Search PubMed⌕ Search

Biomedical subjects

M Drolet

Publications and source records attributed to M Drolet.

At least 37 records · Page 2Linked to original sources
PubMed1993

A protein binding site from the murine c-myc promoter contributes to transcriptional block.

Recent studies have revealed that expression of several eukaryotic genes can be regulated at the level of transcription elongation. As a first step to elucidate the mechanism by which transcription elongation is modulated, several groups have identified sequences necessary for transcriptional block within the c-myc gene. These studies indicated that transcriptional block depends not only on sequences surrounding the sites of block, but also on sequences within the promoter: some deletions within the c-myc promoter eliminated transcriptional block and, with chimeric constructs, transcriptional block was observed when some heterologous promoters but not others were fused to the c-myc termination region. Using a chimeric construct containing the H-2Kb major histocompatibility class gene promoter linked to the c-myc first exon, we show that transcriptional block is increased by the addition of a 25 bp DNA sequence from the c-myc promoter. Similar results are obtained whether this sequence is inserted upstream or downstream of the transcription initiation site. We further show that nuclear factors interact with this sequence in vitro. Interestingly, when a mutated version of this sequence was tested, we observed decreased nuclear factor binding in vitro as well as reduced transcriptional block in nuclear run-on transcription assays. These results suggest that interactions of protein factors with specific nucleotide sequences near the transcription initiation site can affect elongation of transcription at sites located further downstream.

Animals↗
PubMed1992

Mobilization protein-DNA binding and divergent transcription at the transfer origin of the Thiobacillus ferrooxidans pTF1 plasmid.

The possible interaction of the trans-acting mobilization proteins, MobL and MobS, at the cognate origin of transfer (oriT) region of the Thiobacillus ferrooxidans plasmid pTF1 has been investigated. In gel retardation assays with crude protein extracts from overproducing strains, a truncated MobL (c. 28 kDa) as well as its native protein (42 kDa), but not the 11 kDa MobS protein, were found to bind specifically to a 42-mer oligonucleotide which represents the transferred DNA strand of the minimal oriT fragment of pTF1. In vivo, the binding of MobL was studied by monitoring catechol 2,3-dioxygenase (xylE) activities driven by promoters of the divergently transcribed mobL and mobS genes. The mob promoter sequences were found to resemble the Escherichia coli sigma 70-dependent consensus promoter elements. The '-10' recognition sequences of mobL and one of the two mobS promoters overlap except for one base and they are positioned within the putative 'hairpin' structure in the minimal oriT sequence. In accordance with the twin supercoil-domain model of Liu and Wang (1987) which suggests that transcription can generate local variations in DNA superhelicity, we propose a possible physiological role of DNA supercoiling in the transfer origin with reference to divergent transcription of mobL and mobS genes.

Bacterial Proteins↗
PubMed1991

Cloning and nucleotide sequence of the hemA gene of Agrobacterium radiobacter.

The hemA gene of Agrobacterium radiobacter ATCC4718 was identified by hybridization with a hemA probe from Rhizobium meliloti and cloned by complementation of a hemA mutant of Escherichia coli K12. E. coli hemA transformants carrying the hemA gene of Agrobacterium showed delta-aminolevulinic acid synthetase (delta-ALAS) activity in vitro. The hemA gene was carried on a 4.4 kb EcoRI fragment which could be reduced to a 2.6 kb EcoRI-SstI fragment without affecting its complementing or delta-ALAS activity. The sequence of the hemA gene showed an open reading frame of 1215 nucleotides, which could code for a protein of 44,361 Da. This is very close to the molecular weight of the HemA protein obtained using an in vitro coupled transcription-translation system (45,000 Da). Comparison of amino acid sequences of the delta-ALAS of A. radiobacter and Bradyrhizobium japonicum showed strong homology between the two enzymes; less, but still significant, homology was observed when A. radiobacter and human delta-ALAS were compared. Primer extension experiments enabled us to identify two promoters for the hemA gene of A. radiobacter. One of these promoters shows some similarity to the first promoter of the hemA gene of R. meliloti.

5-Aminolevulinate Synthetase↗
PubMed1990

The mobilization and origin of transfer regions of a Thiobacillus ferrooxidans plasmid: relatedness to plasmids RSF1010 and pSC101.

The components for the mobilization function of a plasmid DNA during conjugation include a cis-acting sequence (the origin of transfer, oriT) and a transacting sequence coding for mobilization (Mob) proteins. By genetic and deletion analysis, we have located the mobilization region of pTF1, a cryptic plasmid previously isolated from a Thiobacillus ferrooxidans strain. Within a 2797 bse-pair sequenced region, several open reading frames (ORFs) were predicted; two of the ORFs are divergently transcribed and they encode proteins of calculated molecular masses, 42.6kD (ORF2) and 11.4kD (ORF6). Surprisingly, these protein sequences are substantially similar to two of the previously characterized mobilization proteins of the Escherichia coli IncQ plasmid, RSF1010. Moreover, the pTF1 ORF2 (now designated MobL) sequence is also found to be similar to a presumed mobilization protein of plasmid pSC101. Regions of sequence identity of plasmids pTF1, RSF1010 and pSC101 include their oriT sites. By alkaline agarose gel electrophoresis and DNA sequencing, we have established the location of the relaxation complex nick site within the oriT of pTF1. An identical nick site, which is adjacent to a characteristic 10 base-pair inverted repeat sequence, is also found for plasmid RSF1010. A recombinant plasmid containing a 42 base-pair synthetic piece of DNA encompassing the pTF1 inverted repeat and nick sequence was shown to be oriT-active.

Amino Acid Sequence↗
PubMed1989

Isolation and nucleotide sequence of the hemA gene of Escherichia coli K12.

The hemA gene of Escherichia coli K12 was cloned by complementation of a hemA mutant of this organism. Subcloning of the initial 6.0 kb HindIII fragment allowed the isolation of a 1.5 kb NheI-AvaI fragment which retained the ability to complement the hemA mutant. DNA sequencing by the dideoxy chain terminator method of Sanger showed the presence of an open reading frame (ORF) of 1254 nucleotides, which ends 6 nucleotides beyond the AvaI site. Primer extension experiments showed the existence of a putative transcription initiation site for the hemA gene, located at position 130. A possible promoter sequence was identified upstream from this transcription initiation site, and its functional activity was confirmed by the use of the pK01 promoter-probe vector. Protein synthesis in an in vitro coupled transcription-translation system showed a 46 kDa protein, which corresponds to the mol. wt. of the hemA protein, as deduced from the nucleotide sequence of the gene. No homology was found between the amino acid sequence of the hemA protein of E. coli K12 and known sequences of other delta-aminolevulinic acid synthases (delta-ALAS), suggesting that this protein is different from other delta-ALAS enzymes.

5-Aminolevulinate Synthetase↗
PubMed1988

Nucleotide sequence of the hemB gene of Escherichia coli K12.

The hemB gene of Escherichia coli K12, coding for porphobilinogen synthase (PBG-S; syn., 5-aminolevulinic acid dehydratase, ALA-D), was cloned following insertion of an EcoRI fragment of plasmid F'13 into the mobilizable vector pCR1. The hybrid plasmid carrying the hemB gene was able to complement a hemB mutant of E. coli K12: not only was the PBG-S activity of the mutant restored after the acquisition of the hemB gene, but it was about ten times higher than that of the wild type. Subcloning of the original EcoRI fragment (14.6 kb) enabled us to locate the hemB gene on an NruI-HpaI fragment of about 1.1 kb. The hemB promoter was located toward the NruI end of the fragment, as shown by the use of the pKO promoter-probe series of vectors. Sequencing of the hemB gene indicated the presence of an open reading frame (ORF) of 1051 nucleotides, which should correspond to the HemB protein. Primer extension experiments enabled us to identify the 5' end of the hemB mRNA, and to deduce the -10 and -35 regions of the hemB promoter. Protein synthesis performed by an in vitro coupled transcription-translation system, showed the presence of a protein of about 35 kDa. This is in agreement with the molecular weight of the HemB protein (35.6 kDa), as deduced from the nucleotide sequence of the gene. Comparison of the amino acid sequences of E. coli and human PBG-S allowed the detection of several regions of strong homology between the two proteins. Two of these regions correspond, as expected, to the putative zinc-binding and catalytic sites of the human PBG-S.

Amino Acid Sequence↗
PubMed1987

Molecular cloning and sequencing of the hemD gene of Escherichia coli K-12 and preliminary data on the Uro operon.

DNA of plasmid pSAS1002TH (F' ilv+ hemD+ hemC+ cya+) was used to clone the hemD gene of Escherichia coli K-12. Due to poor transformability of the heme-deficient mutants, the restriction fragments of the F' plasmid were first cloned into a mobilizable derivative of pBR322, pSAS1211LP, which was then mobilized into a hemD recA mutant (E. coli SASX419AN). One recombinant plasmid, carrying a HindIII fragment of about 5 kilobases (kb), was shown to complement the hemD mutant and also a cya mutant of E. coli K-12, as well as a hemC mutant of Salmonella typhimurium LT2. Further subcloning of the insert enabled us to locate the hemD gene to a BamHI-PstI fragment (approximately 2.3 kb) which also carried the hemC gene. The hemD gene occupies a region close to the PstI end, since the deletion of a 0.6-kb fragment from this end resulted in loss of the ability to complement the hemD mutation. The use of the promoter-probe vector pK01 and the results of complementation showed that the hemD gene was transcribed under physiological conditions from the same promoter as the hemC gene, the direction of transcription being hemC-hemD. This allows us to define a new polycistronic operon of E. coli K-12, for which we propose the designation Uro operon. Sequencing of the hemD gene showed the presence of an open reading frame (ORF) of 738 nucleotides which could code for a protein with a molecular weight of 27,766, which should correspond to the hemD protein; the ORF starts with the last nucleotide of the hemC gene, the two genes having different reading frames. An ORF of at least 480 base pairs follows the hemD gene after a few nucleotides. The corresponding gene X, the function of which is unknown, might represent a third member of the Uro operon.

Amino Acid Sequence↗
PubMed1985

Biofeedback control of migraine headaches: a comparison of two approaches.

In order to assess the relative effectiveness of finger warming and temporal blood volume pulse reduction biofeedback in the treatment of migraine, 22 female migraine patients were assigned to one of three experimental conditions: temporal artery constriction feedback, finger temperature feedback, or waiting list. Biofeedback training consisted of 12 sessions over a 6-week period. All patients completed 5 weeks of daily self-monitoring of headache activity (frequency, duration, and intensity) and medication before and after treatment. Treatment credibility was assessed at the end of Sessions 1, 6, and 12. Results showed that temporal constriction and finger temperature biofeedback were equally effective in controlling migraine headaches and produced greater benefits than the waiting list condition. Power analyses indicated that very large sample sizes would have been required to detect any significant differences between the two treatment groups. No significant relationships were found between levels of therapeutic gains and levels of thermal or blood volume pulse self-regulation skills. Likewise, treatment outcome was not found to be related to treatment credibility. Further analyses revealed that changes in headache activity and medication were associated with changes in vasomotor variability. Because blood volume pulse variability was not significantly affected by biofeedback training, questions about its role in the therapeutic mechanism are raised.

Adult↗
PubMed1983

Blood volume pulse biofeedback in the treatment of migraine headache: a controlled evaluation.

In order to evaluate the specific effects of blood volume pulse (BVP) biofeedback in the treatment of migraine headaches, 21 female migraine patients were randomly assigned to one of three experimental conditions: temporal artery constriction feedback, temporal artery dilation feedback, or waiting list. Biofeedback training consisted of 15 sessions over an 8-week period. All patients completed 5 weeks of daily self-monitoring of headache activity and medication before and after treatment. Results showed that constriction and dilation biofeedback were equally effective in controlling migraines and produced greater benefits than the waiting-list condition. No significant relationships were found between therapeutic gains and BVP self-regulation skills. However, further analyses revealed that changes in headache activity and medication were associated with changes in vasomotor variability. The current rationale for the use of BVP biofeedback in the treatment of migraine is questioned and a new one is proposed.

Adolescent↗
PubMed1981

Evaluation of skin temperature biofeedback training at two different sites for migraine.

All combinations of cooling versus warming and finger versus temporal artery were used in the present study, the aim of which was to identify an optimal biofeedback training site and to assess the specific effects of skin temperature biofeedback upon migraine. After an initial 4-week baseline phase, during which daily records of headache activity and medication were kept, 24 migraine patients were randomly assigned to one of the four experimental conditions. Training sessions for all patients were of 50-min duration and occurred once per week for 8 weeks. The headache charts were completed for another 4 weeks after treatment was terminated and again at 6-month follow-up. The results showed significant reductions in migraine activity and drug usage. However, self-regulation of skin temperature in different directions at different sites did not in fact result in significantly different magnitudes of change in migraine activity across groups. The role of nonspecific factors is discussed and a physiological model is proposed to explain how warming and cooling may produce clinical improvement in migraine.

Adult↗
PubMed1975

Carotid endarterectomy: survival rates of 227 patients.

From March 1963 to March 1974, 227 patients with carotid stenosis underwent unilateral or bilateral carotid endarterectomy at l'Hôpital de l'Enfant-Jésus in Québec. Survival during the first 7 years after operation was analysed by life-table methods based on full intervals. The observed probability of death in this group was significantly higher than the probability expected in the general population, by 6.8% in the 1st year and 3.9% in the 2nd year. In the next 3 years the differences were much smaller (0.5, 0.9 and 1.4%). The major causes of death were myocardial infarction and stroke. Women fared somewhat better than men after the 1st year of follow-up. Unexpectedly, patients who underwent unilateral endarterectomy had lower survival rates in each of the first 5 years after operation than patients who underwent staged bilateral operations, whether survival was measured from the date of the first or the second operation.

Adult↗