Insulin regulates the synthesis of several proteins in a variety of tissues. Before techniques were available to quantify the amount of specific mRNAs, insulin was thought to regulate the synthesis of proteins by influencing the rate of translation of a fixed amount of mRNA. A very different interpretation is called for by experiments which show that insulin alters the amount of several specific mRNAs, but little is known about the mechanism. Insulin decreases the rate of synthesis of the critical gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) in both liver and H4IIE heptoma cells. We recently showed that insulin acts directly on H4IIE cells to decrease mRNAPEPCK activity without any other hormone intermediaries. This effect is mediated by the insulin receptor and occurs at insulin concentrations which are well within the physiological range range (10(-12)--10(-9) M). Here we extend these studies to show that insulin specifically inhibits transcription of the PEPCK gene. This inhibition results in a rapid decrease in the concentration of nuclear PEPCK transcripts which is followed, in turn, by a proportionate decline in cytoplasmic mRNAPEPCK and synthesis of the protein.
Diabetes affects many children. Researchers know little about children's perceptions of what type of support they need at school, which was a focus of this study. Group interviews and surveys examined children's perceptions of support in caring for their diabetes (type I diabetes) from school nurses, teachers, and friends. Results indicated the children felt supported at school, but improved flexibility by teachers and nurses (e.g., let me keep my meter with me always) and individualized care plans may improve their ability to manage their diabetes at school. Participating in after-school activities may be difficult for middle school youth. Children reported they needed additional help and support to cope with hypoglycemic episodes.
The glucokinase gene is 15.5-kilobases long, appears to be present as a single copy, and contains 10 exons that range in size from 96 to 977 base pairs. The transcription start site was located 127 nucleotides upstream from the translation initiation codon. The 5' flanking DNA contains several regions similar to dermed promoter elements. These include a probable "TATA box," an Spl binding site, and several elements related to liver-specific gene expression. In addition, we determined that transcription of the glucokinase gene increased at least 20-fold when diabetic rats were treated with insulin for 2 hr.Glucokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) plays a key role in the regulation of glucose homeostasis by catalyzing the first step in glycolysis (1). Expression of the enzyme is limited to hepatocytes and pancreatic ,B cells (2, 3), and it is regulated differently in these two tissues. The hepatic enzyme is induced by insulin and repressed by cAMP (4) whereas in the p cell glucokinase activity is increased by glucose (5). The glucokinase gene is, therefore, of interest both because of its tissue-specific expression and because of the several regulatory processes that can be analyzed. Before the cis-acting DNA elements responsible for the tissuespecific expression and hormonal regulation of this enzyme can be identified and studied, the structure of the glucokinase gene, the transcription unit in each tissue, and the sequence of its 5' flanking DNA must be determined.Glucokinase is thought to be a member of a family of hexokinases that have a common evolutionary origin (6). This concept was based on indirect evidence because none of the complete structures of the mammalian hexokinases were available. We recently deduced the structure of rat liver glucokinase and found that it shares 33% and 53% amino acid sequence identity with yeast hexokinase and the carboxylterminal portion of rat brain hexokinase I, respectively (7). We now have determined the structure of the glucokinase genet as an initial step toward answering how the various hexokinase isozymes are related to each other. Primer-Extension Analysis. A 36-base oligonucleotide (5'-ATGTTCCTGACTCCTGAGGCCACCTGTTGCAGGTGA-3') complementary to sequences near the 5' end of the glucokinase mRNA was synthesized and 5'-end-labeled with [_y-32P]ATP (>5000 Ci/mmol; 1 Ci = 37 GBq) and T4 polynucleotide kinase. The primer (3 x 10s cpm) was annealed in a total volume of 20 gl to 20 pug to poly(A)+ RNA; the annealing buffer contained 20 mM Tris-HCl (pH 7.5), 250 mM NaCl, and 1 mM EDTA. After hybridization for 1 hr at 60TC the reaction mixtures, containing the annealed primer and RNA, were diluted with 130 1A ofa solution containing 50 mM Tris-HCl (pH 7.5), 40 mM KCl, 10 mM dithiothreitol, 3 mM MgCl2, actinomycin D (75 ,ug/ml), deoxyribonucleotides at 0.5 mM each, and 2 units of avian myeloma virus reverse transcriptase (Promega Biotec) and then incubated at 37TC for 1 hr. The products of the reactions were size-fractionated on a 5% polyacrylamide/7 ...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.