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Hassan Y. Naim - One of the best experts on this subject based on the ideXlab platform.
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Heterozygotes Are a Potential New Entity among Homozygotes and Compound Heterozygotes in Congenital Sucrase-Isomaltase Deficiency
Nutrients, 2019Co-Authors: Diab M. Husein, Klauspeter Zimmer, Dalanda Wanes, Lara M. Marten, Hassan Y. NaimAbstract:Congenital sucrase-Isomaltase deficiency (CSID) is an autosomal recessive disorder of carbohydrate maldigestion and malabsorption caused by mutations in the sucrase-Isomaltase (SI) gene. SI, together with maltase-glucoamylase (MGAM), belongs to the enzyme family of disaccharidases required for breakdown of α-glycosidic linkages in the small intestine. The effects of homozygote and compound heterozygote inheritance trait of SI mutations in CSID patients have been well described in former studies. Here we propose the inclusion of heterozygote mutation carriers as a new entity in CSID, possibly presenting with milder symptoms. The hypothesis is supported by recent observations of heterozygote mutation carriers among patients suffering from CSID or patients diagnosed with functional gastrointestinal disorders. Recent studies implicate significant phenotypic heterogeneity depending on the character of the mutation and call for more research regarding the correlation of genetics, function at the cellular and molecular level and clinical presentation. The increased importance of SI gene variants in irritable bowel syndrome (IBS) or other functional gastrointestinal disorders FGIDs and their available symptom relief diets like fermentable oligo-, di-, mono-saccharides and polyols FODMAPs suggest that the heterozygote mutants may affect the disease development and treatment.
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phylogenetic analysis reveals key residues in substrate hydrolysis in the Isomaltase domain of sucrase Isomaltase and its role in starch digestion
Biochimica et Biophysica Acta, 2019Co-Authors: Marcia M Chaudet, Hassan Y. Naim, Mahdi Amiri, Nathalie Marth, David R RoseAbstract:Abstract Background Starch constitutes one of the main sources of nutrition in the human diet and is broken down through a number of stages of digestion. Small intestinal breakdown of starch-derived substrates occurs through the mechanisms of small intestinal brush border enzymes, maltase-glucoamylase and sucrase-Isomaltase. These enzymes each contain two functional enzymatic domains, and though they share sequence and structural similarities due to their evolutionary conservation, they demonstrate distinct substrate preferences and catalytic efficiency. The N-terminal Isomaltase domain of sucrase-Isomaltase has a unique ability to actively hydrolyze isomaltose substrates in contrast to the sucrase, maltase and glucoamylase enzymes. Methods Through phylogenetic analysis, structural comparisons and mutagenesis, we were able to identify specific residues that play a role in the distinct substrate preference. Mutational analysis and comparison with wild-type activity provide evidence that this role is mediated in part by affecting interactions between the sucrase and Isomaltase domains in the intact molecule. Results The sequence analysis revealed three residues proposed to play key roles in Isomaltase specificity. Mutational analysis provided evidence that these residues in Isomaltase can also affect activity in the partner sucrase domain, suggesting a close interaction between the domains. Major conclusions The sucrase and Isomaltase domains are closely interacting in the mature protein. The activity of each is affected by the presence of the other. General Significance: There has been little experimental evidence previously of the effects on activity of interactions between the sucrase-Isomaltase enzyme domains. By extension, similar interactions might be expected in the other intestinal α-glucosidase, maltase-glucoamylase.
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structure function analysis of human sucrase Isomaltase identifies key residues required for catalytic activity
Journal of Biological Chemistry, 2017Co-Authors: Birthe Gericke, Mahdi Amiri, Natalie Schecker, Hassan Y. NaimAbstract:Abstract Sucrase-Isomaltase (SI) is an intestinal membrane-associated α-glucosidase that breaks down di- and oligosaccharides to absorbable monosaccharides. SI has two homologous functional subunits (sucrase and Isomaltase) that both belong to the glycoside hydrolase family 31 (GH31) and differ in substrate specificity. All GH31 enzymes share a consensus sequence harboring an aspartic acid residue as a catalytic nucleophile. Moreover, crystallographic structural analysis of Isomaltase predicts that another aspartic acid residue functions as a proton donor in hydrolysis. Here, we mutagenized the predicted proton donor residues and the nucleophilic catalyst residues in each SI subunit. We expressed these SI variants in COS-1 cells and analyzed their structural, transport, and functional characteristics. All of the mutants revealed expression levels and maturation rates comparable with those of the wild-type species and the corresponding nonmutated subunits were functionally active. Thereby we determined rate and substrate specificity for each single subunit without influence from the other subunit. This approach provides a model for functional analysis of the single subunits within a multidomain protein, achieved without the necessity to express the individual subunits separately. Of note, we also found that glucose product inhibition regulates the activities of both SI subunits. We experimentally confirmed the catalytic function of the predicted proton donor residues, and sequence analysis suggested that these residues are located in a consensus region in many GH31 family members. In summary, these findings reveal the kinetic features specific for each human SI subunit and demonstrate that the activities of these subunits are regulated via product inhibition.
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molecular pathogenicity of novel sucrase Isomaltase mutations found in congenital sucrase Isomaltase deficiency patients
Biochimica et Biophysica Acta, 2017Co-Authors: Birthe Gericke, Mahdi Amiri, Ronald C Scott, Hassan Y. NaimAbstract:Abstract Background & aims Congenital sucrase-Isomaltase deficiency (CSID) is a genetic disorder associated with mutations in the sucrase-Isomaltase ( SI ) gene. The diagnosis of congenital diarrheal disorders like CSID is difficult due to unspecific symptoms and usually requires invasive biopsy sampling of the intestine. Sequencing of the SI gene and molecular analysis of the resulting potentially pathogenic SI protein variants may facilitate a diagnosis in the future. This study aimed to categorize SI mutations based on their functional consequences. Methods cDNAs encoding 13 SI mutants were expressed in COS-1 cells. The molecular pathogenicity of the resulting SI mutants was defined by analyzing their biosynthesis, cellular localization, structure and enzymatic functions. Results Three biosynthetic phenotypes for the novel SI mutations were identified. The first biosynthetic phenotype was defined by mutants that are intracellularly transported in a fashion similar to wild type SI and with normal, but varying, levels of enzymatic activity. The second biosynthetic phenotype was defined by mutants with delayed maturation and trafficking kinetics and reduced activity. The third group of mutants is entirely transport incompetent and functionally inactive. Conclusions The current study unraveled CSID as a multifaceted malabsorption disorder that comprises three major classes of functional and trafficking mutants of SI and established a gradient of mild to severe functional deficits in the enzymatic functions of the enzyme. General significance This novel concept and the existence of mild consequences in a number of SI mutants strongly propose that CSID is an underdiagnosed and a more common intestinal disease than currently known.
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The multiple roles of sucrase-Isomaltase in the intestinal physiology
Molecular and Cellular Pediatrics, 2016Co-Authors: Birthe Gericke, Mahdi Amiri, Hassan Y. NaimAbstract:Osmotic diarrhea and abdominal pain in humans are oftentimes associated with carbohydrate malabsorption in the small intestine due to loss of function of microvillar disaccharidases. Disaccharidases are crucial for the digestion and the subsequent absorption of carbohydrates. This review focuses on sucrase-Isomaltase as the most abundant intestinal disaccharidase and the primary or induced pathological conditions that affect its physiological function. Congenital defects are primary factors which directly influence the transport and function of sucrase-Isomaltase in a healthy epithelium. Based on the mutation type and the pattern of inheritance, a mutation in the sucrase - Isomaltase gene may exert a variety of symptoms ranging from mild to severe. However, structure and function of wild type sucrase-Isomaltase can be also affected by secondary factors which influence its structure and function either specifically via certain inhibitors and therapeutic agents or generally as a part of intestinal pathogenesis, for example in the inflammatory responses. Diagnosis of sucrase-Isomaltase deficiency and discriminating it from other gastrointestinal intolerances can be latent in the patients because of common symptoms observed in all of these cases. Here, we summarize the disorders that implicate the digestive function of sucrase-Isomaltase as well as the diagnostic and therapeutic strategies utilized to restore normal intestinal function.
Buford L Nichols - One of the best experts on this subject based on the ideXlab platform.
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metabolic impacts of maltase deficiencies
Journal of Pediatric Gastroenterology and Nutrition, 2018Co-Authors: Buford L Nichols, Susan S Baker, Roberto QuezadacalvilloAbstract:: The mucosal maltase enzymes are characterized by an activity that produces glucose from linear glucose polymers, assayed with the disaccharide maltose. The related enzyme Isomaltase produces glucose from branched glucose polymers, assayed with palatinose. Maltase and Isomaltase activities are part of the 4 disaccharidases assayed from clinical duodenal biopsy homogenates. The reported maltase activities are more difficult to interpret than lactase or sucrase activities because both the sucrase-Isomaltase and maltase-glucoamylase proteins have overlapping maltase activities. The early work of Dahlqvist identified 4 maltase activities from human small intestinal mucosa. On one peptide, sucrase (maltase Ib) and Isomaltase (maltase Ia) activities shared maltase activities but identified the enzymes as sucrase-Isomaltase. On the other peptide, no distinguishing characteristics of the 2 maltase activities (maltases II and III) were detected and the activities identified as maltase-glucoamylase. The nutritional/clinical importance of small intestinal maltase and Isomaltase activities are due to their crucial role in the digestion of food starches to absorbable free glucose. This review focuses on the interpretation of biopsy maltase activities in the context of reported lactase, sucrase, maltase, and palatinase biopsy assay activity patterns. We present a classification of mucosal maltase deficiencies and novel primary maltase deficiency (Ib, II, III) and provide a clarification of the role of maltase activity assayed from clinically obtained duodenal biopsies, as a path toward future clinical and molecular genomic investigations.
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13c labeled starch breath test in congenital sucrase Isomaltase deficiency
Journal of Pediatric Gastroenterology and Nutrition, 2018Co-Authors: Claudia C Robayotorres, Susan S Baker, Bruce R Hamaker, Antone R. Opekun, Marisela Diazsotomayor, Bruno P Chumpitazi, Buford L NicholsAbstract:ABSTRACTBackground and Hypotheses:Human starch digestion is a multienzyme process involving 6 different enzymes: salivary and pancreatic α-amylase; sucrase and Isomaltase (from sucrose-Isomaltase [SI]), and maltase and glucoamylase (from maltase-glucoamylase [MGAM]). Together these enzymes cleave st
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improved starch digestion of sucrase deficient shrews treated with oral glucoamylase enzyme supplements
Journal of Pediatric Gastroenterology and Nutrition, 2017Co-Authors: Buford L Nichols, Stephen E Avery, Roberto Quezadacalvillo, Shadi B Kilani, Amy Huimei Lin, Douglas G Burrin, Benjamin E Hodges, Shaji K ChackoAbstract:BACKGROUND AND OBJECTIVE Although named because of its sucrose hydrolytic activity, this mucosal enzyme plays a leading role in starch digestion because of its maltase and glucoamylase activities. Sucrase-deficient mutant shrews, Suncus murinus, were used as a model to investigate starch digestion in patients with congenital sucrase-Isomaltase deficiency.Starch digestion is much more complex than sucrose digestion. Six enzyme activities, 2 α-amylases (Amy), and 4 mucosal α-glucosidases (maltases), including maltase-glucoamylase (Mgam) and sucrase-Isomaltase (Si) subunit activities, are needed to digest starch to absorbable free glucose. Amy breaks down insoluble starch to soluble dextrins; mucosal Mgam and Si can either directly digest starch to glucose or convert the post-α-amylolytic dextrins to glucose. Starch digestion is reduced because of sucrase deficiency and oral glucoamylase enzyme supplement can correct the starch maldigestion. The aim of the present study was to measure glucogenesis in suc/suc shrews after feeding of starch and improvement of glucogenesis by oral glucoamylase supplements. METHODS Sucrase mutant (suc/suc) and heterozygous (+/suc) shrews were fed with C-enriched starch diets. Glucogenesis derived from starch was measured as blood C-glucose enrichment and oral recombinant C-terminal Mgam glucoamylase (M20) was supplemented to improve starch digestion. RESULTS After feedings, suc/suc and +/suc shrews had different starch digestions as shown by blood glucose enrichment and the suc/suc had lower total glucose concentrations. Oral supplements of glucoamylase increased suc/suc total blood glucose and quantitative starch digestion to glucose. CONCLUSIONS Sucrase deficiency, in this model of congenital sucrase-Isomaltase deficiency, reduces blood glucose response to starch feeding. Supplementing the diet with oral recombinant glucoamylase significantly improved starch digestion in the sucrase-deficient shrew.
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protein synthesis controls the activity of maltase glucoamylase and sucrase Isomaltase in non intestinal tissues
The FASEB Journal, 2015Co-Authors: Meric Simsek, Roberto Quezadacalvillo, Jacquelin Juarez, Buford L NicholsAbstract:Maltase-Glucoamylase (MGAM) and Sucrase-Isomaltase (SI) are α-glucosidases of the membrane of small intestinal enterocytes responsible for the digestion of dietary carbohydrates into glucose and other monosaccharides. MGAM and SI mRNA expression, protein synthesis and catalytic activity of MGAM and SI are found predominantly in enterocytes; however, they are also found in lower amounts in other non-intestinal tissues, notably blood leukocytes and kidney cells. Little information exists on the functions, the mRNA expression, the nature of the respective proteins and catalytic properties of MGAM and SI in non-intestinal tissues. To correlate the transcription, synthesis and activity of these enzymes, we measured the mRNA expression of MGAM and SI; the catalytic activities of maltase, sucrase, and Isomaltase; and the presence of the respective proteins in small intestine and other tissues such as spleen, kidney and pancreas of 8 weeks old mice. Small intestine showed the highest level of mRNA expression of M...
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inhibition of individual subunits of maltase glucoamylase and sucrase Isomaltase by polyphenols 1045 26
The FASEB Journal, 2014Co-Authors: Meric Simsek, Roberto Quezadacalvillo, Buford L Nichols, Bruce R HamakerAbstract:Some polyphenols inhibit the activities of the four subunits of intestinal α-glucosidases, maltase-glucoamylase (Mgam) and sucrase-Isomaltase (Si). The use of polyphenols as modulators of the rate ...
Roberto Quezadacalvillo - One of the best experts on this subject based on the ideXlab platform.
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metabolic impacts of maltase deficiencies
Journal of Pediatric Gastroenterology and Nutrition, 2018Co-Authors: Buford L Nichols, Susan S Baker, Roberto QuezadacalvilloAbstract:: The mucosal maltase enzymes are characterized by an activity that produces glucose from linear glucose polymers, assayed with the disaccharide maltose. The related enzyme Isomaltase produces glucose from branched glucose polymers, assayed with palatinose. Maltase and Isomaltase activities are part of the 4 disaccharidases assayed from clinical duodenal biopsy homogenates. The reported maltase activities are more difficult to interpret than lactase or sucrase activities because both the sucrase-Isomaltase and maltase-glucoamylase proteins have overlapping maltase activities. The early work of Dahlqvist identified 4 maltase activities from human small intestinal mucosa. On one peptide, sucrase (maltase Ib) and Isomaltase (maltase Ia) activities shared maltase activities but identified the enzymes as sucrase-Isomaltase. On the other peptide, no distinguishing characteristics of the 2 maltase activities (maltases II and III) were detected and the activities identified as maltase-glucoamylase. The nutritional/clinical importance of small intestinal maltase and Isomaltase activities are due to their crucial role in the digestion of food starches to absorbable free glucose. This review focuses on the interpretation of biopsy maltase activities in the context of reported lactase, sucrase, maltase, and palatinase biopsy assay activity patterns. We present a classification of mucosal maltase deficiencies and novel primary maltase deficiency (Ib, II, III) and provide a clarification of the role of maltase activity assayed from clinically obtained duodenal biopsies, as a path toward future clinical and molecular genomic investigations.
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improved starch digestion of sucrase deficient shrews treated with oral glucoamylase enzyme supplements
Journal of Pediatric Gastroenterology and Nutrition, 2017Co-Authors: Buford L Nichols, Stephen E Avery, Roberto Quezadacalvillo, Shadi B Kilani, Amy Huimei Lin, Douglas G Burrin, Benjamin E Hodges, Shaji K ChackoAbstract:BACKGROUND AND OBJECTIVE Although named because of its sucrose hydrolytic activity, this mucosal enzyme plays a leading role in starch digestion because of its maltase and glucoamylase activities. Sucrase-deficient mutant shrews, Suncus murinus, were used as a model to investigate starch digestion in patients with congenital sucrase-Isomaltase deficiency.Starch digestion is much more complex than sucrose digestion. Six enzyme activities, 2 α-amylases (Amy), and 4 mucosal α-glucosidases (maltases), including maltase-glucoamylase (Mgam) and sucrase-Isomaltase (Si) subunit activities, are needed to digest starch to absorbable free glucose. Amy breaks down insoluble starch to soluble dextrins; mucosal Mgam and Si can either directly digest starch to glucose or convert the post-α-amylolytic dextrins to glucose. Starch digestion is reduced because of sucrase deficiency and oral glucoamylase enzyme supplement can correct the starch maldigestion. The aim of the present study was to measure glucogenesis in suc/suc shrews after feeding of starch and improvement of glucogenesis by oral glucoamylase supplements. METHODS Sucrase mutant (suc/suc) and heterozygous (+/suc) shrews were fed with C-enriched starch diets. Glucogenesis derived from starch was measured as blood C-glucose enrichment and oral recombinant C-terminal Mgam glucoamylase (M20) was supplemented to improve starch digestion. RESULTS After feedings, suc/suc and +/suc shrews had different starch digestions as shown by blood glucose enrichment and the suc/suc had lower total glucose concentrations. Oral supplements of glucoamylase increased suc/suc total blood glucose and quantitative starch digestion to glucose. CONCLUSIONS Sucrase deficiency, in this model of congenital sucrase-Isomaltase deficiency, reduces blood glucose response to starch feeding. Supplementing the diet with oral recombinant glucoamylase significantly improved starch digestion in the sucrase-deficient shrew.
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protein synthesis controls the activity of maltase glucoamylase and sucrase Isomaltase in non intestinal tissues
The FASEB Journal, 2015Co-Authors: Meric Simsek, Roberto Quezadacalvillo, Jacquelin Juarez, Buford L NicholsAbstract:Maltase-Glucoamylase (MGAM) and Sucrase-Isomaltase (SI) are α-glucosidases of the membrane of small intestinal enterocytes responsible for the digestion of dietary carbohydrates into glucose and other monosaccharides. MGAM and SI mRNA expression, protein synthesis and catalytic activity of MGAM and SI are found predominantly in enterocytes; however, they are also found in lower amounts in other non-intestinal tissues, notably blood leukocytes and kidney cells. Little information exists on the functions, the mRNA expression, the nature of the respective proteins and catalytic properties of MGAM and SI in non-intestinal tissues. To correlate the transcription, synthesis and activity of these enzymes, we measured the mRNA expression of MGAM and SI; the catalytic activities of maltase, sucrase, and Isomaltase; and the presence of the respective proteins in small intestine and other tissues such as spleen, kidney and pancreas of 8 weeks old mice. Small intestine showed the highest level of mRNA expression of M...
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inhibition of individual subunits of maltase glucoamylase and sucrase Isomaltase by polyphenols 1045 26
The FASEB Journal, 2014Co-Authors: Meric Simsek, Roberto Quezadacalvillo, Buford L Nichols, Bruce R HamakerAbstract:Some polyphenols inhibit the activities of the four subunits of intestinal α-glucosidases, maltase-glucoamylase (Mgam) and sucrase-Isomaltase (Si). The use of polyphenols as modulators of the rate ...
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different sucrose Isomaltase response of caco 2 cells to glucose and maltose suggests dietary maltose sensing
Journal of Clinical Biochemistry and Nutrition, 2014Co-Authors: Minwen Cheng, Roberto Quezadacalvillo, Buford L Nichols, Mohammad Chegeni, Keehong Kim, Genyi Zhang, Mustapha Benmoussa, Bruce R HamakerAbstract:Using the small intestine enterocyte Caco-2 cell model, sucrase-Isomaltase (SI, the mucosal α-glucosidase complex) expression and modification were examined relative to exposure to different mono- and disaccharide glycemic carbohydrates. Caco-2/TC7 cells were grown on porous supports to post-confluence for complete differentiation, and dietary carbohydrate molecules of glucose, sucrose (disaccharide of glucose and fructose), maltose (disaccharide of two glucoses α-1,4 linked), and isomaltose (disaccharide of two glucoses α-1,6 linked) were used to treat the cells. qRT-PCR results showed that all the carbohydrate molecules induced the expression of the SI gene, though maltose (and isomaltose) showed an incremental increase in mRNA levels over time that glucose did not. Western blot analysis of the SI protein revealed that only maltose treatment induced a higher molecular weight band (Mw ~245 kDa), also at higher expression level, suggesting post-translational processing of SI, and more importantly a sensing of maltose. Further work is warranted regarding this putative sensing response as a potential control point for starch digestion and glucose generation in the small intestine.
Matthew A Lines - One of the best experts on this subject based on the ideXlab platform.
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mg 107 congenital sucrase Isomaltase deficiency identification of the common inuit founder mutation
Journal of Medical Genetics, 2015Co-Authors: Julien L Marcadier, Margaret Boland, Ronald C Scott, Kheirie Issa, Adam D Mcintyre, Robert A Hegele, Michael T Geraghty, Matthew A LinesAbstract:Objective Congenital sucrase-Isomaltase deficiency (CSID) is a rare hereditary cause of chronic diarrhoea in children. Persons with CSID lack the intestinal brush-border enzyme required for digestion of di- and oligosaccharides, including sucrose and isomaltose. Malabsorption results in abdominal pain, distention, copious diarrhoea, and failure to thrive. If recognised, dietary avoidance of the offending carbohydrates is highly effective. Although CSID is known to be highly prevalent (˜5–10%) in several Inuit populations, the genetic basis for this condition has not been described. Methods We sequenced the sucrase-Isomaltase gene, SI, in a single Inuit CSID proband with severe fermentative diarrhoea and failure to thrive. We then genotyped a further 128 anonymized Inuit control individuals from a variety of circumpolar locales to assess for a possible founder effect. Results We identified a novel, homozygous frameshift mutation, c.273_274delAG (p. Gly92Leufs*8) in exon 4 of SI in the proband, that is predicted to result in complete absence of functional protein product. This change is indeed very common among Inuit control specimens, with an observed allele frequency of 0.17 (95% confidence interval 0.13–0.22). The predicted Hardy-Weinberg prevalence of CSID in the Inuit, based on this single founder allele is ˜3% (95% confidence interval 1.6–5.0%), comparable with previous estimates. Interpretation Targeted mutation testing for the c.273_274delAG allele should afford a simple and minimally invasive means of diagnosing CSID in persons of Inuit descent. Because CSID is a readily treatable disorder, such testing should be considered at an early stage in the assessment of Inuit patients with chronic diarrhoea.
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congenital sucrase Isomaltase deficiency identification of a common inuit founder mutation
Canadian Medical Association Journal, 2015Co-Authors: Julien L Marcadier, Margaret Boland, Ronald C Scott, Kheirie Issa, Adam D Mcintyre, Robert A Hegele, Michael T Geraghty, Matthew A LinesAbstract:Background Congenital sucrase-Isomaltase deficiency is a rare hereditary cause of chronic diarrhea in children. People with this condition lack the intestinal brush-border enzyme required for digestion of di- and oligosaccharides, including sucrose and isomaltose, leading to malabsorption. Although the condition is known to be highly prevalent (about 5%-10%) in several Inuit populations, the genetic basis for this has not been described. We sought to identify a common mutation for congenital sucrase-Isomaltase deficiency in the Inuit population. Methods We sequenced the sucrase-Isomaltase gene, SI, in a single Inuit proband with congenital sucrase-Isomaltase deficiency who had severe fermentative diarrhea and failure to thrive. We then genotyped a further 128 anonymized Inuit controls from a variety of locales in the Canadian Arctic to assess for a possible founder effect. Results In the proband, we identified a novel, homozygous frameshift mutation, c.273_274delAG (p.Gly92Leufs*8), predicted to result in complete absence of a functional protein product. This change was very common among the Inuit controls, with an observed allele frequency of 17.2% (95% confidence interval [CI] 12.6%-21.8%). The predicted Hardy-Weinberg prevalence of congenital sucrase-Isomaltase deficiency in Inuit people, based on this single founder allele, is 3.0% (95% CI 1.4%-4.5%), which is comparable with previous estimates. Interpretation We found a common mutation, SI c.273_274delAG, to be responsible for the high prevalence of congenital sucrase-Isomaltase deficiency among Inuit people. Targeted mutation testing for this allele should afford a simple and minimally invasive means of diagnosing this condition in Inuit patients with chronic diarrhea.
Monique Rousset - One of the best experts on this subject based on the ideXlab platform.
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differential expression of sucrase Isomaltase in clones isolated from early and late passages of the cell line caco 2 evidence for glucose dependent negative regulation
Journal of Cell Science, 1994Co-Authors: Isabelle Chantret, E. Dussaulx, Alain Barbat, Alain Zweibaum, Annie Rodolosse, Edith Brotlaroche, Monique RoussetAbstract:The expression of the brush border-associated hydrolase sucrase-Isomaltase was shown to increase from early to late passages of Caco-2 cells, concomitant with a decrease in the rates of glucose consumption. Twenty-six clones were isolated from early (P29) and late (P198) passages of the cell line. These clones show considerable and inverse differences in the levels of sucrase activities and rates of glucose consumption, without marked changes in other features of enterocytic differentiation of the cells (presence of an apical brush border, levels of expression of other brush border-associated hydrolases). Clones with low sucrase-Isomaltase expression show a mosaic expression of the enzyme and a 38-fold higher rate of glucose consumption than clones with high sucrase-Isomaltase expression. The clones with high expression show an homogeneous apical distribution of the enzyme and 70-fold and 35-fold higher levels of sucrase activities and sucrase-Isomaltase mRNA, respectively. In contrast no differences were found from one clone to another in the enrichment of sucrase activity in brush border-enriched fractions as compared to cell homogenates. Switch to low glucose-containing medium (1 mM versus 25 mM in standard culture conditions) of cells with low sucrase-Isomaltase results in an increased and more homogeneous expression of the enzyme and a tenfold augmentation of the levels of sucrase-Isomaltase mRNA and sucrase activity. These results show that glucose interferes with the expression of sucrase-Isomaltase in Caco-2 cells at the mRNA level.
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sequence of the complete cdna and the 5 structure of the human sucrase Isomaltase gene possible homology with a yeast glucoamylase
Biochemical Journal, 1992Co-Authors: I Chantret, Ned Mantei, Michel Lacasa, G Chevalier, J Ruf, I Islam, Yvonne J K Edwards, Dallas M Swallow, Monique RoussetAbstract:The complete sequence of the 6 kb cDNA and the 5' genomic structure are reported for the gene coding for the human intestinal brush border hydrolase sucrase-Isomaltase. The human sucrase-Isomaltase cDNA shows a high level of identity (83%) with that of the rabbit enzyme, indicating that the protein shares the same structural domains in both species. In addition to the previously reported homology with lysosomal alpha-glucosidase, the sucrase and Isomaltase subunits also appear to be homologous to a yeast glucoamylase. A 14 kb human genomic clone has been isolated which includes the first three exons and the first two introns of the gene, as well as 9.5 kb 5' to the major start site of transcription. The first exon comprises 62 bp of untranslated sequence and the second starts exactly at the initiation ATG codon. Typical CAAT and TATA boxes are seen upstream of the first exon. A genetic polymorphism is described which involves a PstI site in the second intron. Southern blotting, sequencing and mRNA studies indicate that the structures of the sucrase-Isomaltase gene and its mRNA are unaltered in the two human colon cancer cell lines Caco-2 and HT-29 in comparison with normal human small intestine.
