intron insertion site and thus might be involved in intron mobility. This is further supported by the fact that this ENase activity is only detected in a strain containing the aI5 alpha intron. Furthermore, similar to other ENases encoded by mobile mt introns of yeast, the ENase generates a cut with a four-base 3'-OH overhang. Thus, intron aI5 alpha represents a characteristic member of the family of mobile group-I introns.

Base Sequence↗
PubMed1992

Sequence of a 12.7 kb segment of yeast chromosome II identifies a PDR-like gene and several new open reading frames.

A 12,684 bp DNA fragment, between FUS3 and the centromere, from the left arm of chromosome II of Saccharomyces cerevisiae was sequenced as part of the European project to sequence the whole chromosome. This segment contains at least five complete new open reading frames (ORFs) and the beginning (191 first 5' codons) of an ORF whose putative translational product is highly similar to the multidrug resistance PDR1 gene previously characterized by Balzi et al. (1987) on chromosome VII.

Amino Acid Sequence↗
PubMed1992

Connections between RNA splicing and DNA intron mobility in yeast mitochondria: RNA maturase and DNA endonuclease switching experiments.

The intron-encoded proteins bI4 RNA maturase and aI4 DNA endonuclease can be faithfully expressed in yeast cytoplasm from engineered forms of their mitochondrial coding sequences. In this work we studied the relationships between these two activities associated with two homologous intron-encoded proteins: the bI4 RNA maturase encoded in the fourth intron of the cytochrome b gene and the aI4 DNA endonuclease (I-SceII) encoded in the fourth intron of the gene coding for the subunit I of cytochrome oxidase. Taking advantage of both the high recombinogenic properties of yeast and the similarities between the two genes, we constructed in vivo a family of hybrid genes carrying parts of both RNA maturase and DNA endonuclease coding sequences. The presence of a sequence coding for a mitochondrial targeting peptide upstream from these hybrid genes allowed us to study the properties of their translation products within the mitochondria in vivo. We thus could analyze the ability of the recombinant proteins to complement RNA maturase deficiencies in different strains. Many combinations of the two parental intronic sequences were found in the recombinants. Their structural and functional analysis revealed the following features. (i) The N-terminal half of the bI4 RNA maturase could be replaced in total by its equivalent from the aI4 DNA endonuclease without affecting the RNA maturase activity. In contrast, replacing the C-terminal half of the bI4 RNA maturase with its equivalent from the aI4 DNA endonuclease led to a very weak RNA maturase activity, indicating that this region is more differentiated and linked to the maturase activity. (ii) None of the hybrid proteins carrying an RNA maturase activity kept the DNA endonuclease activity, suggesting that the latter requires the integrity of the aI4 protein. These observations are interesting because the aI4 DNA endonuclease is known to promote the propagation, at the DNA level, of the aI4 intron, whereas the bI4 RNA maturase, which is required for the splicing of its coding intron, also controls the splicing process of the aI4 intron. We propose a scenario for the evolution of these intronic proteins that relies on a switch from DNA endonuclease to RNA maturase activity.

Amino Acid Sequence↗
PubMed1991

A new specific DNA endonuclease activity in yeast mitochondria.

Two group I intron-encoded proteins from the yeast mitochondrial genome have already been shown to have a specific DNA endonuclease activity. This activity mediates intron insertion by cleaving the DNA sequence corresponding to the splice junction of an intronless strain. We have discovered in mitochondrial extracts from the yeast strain 777-3A a new DNA endonuclease activity which cleaves the fused exon A3-exon A4 junction sequence of the CO XI gene.

Base Sequence↗
PubMed1990

In vivo and in vitro analyses of an intron-encoded DNA endonuclease from yeast mitochondria. Recognition site by site-directed mutagenesis.

The pal 4 nuclease (termed I-Sce II) is encoded in the group I al 4 intron of the COX I gene of Saccharomyces cerevisiae. It introduces a specific double-strand break at the junction of the two exons A4-A5 and thus mediates the insertion of the intron into an intronless strain. To define the sequence recognized by pal 4 we introduced 35 single mutations in its target sequence and examined their cleavage properties either in vivo in E. coli (when different forms of the pal 4 proteins were artificially produced) or in vitro with mitochondrial extracts of a mutant yeast strain blocked in the splicing of the al 4 intron. We also detected the pal 4 DNA endonuclease activity in extracts of the wild type strain. The results suggest that 6 to 9 noncontiguous bases in the 17 base-pair region examined are necessary for pal 4 nuclease to bind and cleave its recognition site. We observed that the pal 4 nuclease specificity can be significantly different with the different forms of the protein thus explaining why only some forms are highly toxic in E. coli. This study shows that pal 4 recognition site is a complex phenomenon and this might have evolutionary implications on the transfer properties of the intron.

Base Sequence↗
PubMed1990

Expression in Saccharomyces cerevisiae of a gene associated with cytoplasmic male sterility from maize: respiratory dysfunction and uncoupling of yeast mitochondria.

We asked whether the mitochondrial T-urf13 gene, associated with the male sterility phenotype of T cytoplasm in maize, can be expressed in Saccharomyces cerevisiae and whether this expression can mimic the effects observed in maize. We introduced the universal code equivalent of the T-urf13 gene into the S. cerevisiae nucleus by transformation and directed its translation product into mitochondria by means of a fusion with the targeting presequence from Neurospora crassa AT-Pase subunit 9. We show that expression of the universal code equivalent of the T-urf13 gene in the yeast nucleus does indeed mimic its effects in maize: respiratory growth of yeast is inhibited, respiration-deficient cytoplasmic mutants accumulate and NADH oxidation of isolated mitochondria is uncoupled. All these effects are observed only if the mitochondrial targeting peptide and methomyl or HmT toxin are present.

Chimera↗
PubMed1989

Mobile introns: definition of terms and recommended nomenclature.

A number of introns in mitochondrial, chloroplast, nuclear or prokaryotic genes have recently been shown to encode double-strand sequence-specific endonucleases. Such introns are mobile genetic elements that insert themselves at or near the cleaved sites. A uniform nomenclature to designate the molecular elements involved in the phenomenon of intron mobility is proposed.

Base Sequence↗
PubMed1989

Site-specific DNA endonuclease and RNA maturase activities of two homologous intron-encoded proteins from yeast mitochondria.

Two introns of the mitochondrial genome 777-3A of S. cerevisiae, bl4 in cob and al4 in coxl genes, contain ORFs that can be translated into two homologous proteins. We changed the UGA, AUA, and CUN codons of these ORFs to the universal genetic code, in order to study the functions of their translated products in E. coli and in yeast, by retargeting the nuclear encoded protein into mitochondria. The p27bl4 protein has been shown to be required for the splicing of both introns bl4 and al4. The homologous p28al4 protein is highly toxic to E. coli. It can specifically cleave double-stranded DNA at a sequence representing the junction of the two fused flanking exons. We present evidence that this system is a good model for studying the role of mitochondrial intron-encoded proteins in the rearrangement of genetic information at both the RNA (RNA splicing-bl4 maturase) and DNA levels (intron transposition-al4 transposase).

Base Sequence↗