Cloning E. coli genes by oligonucleotide hybridization.
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Cloning, Molecular↗
Biomedical subjects
F Soubrier
Publications and source records attributed to F Soubrier.
Renin secretion from malignant pulmonary metastatic tumour cells of vascular origin.
1. A pulmonary chemodectoma/glomangiosarcoma that had metastasized from the thigh was studied after removal from a 22 year old Algerian patient with hypertension, high plasma prorenin and signs of secondary aldosteronism. 2. Renin and renin mRNA were localized in sections of the tumour tissue using monoclonal anti-human renin antibody and human renin cDNA probe, respectively. 3. The cells grew prolifically in culture, but, even though their renin content was similar to that of transfected human juxtaglomerular cell tumour cells (approximately 1 pg/microgram DNA), their rate of secretion of renin was much lower (0.05-0.15 cf. 0.5-1.5 pg/h per microgram DNA). 4. Forskolin (10 mumol/l for 24 h) increased secretion of renin from 1.9 +/- 0.36 to 4.1 +/- 0.64 pg/ml per h of culture (P less than 0.001, n = 11), consistent with cAMP being a second messenger in the secretory mechanism. 5. The cells should provide valuable information about intracellular mechanisms for the regulation of renin synthesis and secretion.
Adolescent↗
Segmental homology between the promoter region of the human renin gene and the mouse ren1 and ren2 promoter regions.
We have isolated and determined the nucleotide (nt) sequence of the 5' region of the human renin gene (h-ren). Two TATA boxes and a CAAT box were found. Start point determination has shown that only the proximal TATA box was used as the transcription initiation signal, both in the kidney and in a renin-secreting tumor. Comparison of the sequence of the 500-bp region upstream from the transcription start point with the corresponding regions of the ren1 and ren2 genes of the Swiss mouse revealed no overall homology between the human and mouse renin sequences. Only very short sequences of high homology ranging in size from 10 to 18 nt were found in the sequenced regions. By hybridization experiments, we have detected a region upstream from each mouse renin gene related to the h-ren; analysis of the nt sequence of this region reveals that they belong to the Alu family of repetitive DNA.
Animals↗
Myocardial recruitment during ANF mRNA increase with volume overload in the rat.
A synthetic oligonucleotide probe complementary to messenger ribonucleic acid (mRNA) encoding for the C-terminal portion of atrial natriuretic factor (ANF) has been used to study the expression of the ANF gene in rat myocardium. Four experimental models were studied: binephrectomy (for 48 h); ligature of both ureters (for 48 h); deoxycorticosterone acetate-salt (for 3 wk); and aortocaval fistula (for 2 wk). Analysis of atrial RNA by gel-blot hybridization detected a single band, corresponding in length to that of mRNA coding for ANF. Such an mRNA was also detected in ventricular RNA but was 1/50th as abundant. In the four experimental groups ANF mRNA was increased significantly as compared with controls. In all rats there was no significant difference in the ANF mRNA content between the left and the right atrium. Each experimental condition was accompanied by a highly significant increase in ANF gene expression in the left ventricle, where all of the ventricular tissue could be recruited and with a negative gradient from the base to the apex of the left ventricle. These data were confirmed by in situ hybridization. Thus all of the atrial and ventricular myocardium can express the ANF gene. Recruitment increases in response to passive stretch of the cardiac chambers.
Animals↗
Molecular cloning and nucleotide sequence of a human renin cDNA fragment.
We have studied human renin messenger RNA by hybridization with the mouse submaxillary gland (SMG) renin cDNA probe. The human kidney messenger RNA is about 1.6 kilobase (kb) long, similarly to the mouse SMG renin mRNA. A kidney renin cDNA clone of 1.1 kb length was obtained. A comparison of nucleotide sequences of mouse and human cDNA clones reveals conservation of residues involved in catalytic mechanisms and a potential glycosylation site. The human renin molecular probe allowed us to study renin expression in human chorionic tissue. The chorionic and kidney renin messenger RNAs are similar in length. The Southern blot analysis reveals the presence of a single renin gene in human DNA.
Amino Acid Sequence↗
Mouse submaxillary renin: a useful model for the study of renal renin.
The submaxillary gland of mouse contains a renin-like enzyme which represents as much as 5% of the total protein content. Its physico-chemical and enzymatic characteristics are similar to those of renal renin. Recently, the amino-acid sequence of the submaxillary pre-prorenin molecule has been deduced from the nucleotide sequence of the renin structural gene. A model for pre-prorenin processing into active renin has been proposed. Comparison of the structures of mouse submaxillary renin and of aspartyl proteases shows that renin belongs to this class of proteins and shares a similar catalytic site. Although the structure of renal renin is not yet known, preliminary studies suggest that the renal pro-enzyme is processed as the submaxillary enzyme. However, glycosylation would occur in the case of renal renin, whereas submaxillary renin is not glycosylated. Genetic studies and DNA hybridization experiments in mouse with high or low renin content in the submaxillary gland show that submaxillary renin in high renin producing strains results from a gene duplication. Submaxillary renin is therefore an isoenzyme and a useful model for the study of renal renin.
Amino Acid Sequence↗
Biochemical and immunological characterization of ectopic tumoral renin.
Biochemical and immunological characteristics of renin secreted by two malignant renin-secreting tumors [pulmonary (PT) and paraovarian (POT)] were studied. They both contain inactive renin (IR), as renin activity of tumoral extracts was able to be increased after acid activation or trypsin treatment (10.1 to 20.8 Goldblatt units/g tissue for PT and 1.4 to 3.71 for POT). Renin activity after activation reached the value obtained by direct RIA of human renin (23 and 3.4, respectively), as both forms are recognized by renin antiserum. Both enzymatic activities could be completely inhibited by renin antiserum. Displacement curves for the two tumoral renins paralleled the MRC renin in the direct RIA. After chromatography on affigel blue, active renin was not bound to the gel, and inactive renin eluted only with 1 M NaCl. On pepstatin A Sepharose and CBL-pepstatin Sepharose (an N-modified-pepstatin), a separation of the two forms of pulmonary renin was obtained; inactive renin eluted with breakthrough proteins, whereas active renin was strongly bound to the gel. After this affinity chromatography, the molecular weights of inactive and active renin, determined on Ultrogel, were very close (46,000 and 42,500). We conclude that 1) ectopic renin in these cases in similar to the renal enzyme; 2) renin can be secreted in an inactive form, supporting the hypothesis of an inactive initial state of renin; and 3) molecular weight differences between the two forms are very slight.
Adenocarcinoma↗
[Diagnostic value of computerized tomography in severe renal tissue infections (author's transl)].
A patient with severe acute pyonephritis and septicaemia had persistent symptoms of infection despite appropriate chemotherapy. Although no obstruction was seen on ascending ureteropyelography, exploratory lumbar incision was considered. However, computerized tomography (CT scan) failed to show any obstacle or collection of pus but indicated diffuse infection of kidney. Medical treatment was continued and the patient recovered. This case and a review of the literature has prompted the authors to compare CT scan with conventional contrast-radiography methods in septicaemias of renal origin. It appeared that intravenous and ascending uretero-pyelography are often at fault, and the latter is known to carry iatrogenic risks. CT scan is of considerable help when looking for foci of infection and is proposed as a deciding examination in septicaemias with or without signs suggestive of renal lesions.
Acute Disease↗
A monoclonal antibody for immunopurification of human renin.
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Animals↗
Activation of renin in an anaplastic pulmonary adenocarcinoma.
1. Biochemical characteristics of a renin-like enzyme secreted by a pulmonary adenocarcinoma have been studied. A very high renin content was revealed by both enzymatic (10.4 Goldblatt units/g of tissue) and direct radioimmunoassay of immunoreactive renin (23 Goldblatt units/g of tissue). 2. The higher value by direct radioimmunoassay suggested the presence of an inactive form of the enzyme. Indeed 40--100% activation occurred with treatment by trypsin or pepsin or prolonged dialysis at pH 7.4 with or without prior acid dialysis. This neutral activation was completely abolished by a serine proteinase inhibitor. 3. A large fraction of the renin in this tumour is inactive. In comparison with other prohormones produced in tumours the findings support strongly the proposition that renin passes through a proenzyme step in synthesis.
Adenocarcinoma↗
Monoclonal antibody against human renin.
By spleen cell fusion with NS1 myeloma, a mouse hybridoma was obtained which secretes an antibody directed against human renin. This monoclonal antibody recognizes human and monkey renin, but neither hog nor mouse. Preliminary experiments demonstrate the potential of this antibody for renin immunopurification and characterization.
Animals↗
[Normal pulmonary capillary pressure in hemodynamic pulmonary edema].
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Blood Pressure↗
Nonproportional changes in plasma renin concentration, renal renin content, and rat renin messenger RNA.
The expression of the renin gene in rat kidneys was studied using mouse submaxillary gland renin complementary DNA. The length of rat renin messenger RNA (mRNA) was approximately 1600 nucleotides, similar to that of mouse submaxillary gland and kidney renin mRNA. Rat renin mRNA was quantified by a radiodensitometric complementary DNA hybridization assay. The effects of intense long-term stimulation and short-term inhibition of renin secretion on plasma renin concentration, renal renin concentration, and renin mRNA content were compared with those of controls. After 15 days of sodium depletion and captopril treatment, plasma renin concentration increased 46-fold, renal renin concentration only 1.5-fold, and renin mRNA content increased about threefold. Following a 1-hour infusion of angiotensin II in sodium-depleted and captopril-treated rats, plasma renin concentration decreased by 84% whereas no significant changes in either renal renin concentration or renin mRNA content were observed. These results show that sodium depletion and captopril treatment increase the level of renin gene transcription and renin biosynthesis. However, there are nonproportional changes in plasma renin levels, renal renin content, and its mRNA. These results suggest that newly synthesized renin is not stored in the kidney but is rapidly secreted into the blood. Short-term inhibition of plasma renin concentration by angiotensin II is most likely mediated by posttranslational mechanisms.
Angiotensin II↗
[Prevalence of factor V Leiden, hyperhomocysteinemia, prothrombin G20210A, and methylene tetrahydrofolate reductase C677T mutations in obstetrical complications].
The etiology and pathogenesis of intrauterine fetal death, preeclampsia or fetal growth retardation remain still unknown in many cases. However, placental thrombosis and/or infarction might lead to inadequate maternal-fetal circulation. So, the relevance of an additional thrombotic risk factor that enhances the physiological hypercoagulable state of gestation has been suggested in the development of these adverse outcomes of pregnancy. Several genetic mutations are newly recognized associated with an increased frequency of venous thrombosis: mutation of adenine to guanine at nucleotide 506 in the factor V gene, mutation of cytosine at nucleotide 677 in the methylenetetrahydrofolate gene and mutation of guanine to adenine at nucleotide 20210 in the prothrombin gene. In this issue, a review of literature has allowed us to evaluate the prevalence of these genetic predisposing thrombotic factors with the development of obstetrical complications. Furthermore, therapeutic approach is considered.
Factor V↗
[Mutation in the MSH2 gene in Muir-Torre syndrome].
BACKGROUND: The Muir-Torre syndrome is an autosomal dominant hereditary condition predisposing to cancer. It is characterized by skin tumors associated with adenocarcinoma of the colon or other neoplasias observed in the context of hereditary non-polyposis colorectal cancer (HNPCC). The Muir-Torre syndrome is also characterized by the frequent presence of multiple colonic polyps and the relatively moderate aggressivity of the tumors. CASE REPORT: We studied a family with Muir-Torre syndrome. We sequenced the exons of the hMSH2 gene in this family and identified heterozygous germinal mutation by G insert at position 2427 (2427insG). This mutation changes the lecture phase producing a premature codon stop. DISCUSSION: Our study confirms the predominant responsibility of the hMSH2 gene in Tuir-Torre syndrome. This clinical case and data reported in the literature demonstrate the importance of searching for a history of non-polyposis colorectal cancer in patients and relatives and the unstable genome characteristic of these tumors found in sebaceous tumors or keratoacanthomas. Sequencing the hMSH2 gene should be a priority when clinical features, history and laboratory tests suggest Muir-Torre syndrome.
Adenocarcinoma↗
[Comparison of three reactants for the detection of activated protein C resistance due to mutation of factor V Leiden during pregnancy].
The presence of the R506Q mutation of the factor V gene is associated with an increased risk of thromboembolism, particularly during pregnancy. Recently, its involvement in the development of obstetrical complications, such as preeclampsia and fetal losses, has been evoked. The resulting factor VQ506 (factor V Leiden) has arginine 506 replaced by glutamine at the factor Va cleavage site for activated protein C (APC) which induces APC-resistance. During pregnancy, an acquired resistance to APC is observed without the presence of the factor V Leiden mutation which leads to an inappropriate realization of the more expensive DNA analysis. This resistance is at least partly explained by an increase of the factor VIII. In this study, we have compared three reagents: the original test Coatest APC Resistance (Chromogenix) and two modified tests using factor V depleted plasma: Coatest APC Resistance V (Chromogenix) and Accélérimat (BioMérieux). The last test is not influenced by the factor VIII by the adjunction of activated factor X. For each test, the coefficient of discrimination, between carrier and non-carrier of the R506Q mutation of the factor V gene, has been determined on 43 pregnant women (33 non-carriers and 11 heterozygotes) and 51 unselected non pregnant patients with clinically suspected thrombosis (40 non-carriers and 11 heterozygotes). The predilution of the patient's plasma with factor V deficient plasma (Coatest APC Resistance V and Accélérimat) enhances the discrimination between carriers and non-carriers in both groups. However, using Coatest APC Resistance V, a significant difference of results is observed between the two populations in the non-carriers patients. Thus, Accelerimat is probably more efficient than Coatest APC Resistance for the detection of the factor VQ506 during pregnancy.
Activated Protein C Resistance↗
Genetic variability in the renin-angiotensin system: prevalence of alleles and genotypes.
BACKGROUND: Variants of the human genes coding for renin, angiotensin-converting enzyme (ACE), angiotensinogen (AGT) and the angiotensin II type-I receptor (AT1R) are inconsistently associated with cardiovascular-renal disease, possibly because of genetic differences in the background populations. METHODS: This systematic review of the literature investigated genetic variation in the renin system according to race, sex and age. Across studies with relevant information, multivariate analyses also accounted for the methods of genotyping and the enrollment of subjects as controls, cases or groups studied cross-sectionally. RESULTS: The 185 reviewed reports included 64978 subjects. In five studies (n=989) on the renin gene, the Hind III and Taq I polymorphisms varied with the groups' average age, whereas the Bg I, Bg II and Hind III but not Taq I sites differed according to race. Among 135 studies (n=44697) on the deletion/insertion (D/I) polymorphism of the ACE gene, the frequency of the D allele was 54.0%. Its prevalence was not related to sex and black race, was 49.9% lower in Asians, 10.0% lower in studies relying on I-specific primers, 4.9% higher for each 25-year increment in the average age of the groups studied, and 16.7% higher in cases than controls. Among 12 studies (n=4952) on the T174-->M variant of the AGT gene, the M174 frequency was 11.0%, did not vary according to sex and enrollment group, was 56.7% lower in blacks and 39.5% lower for each 25-year increase in the groups' mean age. Across 44 studies (n=16713) on the M235-->T substitution in the AGT gene, the T235 prevalence was 51.6%. Its frequency was not related to sex and the method of genotyping, tended to be 7.5% lower for each 25-year increase in average age, was from 4.6 to 6.6 times higher in nonwhites than whites and 13.2% higher in cases than controls. Among 13 studies (n=4332) on the A1166-->C variant of the AT1R gene, the C1166 allelic frequency was 25.7%. Its prevalence was independent of the enrollment group, 77.4% lower in Asians, and nearly doubled for each 25-year increment in age. CONCLUSION: With adjustments applied for the subjects' enrollment group and the methods of genotyping, genetic polymorphism in the renin system varies according to race and age, but not sex. One possible application of the present results is to provide allelic and genotypic frequencies, which could be used to assess power, to perform sample size calculations, or to predict selection bias in future studies.
Age Factors↗