Localization of human homeodomain transcription factor insulin promoter factor 1 (IPF1) to chromosome band 13q12.1.
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Base Sequence↗
Biomedical subjects
G I Bell
Publications and source records attributed to G I Bell.
Human fructose-1,6-bisphosphatase gene (FBP1): exon-intron organization, localization to chromosome bands 9q22.2-q22.3, and mutation screening in subjects with fructose-1,6-bisphosphatase deficiency.
Fructose-1,6-bisphosphatase (EC 3.1.3.11) is a key regulatory enzyme of gluconeogenesis that catalyzes the hydrolysis of fructose-1,6-bisphosphate to generate fructose-6-phosphate and inorganic phosphate. Deficiency of fructose-1,6-bisphosphatase is associated with fasting hypoglycemia and metabolic acidosis because of impaired gluconeogenesis. We have cloned and characterized the human liver fructose-1,6-bisphosphatase gene (FBP1). FBP1, localized to chromosome bands 9q22.2-q22.3 by fluorescence in situ hybridization, consists of seven exons that span > 31 kb, and the six introns are in the same position as in the rat gene. FBP1 was screened for mutations in two subjects with fructose-1,6-bisphosphatase deficiency. Four nucleotide substitutions were identified, two of which were silent mutations in the codons for Ala-216 (GCT-->GCC) and Gly-319 (GGG-->GGA). The other substitutions were in intron 3, a C-->T substitution 7 nucleotides downstream from the splice donor site, and in the promoter region, an A-->T substitution 188 nucleotides upstream from the start of transcription. These nucleotide substitutions were also found in normal unaffected subjects and thus are not the cause of fructose-1,6-bisphosphatase deficiency in the two subjects studied. The molecular basis of hepatic fructose-1,6-bisphosphatase deficiency in these subjects remains undetermined but could result from unidentified mutations in the promoter that decrease expression or from mutations in another gene that indirectly lead to decreased fructose-1,6-bisphosphatase activity.
Amino Acid Sequence↗
Cloning of rat and mouse P2Y purinoceptors.
The cellular responses to ATP are mediated by specific high-affinity receptors designated as P2 purinoceptors, five subclasses of which have been defined pharmacologically-P2X, P2Y, P2U, P2T, and P2Z. A cDNA clone encoding a rat P2Y purinoceptor was isolated from an insulinoma cDNA library. The 373-amino acid rat P2Y purinoceptor sequence has 85.7% and 37.8% identity with chicken P2Y and rat P2U purinoceptors, respectively. The sequence of the mouse P2Y purinoceptor was also determined and there was 97.1% amino acid identity with the corresponding rat sequence. RNA blotting studies showed that rat P2Y purinoceptor mRNA was expressed at variable levels in many tissues including heart, brain, spleen, lung, liver, skeletal muscle and kidney, although it was not detected in testis. The cloned rat P2Y purinoceptor was expressed in Xenopus laevis oocytes and possessed the properties expected for this receptor subtype.
Adenosine Triphosphate↗
Localization of human somatostatin receptor 5 gene (SSTR5) to chromosome band 16p13.3 by fluorescence in situ hybridization.
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Chromosome Mapping↗
Inhibition of cell proliferation by the somatostatin analogue RC-160 is mediated by somatostatin receptor subtypes SSTR2 and SSTR5 through different mechanisms.
Effects of the stable somatostatin analogue RC-160 on cell proliferation, tyrosine phosphatase activity, and intracellular calcium concentration were investigated in CHO cells expressing the five somatostatin receptor subtypes SSTR1 to -5. Binding experiments were performed on crude membranes by using [125I-labeled Tyr11] somatostatin-14; RC-160 exhibited moderate-to-high affinities for SSTR2, -3, and -5 (IC50, 0.17, 0.1 and 21 nM, respectively) and low affinity for SSTR1 and -4 (IC50, 200 and 620 nM, respectively). Cell proliferation was induced in CHO cells by 10% (vol/vol) fetal calf serum, 1 microM insulin, or 0.1 microM cholecystokinin (CCK)-8; RC-160 inhibited serum-induced proliferation of CHO cells expressing SSTR2 and SSTR5 (EC50, 53 and 150 pM, respectively) but had no effect on growth of cells expressing SSTR1, -3, or -4. In SSTR2-expressing cells, orthovanadate suppressed the growth inhibitory effect of RC-160. This analogue inhibited insulin-induced proliferation and rapidly stimulated the activity of a tyrosine phosphatase in only this cellular clone. This latter effect was observed at doses of RC-160 (EC50, 4.6 pM) similar to those required to inhibit growth (EC50, 53 pM) and binding to the receptor (IC50, 170 pM), implicating tyrosine phosphatase as a transducer of the growth inhibition signal in SSTR2-expressing cells. In SSTR5-expressing cells, the phosphatase pathway was not involved in the inhibitory effect of RC-160 on cell growth, since this action was not influenced by tyrosine and serine/threonine phosphatase inhibitors. In addition, in SSTR5-expressing cells, RC-160 inhibited CCK-stimulated intracellular calcium mobilization at doses (EC50, 0.35 nM) similar to those necessary to inhibit somatostatin-14 binding (IC50, 21 nM) and CCK-induced cell proliferation (EC50, 1.1 nM). This suggests that the inositol phospholipid/calcium pathway could be involved in the antiproliferative effect of RC-160 mediated by SSTR5 in these cells. RC-160 had no effect on the basal or carbachol-stimulated calcium concentration in cells expressing SSTR1 to -4. Thus, we conclude that SSTR2 and SSTR5 bind RC-160 with high affinity and mediate the RC-160-induced inhibition of cell growth by distinct mechanisms.
Amino Acid Sequence↗
A tyrosine phosphatase is associated with the somatostatin receptor.
Regulation of tyrosine phosphorylation is thought to be an essential step in signal transduction mechanisms that mediate cellular responses. In pancreatic tumour cells we demonstrated that somatostatin analogues inhibited cell proliferation and stimulated a membrane protein tyrosine phosphatase (PTP) activity at concentrations at which they bind to the somatostatin receptor. To elucidate the role of PTP in the signal transduction pathway activated by somatostatin receptors we first studied the interaction of PTP with the somatostatin receptor at the membrane. We purified somatostatin receptors by immunoaffinity from pancreatic membranes that strongly expressed the type 2 somatostatin receptor sstr2. We identified the receptor as an 87 kDa protein. We demonstrated that a PTP activity co-purified with somatostatin receptors. The PTP was identified as a 66 kDa protein immunoreactive to antibodies against SHPTP1. These antibodies immunoprecipitated somatostatin receptors either occupied or unoccupied by ligand indicating that SHPTP1 is associated with somatostatin receptors. We then expressed sstr2A in monkey kidney COS-7 cells and mouse NIH/3T3 fibroblasts and demonstrated that somatostatin analogues (RC 160, octreotide and BIM 23014) which exhibited high affinity for sstr2 stimulated a PTP activity and inhibited cell proliferation in proportion to their affinities for sstr2. Under the same conditions these analogues have no effect on the growth of cells expressing sstr1. All these results suggest that a PTP related to SHPTP1 is associated with somatostatin receptors and may be involved in the negative growth signal promoted by sstr2.
3T3 Cells↗
Molecular biology of somatostatin receptors.
The diverse physiological effects of somatostatin are mediated by a family of cell surface receptors that bind somatostatin selectively and with high affinity. The somatostatin receptors are members of the seven transmembrane segment receptor superfamily and molecular cloning studies have identified five types, designated sstr1-5. The human somatostatin receptors vary in size from 364 (sstr5) to 418 (sstr3) amino acids with 46-61% amino acid identity between receptors, and 105 amino acids are invariant. The sequences of the seven putative alpha-helical membrane-spanning domains are more highly conserved than those of the extracellular N- and intracellular C-terminal domains. Two forms of sstr2 have been identified in the mouse, sstr2A and sstr2B, which differ in size and sequence of the intracellular C-terminal domain. These two forms of sstr2 are products of a common gene and are generated by alternative splicing with sstr2A and sstr2B being the products of the unspliced and spliced forms, respectively, of sstr2 mRNA. Thus, functional diversity within the somatostatin receptor family may result from the expression of multiple types as well as from alternative splicing. The five somatostatin receptors have distinct patterns of expression in the central nervous system and peripheral tissues. They have also been expressed in vitro and shown to have different pharmacological properties. Somatostatin analogues selective for sstr2, sstr3 and sstr5 have been identified which will facilitate in vivo studies of the functions of these somatostatin receptors. Such studies to date suggest that sstr2 mediates inhibition of growth hormone secretion and sstr5 mediates inhibition of insulin secretion. The molecular cloning and functional characterization of the somatostatin receptor family is a first step in elucidating the diverse effects of somatostatin on cellular functions.
Amino Acid Sequence↗
Identification of two novel amino acid polymorphisms in beta-cell/liver (GLUT2) glucose transporter in Japanese subjects.
The beta-cell/liver glucose transporter (GLUT2) gene was screened for mutations using single-strand conformation polymorphism analysis (SSCP) in 30 Japanese subjects with non-insulin dependent diabetes mellitus (NIDDM). Analysis of all exons and adjacent intron regions identified six SSCP polymorphisms, three of which resulted in amino acid substitutions: V101I, T110I and G519E. The V101I and G519E, substitutions represent new polymorphisms in this gene. The six polymorphisms were observed in both NIDDM and control groups and there were no significant differences in allele frequencies between groups. A portion of the insulin receptor substrate 1 gene in 30 NIDDM subjects and in normal control subjects was also screened for mutations. Two SSCP variants that change the sequence of the protein, delta S686/687 (deletion of the codons for serine-686 and 687) and G972R, were identified in two different NIDDM subjects, both whom were also heterozygous for the V101I polymorphisms in GLUT2. The GLUT2 and IRS1 amino acid polymorphisms did not show a simple pattern of co-inheritance with NIDDM in the families of these subjects suggesting that neither polymorphism is sufficient to cause NIDDM but may increase diabetes-susceptibility through their interaction with other loci and environmental factors.
Amino Acid Sequence↗
Human hexokinase II gene: exon-intron organization, mutation screening in NIDDM, and its relationship to muscle hexokinase activity.
In muscle, hexokinase II (HK2) regulates phosphorylation of glucose to glucose 6-phosphate, which has been reported to be impaired in patients with non-insulin-dependent diabetes mellitus (NIDDM). Here we report decreased HK2 enzyme activity in skeletal muscle biopsies from patients with impaired glucose tolerance compared with healthy control subjects (2.7 +/- 0.9 vs 4.9 +/- 1.1 nmol.min-1.mg protein-1). Therefore, mutations in the HK2 gene could contribute to skeletal muscle insulin resistance in NIDDM. To address this question, we first determined the exon-intron structure of the human HK2 gene and using this information, we screened all 18 exons with single-strand conformation polymorphism technique in 80 Finnish NIDDM patients. Nine nucleotide substitutions were found, one of which was a missense mutation (Gln142-His142) in exon 4. In human muscle, a single HK2 mRNA transcript with a size of approximately 5500 nucleotides was detected with Northern blot analysis. We also describe an HK2 pseudogene (HK2P1), which was mapped to chromosome 4, band q26, by fluorescence in situ hybridization to metaphase chromosomes. The clinical characteristics and HK2 enzyme activities of the subjects with either Gln or His at residue 142 did not differ from each other. Instead, HK2 activity correlated inversely with fasting blood glucose levels, suggesting that changes in HK2 activity could be secondary to other metabolic abnormalities (r = 0.55; p < 0.0003; n = 39). In conclusion; the data suggest that impaired HK2 activity in prediabetic individuals is a consequence of impaired glucose tolerance rather than of a genetic abnormality. The data thus seem to rule out mutations in the HK2 gene as a major cause of inherited insulin resistance in NIDDM.
Aged↗
Linkage studies in NIDDM with markers near the sulphonylurea receptor gene.
The high affinity receptor for sulphonylureas, expressed on the beta cells of the pancreas, plays a crucial role in the control of insulin secretion. Mutations in the cytoplasmic domain of the sulphonylurea receptor (SUR) gene that disrupt the regulation of insulin secretion have been previously described. In the present study, the potential role of genetic variation in the SUR gene has been investigated in non-insulin-dependent diabetes mellitus (NIDDM) through linkage studies with microsatellite markers tightly linked to the SUR gene. The microsatellite markers were typed in 346 Mexican-American NIDDM affected sib pairs derived from 176 families and an additional 110 ethnically and geographically matched control subjects. No evidence of linkage, based on allele sharing, or association based on allele frequencies in patients and control subjects, for any microsatellite marker and NIDDM was observed in this population. These results suggest that genetic variation in the SUR gene does not play a major role in susceptibility to NIDDM in the Mexican-American population.
ATP-Binding Cassette Transporters↗
Candidate gene studies in pedigrees with maturity-onset diabetes of the young not linked with glucokinase.
Maturity-onset diabetes of the young (MODY) is a form of non-insulin-dependent diabetes mellitus characterised by an early age of onset and an autosomal dominant mode of inheritance. Only a proportion of cases are due to mutations in the glucokinase gene. We have studied five Caucasian MODY families, including the first MODY family to be described, with five candidate genes implicated in regulation of insulin secretion. The affected subjects showed more marked hyperglycaemia than that found in subjects with glucokinase mutations. We assessed polymorphic markers close to the genes for glucokinase, hexokinase II, adenosine deaminase, pituitary adenylate cyclase-activating polypeptide receptor, and glucagon-like peptide-1 receptor. Linkage analysis with diabetes gave cumulative log of the odds (LOD) scores of less than -3, implying that mutations in these genes are unlikely to provide a major genetic contribution to this form of MODY.
Adolescent↗
Characterization of Japanese families with early-onset type 2 (non-insulin dependent) diabetes mellitus and screening for mutations in the glucokinase and mitochondrial tRNA(Leu(UUR)) genes.
Genetic linkage studies of families with early-onset type 2 diabetes have facilitated the identification of diabetes-susceptibility genes. In order to assess the feasibility of using linkage approaches to identify genes responsible for the development of type 2 diabetes in Japanese subjects, we examined our clinical records for multigenerational families suitable for genetic studies. We identified 16 families in which at least one subject was diagnosed with type 2 diabetes before 25 years of age. Seven of these families had a pattern of inheritance consistent with a diagnosis of maturity-onset diabetes of the young (MODY) and nine families showed a complex pattern of inheritance of type 2 diabetes with transmission of diabetes-susceptibility genes from both parents. The glucokinase and mitochondrial tRNA(Leu(UUR)) genes were screened for mutations in at least one affected subject from each family in order to assess the contribution of mutations in these genes to the development of the diabetes. No mutations were found, which suggests that the diabetes in these families resulted from mutations in other genes.
Adolescent↗
Inositol 1,4,5-trisphosphate receptors, secretory granules and secretion in endocrine and neuroendocrine cells.
Recent studies have revealed the presence of inositol 1,4,5-trisphosphate receptors in the secretory granules of endocrine and neuroendocrine cells. This distribution suggests that inositol 1,4,5-trisphosphate-regulated release of granule stores of Ca2+ might facilitate the secretory process. In addition, inositol 1,4,5-trisphosphate receptors might participate directly in the biogenesis of secretory granules. The presence of inositol 1,4,5-trisphosphate receptors in synaptic nerve terminals raises the possibility that they might also be involved in the control of neurotransmitter release.
Animals↗
Molecular properties of somatostatin receptors.
The neuropeptide somatostatin is widely distributed in the central nervous system and in peripheral tissues and may be involved in the regulation of a number of physiological functions including movement and cognition. Somatostatin may also have a role in the development of the central nervous system, in particular, the cerebellum and spinal cord. Somatostatin induces its actions by interacting with a family of membrane associated receptors. Recently, five somatostatin receptors have been cloned and referred to as SSTR1-SSTR5. The distribution of the expression of the mRNAs for these receptors are distinct but overlapping. Preliminary pharmacological analysis of these receptors may lead to the development of selective ligands at these receptors. These compounds may be useful in identifying the selective functions of these receptor subtypes. Some somatostatin analogues have antiproliferative actions and are used presently to treat carcinoids. Development of subtype selective somatostatin analogues could be helpful in further identifying somatostatin receptor-expressing tumors and in the treatment of cancer. The cloning of these receptors has now opened up the possibility of more clearly investigating the functions of somatostatin in the brain and peripheral tissues and will facilitate the generation of new somatostatin drugs that may be employed for the treatment of a number of diseases.
Animals↗
Classification and nomenclature of somatostatin receptors.
There is considerable controversy about the classification and nomenclature of somatostatin receptors. To date, five distinct receptor genes have been cloned and named chronologically according to their respective publication dates, but two were unfortunately given the same appellation (SSTR4). Consensually, a nomenclature for the recombinant receptors has been agreed according to IUPHAR guidelines (sst1, sst2, sst3, sst4, and sst5). However, a more informative classification is to be preferred for the future, employing all classification criteria in an integrated scheme. It is already apparent that the five recombinant receptors fall into two classes or groups, on the basis of not only structure but also pharmacological characteristics. One class (already referred to by some as SRIF1) appears to comprise sst2, sst3 and sst5 receptor subtypes. The other class (SRIF2) appears to comprise the other two recombinant receptor subtypes (sst1 and sst4). This promising approach is discussed but it is acknowledged that much more data from endogenous receptors in whole tissues are needed before further recommendations on somatostatin receptor nomenclature can be made.
Amino Acid Sequence↗
Linkage and association between insulin-dependent diabetes mellitus (IDDM) susceptibility and markers near the glucokinase gene on chromosome 7.
Insulin-dependent diabetes mellitus (IDDM) is characterized by autoimmune destruction of the insulin secreting beta-cells of the pancreas and subsequent disruption of glucose metabolism. The tendency of IDDM to cluster in families and the modest (36%) concordance rate in monozygotic twins indicates that both genetic and environmental factors contribute to IDDM susceptibility. Recent genome-wide searches using the affected sib-pair (ASP) approach have provided evidence for novel loci, in addition to HLA (IDDM1) and insulin (IDDM2), which show evidence of linkage to IDDM (P < 0.05). We have evaluated 35 microsatellite marker loci on human chromosome 7 for linkage to IDDM in 339 affected sib-pair families. Increased sharing of parental haplotypes in affected sib-pairs was detected for two microsatellite markers flanking glucokinase (GCK). Preferential transmission of alleles to affected offspring was observed at one of these marker loci, GCK3, indicating linkage disequilibrium between the marker and a disease susceptibility locus. This combination of linkage and disease association suggests that glucokinase, or a gene in the vicinity, plays an important part in IDDM susceptibility.
Alleles↗
Structure and function of ASP, the human homolog of the mouse agouti gene.
The mouse agouti coat color gene encodes a novel paracrine signaling molecule whose pulsatile expression produces a characteristic pattern of banded pigment in individual hairs. Several spontaneous agouti alleles produce adult-onset obesity and diabetes, and have provided important single-gene animal models for alterations in energy metabolism. Utilizing linkage groups conserved between mice and humans, we have cloned the human homolog of the mouse agouti gene from a human chromosome 20 yeast artificial chromosome known to contain S-adenosyl homocysteine hydrolase (AHCY). The human agouti gene, named Agouti Signaling Protein (ASP), encodes a 132 amino acid protein, the mRNA for which is expressed in testis, ovary, and heart, and at lower levels in liver, kidney, and foreskin. As predicted by the interactions of mouse agouti with the extension gene (which encodes the melanocyte receptor for alpha-melanocyte stimulating hormone [alpha-MSH]), expression of ASP in transgenic mice produces a yellow coat, and expression of ASP in cell culture blocks the alpha-MSH-stimulated accumulation of cAMP in mouse melanoma cells. The localization of ASP relative to other loci on chromosome 20 excludes it as a candidate for the MODY1 locus, a gene responsible for one form of early-onset non-insulin-dependent diabetes mellitus or maturity-onset diabetes of the young. The expression of ASP in human tissues suggests a function for agouti homologs in species that do not exhibit the characteristic phenotype of banded hairs.
Agouti Signaling Protein↗
Clinical characteristics of subjects with a missense mutation in glucokinase.
The clinical characteristics of subjects with a missense glucokinase mutation, gly299-->arg, were studied in a large pedigree, BX, initially characterized by some members having Maturity Onset Diabetes of the Young (MODY). Glucose tolerance, beta cell function and insulin sensitivity were measured with Homeostasis Model Assessment (HOMA) and with a 'Continuous Infusion of Glucose with Model Assessment' (CIGMA) test. Diabetic complications were clinically assessed. Subjects with glucokinase gly299-->arg were the same age, height, and obesity as the subjects without the mutation. Diabetes was usually asymptomatic at diagnosis and was treated with diet alone in 15 of the 18 subjects. Five of the 11 adult females had been diagnosed when they developed gestational diabetes. The fasting plasma glucose concentrations at the time of study were 4.3-12.6 mmol l-1, with the higher levels being in the more obese (p < 0.05) and in the older subjects (p < 0.05). In subjects with the mutation, beta cell function was impaired, being geometric mean 63% (normal-100%) compared with 126% in the subjects without the mutation (p < 0.001) measured by HOMA and in a subset assessed by CIGMA 59% and 127% (p < 0.01), respectively. There was no difference in fasting insulin concentrations, insulin sensitivity, lipid concentrations or blood pressure between the groups. The haemoglobin A1c was raised (mean 6.5% compared with 5.5% in the subjects without the mutation), but microvascular and macrovascular complications were uncommon. The subjects with the mutation did not have microalbuminuria but had an impaired vibration perception threshold compared with subjects without the mutation.(ABSTRACT TRUNCATED AT 250 WORDS)
Adolescent↗