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J Pelletier

Publications and source records attributed to J Pelletier.

At least 109 records · Page 6Linked to original sources
PubMed1996

Germline WT1 mutations in Wilms' tumor patients: preliminary results.

We conducted a comparative study of the prevalence of germline WT1 mutations in patients with Wilms' tumor. Patients in Group 1 have familial Wilms' tumor, bilateral disease, associated urogenital anomalies, and/or second cancers. Those in Group 2 are unilateral, sporadic Wilms' patients without other associated conditions. Patients with aniridia or Denys-Drash syndrome are known to have WT1 alterations, and are excluded from this study. Preliminary results on 96 subjects show that the overall germline WT1 mutation frequency is low (< 5%). The work to date establishes the feasibility of identifying patients with germline WT1 mutations and, in the future, offering genetic predisposition testing to at-risk relatives. However, genetic predisposition testing of children for WT1 mutations raises many ethical, legal, and psychosocial issues; research is needed to evaluate risks and benefits.

Base Sequence↗
PubMed1996

Abnormal gonadal differentiation in two subjects with ambiguous genitalia, Mullerian structures, and normally developed testes: evidence for a defect in gonadal ridge development.

Among a group of patients with abnormal sexual differentiation, we have identified two subjects who had a 46,XY karyotype, ambiguous genitalia, and well-developed Müllerian structures, but normal appearing testes. The presence of ambiguous genitalia and persistent Müllerian structures implied both Leydig cell and Sertoli cell dysfunction, hence, gonadal dysgenesis. However, the normal testicular histology suggested that the underlying abnormality was not a defect in testis determination itself but an abnormality in timing of gonadal ridge and testis development. In one of the two subjects genomic DNA was available. The sequence of the SRY gene was normal. Because rare patients with partial androgen insensitivity may have a similar phenotype, the AR gene was evaluated by denaturing gradient gel electrophoresis (DGGE) and was normal. Some subjects with mutation of the WT1 gene or with deletion of the distal short arm of chromosome 9 may have similar phenotypes. The WT1 gene was studied by single-strand conformation polymorphism (SSCP) analysis and was normal. In addition, there was no loss of heterozygosity of polymorphic markers in distal 9p. The gene for Müllerian inhibiting substance (MIS) was also studied by SSCP and was normal. Although the exact mechanism for the defect in the two subjects is unknown, it may be due to an abnormality in a gene or genes involved in the timing of gonadal ridge development.

Cell Differentiation↗
PubMed1996

The renal brush border membrane sodium/sulfate cotransporter functions in situ as a homotetramer.

The functional molecular size of the renal Na+/SO4(2-) cotransporter was analysed with the radiation inactivation and fragmentation method. Purified brush border membrane vesicles preserved in a cryoprotective medium were exposed to gamma-radiations. Initial rates of SO4(2-) influx into these vesicles were estimated with membranes irradiated with 0, 4 and 8 Mrad. In each case, SO4(2-) uptake by irradiated membranes was significantly reduced but remained linear during the first 5 sec of incubation. To avoid artifacts arising from a decrease in the driving force caused by modifications in membrane permeability, this incubation period was chosen to measure the effect of irradiation on the SO4(2-) transport activity. Increasing irradiation doses resulted in a monoexponential decrease in transport activity allowing the molecular size to be estimated at 238 +/- 6 kDa (SD, n = 3). Recently, a cDNA for the Na+/SO4(2-) cotransporter was cloned and expressed in Xenopus laevis oocytes (Markovich D. et al. (1993) Proc. Natl Acad. Sci. U.S.A. 90, 8073-8077). The deduced amino acid sequence of this cotransporter predicts a molecular weight of 66 kDa. We suggest that the in situ activity of the renal brush border membrane Na+/SO4(2-) cotransporter requires the presence of four intact and identical subunits arranged as a homotetramer.

Animals↗
PubMed1996

Evidence for a familial Wilms' tumour gene (FWT1) on chromosome 17q12-q21.

Wilms' tumour (WT) is one of the most common solid tumours of childhood, occurring in 1 in 10,000 children and accounting for 8% of childhood cancers. It is believed to result from malignant transformation of abnormally persistent renal stem cells (nephrogenic rests) which retain embryonic differentiation potential. Although WT is usually sporadic, approximately one percent occur in families in which susceptibility appears to be inherited as an autosomal dominant trait with incomplete penetrance. Predisposition to other cancers or to the developmental abnormalities associated with sporadic WT is not usually apparent in WT families. The WT1 gene at 11p13 (ref.2), and additional genes on chromosomes 11p15 (ref. 3) and 16q (ref. 4) have been implicated in the development of WT but are not responsible for familial WT. We have carried out a genome linkage search in a large Canadian family with seven confirmed cases of WT. Our results provide strong evidence for the localisation of a familial WT predisposition gene, FWT1, to an 18-centimorgan (cM) interval on chromosome 17q12-q21.

Child↗
PubMed1996

[Reflex epilepsy with seizures induced by mental calculation, playing chess and scrabble].

Reflex seizures induced by higher mental activity is rare. We report the case of a young man with myoclonic jerks and generalized tonico-clonic convulsion precipitated by calculation, playing chess and scrabble. Routine EEG, including hyperventilation and photic stimulation, showed no abnormality. Tests stimulation procedures, including spatial tasks, induced focal and generalized EEG spike-wave complex and myoclonic jerks. Valproate was effective in reducing epileptic seizures during a follow-up period of three years. Comparison of our case with previously reported reflex epilepsy with seizures induced by higher mental activity is discussed.

Adult↗
PubMed1995

Antagonism of WT1 activity by protein self-association.

Germline loss-of-function mutations at the Wilms tumor (WT) suppressor locus WT1 are associated with a predisposition to WTs and mild genital system anomalies. In contrast, germ-line missense mutations within the WT1 gene encoding the DNA-binding domain often yield a more severe phenotype consisting of WT, sexual ambiguity, and renal nephropathy. In this report, we demonstrate that the products of mutant alleles that impair DNA recognition can antagonize WT1-mediated transcriptional repression. We demonstrate that WT1 can self-associate in vitro and in vivo and that the responsible domain maps to the amino-terminal region of the protein. Oligomers of full-length protein form less efficiently or produce less stable complexes than oligomers between truncated polypeptides and full-length protein. Our data suggest a molecular mechanism to explain how WT1 mutations may act in deregulating cellular proliferation and differentiation.

Alleles↗
PubMed1995

The human steroidogenic acute regulatory (StAR) gene is expressed in the urogenital system and encodes a mitochondrial polypeptide.

The first enzymatic step in the biosynthesis of steroid hormones occurs in the mitochondrial inner membrane and is dependent on the mobilization of cholesterol from cellular stores. We report on the isolation of a human cDNA which encodes a mitochondrial protein called steroidogenic acute regulatory (StAR) protein, implicated in transport of cholesterol into mitochondria. Nucleotide and predicted amino acid sequence analyses indicate that the human and murine polypeptides are highly conserved, sharing 87% identity with an overall homology of 92%. Analysis of the distribution of StAR mRNA transcripts in human tissues by Northern blotting reveals several mRNA species, the most abundant of which is a 1.8 kb mRNA transcript present in testes, ovaries and kidneys. Using in vitro translated protein, we demonstrate that the StAR gene product can be efficiently imported into exogenously added mitochondria.

Amino Acid Sequence↗
PubMed1995

Repression of the retinoic acid receptor-alpha gene by the Wilms' tumor suppressor gene product, wt1.

The Wilms' tumor (WT) suppressor gene, WT1, encodes a zinc finger DNA binding protein (wt1) which functions as a transcriptional regulator. Germline WT1 mutations predispose to WTs and in many cases are associated with urogenital anomalies. Identification of wt1 downstream targets is essential to understanding regulatory processes involved in development of this system. In this study, we demonstrate that wt1 can repress transcription of the retinoic acid receptor-alpha 1 (RAR-alpha 1) promoter. Transient transfection, deletion mutagenesis, and mobility shift assays suggest that wt1 mediates repression of the human RAR-alpha 1 promoter through a GC-rich DNA binding motif (5'-GCGGGGGCG-3'), at positions -111 to -120 bp (relative to the transcription initiation site). In contrast, the murine RAR-alpha 1 promoter contains a cryptic binding motif and is not responsive to wt1. These results indicate that some wt1-regulatory pathways are not conserved across species, suggesting a molecular basis for differences in phenotypes between humans and mice harboring WT1 lesions.

Base Sequence↗
PubMed1995

Clonal expansion and attenuated apoptosis in Wilms' tumors are associated with p53 gene mutations.

The p53 gene product is required for activation of an apoptotic pathway triggered by oncogenes and cytotoxic agents. Wilms' tumor, a pediatric renal malignancy, provides a paradigm for evaluating genetic events involved in tumor progression. This malignancy is generally not associated with p53 mutations, and even in advanced disease states is quite responsive to current treatment regimens. The anaplastic histological variant of Wilms' tumor, however, is frequently associated with p53 gene mutations and shows poor prognosis. We analyzed seven Wilms' tumors for which we had paired samples from nonanaplastic and anaplastic regions. p53 mutations were detected in six of these tumors, five of which demonstrated mutations restricted to anaplastic regions. Nonanaplastic cells of the sixth sample were heterozygous for a p53 mutation, whereas the anaplastic area of this tumor showed reduction to homozygosity. These results indicate that progression to anaplasia is associated with clonal expansion of cells which have acquired a p53 mutation. We demonstrated that tumor cells with p53 mutations show attenuated apoptosis, suggesting that such lesions may provide a selective advantage in vivo by decreasing cell death.

Anaplasia↗
PubMed1995

Changes in LHbeta-gene and FSHbeta-gene expression in the ram pars tuberalis according to season and castration.

Luteinizing hormone beta (LHbeta) and follicle stimulating hormone beta (FSHbeta) subunits and their mRNAs were studied in the ram pars tuberalis following different seasonal (winter vs summer) and experimental (intact vs castrated animals) conditions. Hormone-containing cells were identified by immunohistochemistry, using homologous double-stranded 35S-cDNAs. The labelling was quantified by image analysis. Immunohistochemical staining showed that cells containing LHbeta and FSHbeta were localized mainly in the ventral part of the pars tuberalis but that, in the summer, additional LHbeta containing cells were present in the dorsal part in intact rams. On the other hand, LHbeta-mRNA labelling was found in the whole pars tuberalis in wethers but only in the ventral part in intact rams. The magnitude of LHbeta-mRNA labelling was significantly greater in summer than in winter rams, and in castrated than in intact animals (P<0.001). However, the number of labelled cells was found to be the greatest in the winter (P<0.001) and was not affected by castration. FSHbeta-mRNA expression was similar to that of LHbeta-mRNA except that the level and extent were considerably lower. Thus, our results show an increase in the magnitude of gonadotropin beta subunit-mRNA in the summer and following castration; this increase appears to involve the entire pars tuberalis.

Animals↗
PubMed1995

Nephropathy with Wilms tumour or gonadal dysgenesis: incomplete Denys-Drash syndrome or separate diseases?

UNLABELLED: We report three children, one presenting with nephropathy, bilateral Wilms tumour (WT) and cryptorchism, one with combined nephropathy and gonadal dysgenesis and one with nephropathy which developed 13 years after a WT. The first case was recognized as typical Denys-Drash syndrome (DDS) which is characterized by the combination of nephropathy, intersex disorders and WT. The two other patients, who did not express the full spectrum of the syndrome, were older than 10 years, when they reached and stage renal failure. The fact that nephropathy in childhood is combined with such rare diseases like gonadal dysgenesis and/or WT, supports the concept of a common aetiology with DDS. Therefore, the patients were analysed for possible Wilms tumour suppressor gene (WT1) mutations. In all three individuals mutations in the heterozygous configuration could be demonstrated. CONCLUSION: These results provide evidence that incomplete and complete DDS are diseases of the same spectrum. WT1 analysis of more children with two symptoms of the triad of DDS should be helpful in establishing genotype-phenotype correlations and in understanding differences in the clinical picture of DDS.

Adolescent↗
PubMed1995

Sequence and structural requirements for high-affinity DNA binding by the WT1 gene product.

The Wilms' tumor suppressor gene, WT1, encodes a zinc finger polypeptide which plays a key role regulating cell growth and differentiation in the urogenital system. Using the whole-genome PCR approach, we searched murine genomic DNA for high-affinity WT1 binding sites and identified a 10-bp motif 5'GCGTGGGAGT3' which we term WTE). The WTE motif is similar to the consensus binding sequence 5'GCG(G/T)GGGCG3' recognized by EGR-1 and is also suggested to function as a binding site for WT1, setting up a competitive regulatory loop. To evaluate the underlying biochemical basis for such competition, we compared the binding affinities of WT1 and EGR1 for both sequences. WT1 shows a 20- to 30-fold-higher affinity for the WTE sequence compared with that of the EGR-1 binding motif. Mutational analysis of the WTE motif revealed a significant contribution to binding affinity by the adenine nucleotide at the eighth position (5'GCGTGGGAGT3') as well as by the 3'-most thymine (5'GCGTGGGAGT3'), whereas mutations in either flanking nucleotides or other nucleotides in the core sequence did not significantly affect the specific binding affinity. Mutations within WT1 zinc fingers II to IV abolished the sequence-specific binding of WT1 to WTE, whereas alterations within the first WT1 zinc finger reduced the binding affinity approximately 10-fold but did not abolish sequence recognition. We have thus identified a WT1 target, which, although similar in sequence to the EGR-1 motif, shows a 20- to 30-fold-higher affinity for WT1. These results suggest that physiological action of WT1 is mediated by binding sites of significantly higher affinity than the 9-bp EGR-1 binding motif. The role of the thymine base in contributing to binding affinity is discussed in the context of recent structural analysis.

Base Sequence↗
PubMed1995

An efficient strategy to isolate full-length cDNAs based on an mRNA cap retention procedure (CAPture).

The ability to generate cDNA libraries is one of the most fundamental procedures in contemporary molecular biology. One of the major drawbacks of current methods is that most cDNAs present in any given library are incomplete, rendering the characterization of genes an inefficient and time-consuming task. We have developed an affinity selection procedure using a fusion protein containing the murine cap-binding protein (eukaryotic initiation factor 4E), coupled to a solid support matrix, that allows for the purification of mRNAs via the 5' cap structure. When combined with a single-strand-specific RNase digestion step, specific retention of complete cDNA-RNA duplexes following first-strand synthesis is achieved. This method can be used to generate cDNA libraries in which polyadenylated and nonpolyadenylated mRNAs are equally represented and to enrich for full-length or 5'-end clones, thus facilitating cDNA cloning and promoter mapping.

Amino Acid Sequence↗
PubMed1994

Fine structure analysis of the WT1 gene in sporadic Wilms tumors.

Molecular genetic studies indicate that the etiology of Wilms tumor (WT) is complex, involving at least three loci. Germ-line mutations in the tumor suppressor gene, WT1, have been documented in children with WTs and urogenital developmental anomalies. Sporadic tumors constitute the majority (> 90%) of WT cases and previous molecular analyses of the WT1 gene have focused only on the DNA-binding domain. Using the single-strand conformational polymorphism (SSCP) assay, we analyzed the structural integrity of the entire WT1 gene in 98 sporadic WTs. By PCR-SSCP we find that mutations in the WT1 gene are rare, occurring in only six tumors analyzed. In one sample, two independent intragenic mutations inactivated both WT1 alleles, providing a singular example of two different somatic alterations restricted to the WT1 gene. This case is consistent with the existence of only one tumor suppressor gene at 11p13 involved in the pathogenesis of WTs. Our data, together with the previously ascertained occurrence of large deletions/insertions in WT1, define the frequency at which the WT1 gene is altered in sporadic tumors.

Amino Acid Sequence↗
PubMed1994

Women's deficiency in water-level representation: present in visual conditions yet absent in haptic contexts.

The present experiment studied horizontality representation among men and women performing the water-level task either in visual conditions (the subjects saw outlines of tilted containers in which they respectively drew or set the water line) or in partially and totally haptic conditions (respectively, the subjects both saw and felt or merely felt the contours of the containers cut out of metal plates, and positioned a rod on the underside of the plates to indicate the water line). It was expected that, as the visual components of the setting were replaced by haptic ones, a reduction of the typical gender difference in proficiency would ensue. It was found that men surpassed women under visual conditions, whereas both genders were equivalent in haptic conditions. There were no gender or condition differences in a control task in which a line had to be placed horizontally in tilted containers. Forced reliance on proprioceptive cues among both men and women under the totally haptic condition was contrasted with the critical role played in visual conditions by visual references; those used by men were correct, whereas the ones used by women were incorrect.

Adult↗