SCD | SCTD | SDG | SGDDM | SHBG | UMD | Main Page
 
MOLECULAR GENETICS OF HERITABLE HUMAN DISORDERS
     
Janice Y. Chou, Ph.D., Principal Investigator
Li-Yaun Chen, Ph.D.,Postdoctoral Fellow
Abhijit Ghosh, Ph.D., Postdoctoral Fellow
Jeng-Jer Shieh, Ph.D., Postdoctoral Fellow
Mao-Sen Sun, M.D., Ph.D., Postdoctoral Fellow
Hisayuki Hiraiwa, M.D., former Postdoctoral Fellow
Mugen Terzioglu, former Predoctoral Fellow
Chi-Jiunn Pan, Senior Research Assistant
Peter Chiang, Ph.D., Guest Researcher
Moshe Zilberstein, M.D., Guest Researcher
Janice Chou
 
The goal of the Section on Cellular Differentiation is to understand the biology and pathogenesis of type 1 glycogen storage disease (GSD-1), a group of autosomal recessive disorders, and to develop novel therapies for these disorders. GSD-1 is caused by a deficiency in the endoplasmic reticulum-bound glucose-6-phosphatase (G6Pase) system that comprises two integral membrane proteins, G6Pase and the associated glucose-6-phosphate transporter (G6PT). G6PT translocates G6P, the product of gluconeogenesis and glycogenolysis, from cytoplasm to the lumen of the endoplasmic reticulum (ER), and, inside the ER, G6Pase catalyzes the hydrolysis of G6P to produce glucose and phosphate. Therefore, G6PT and G6Pase work in concert to maintain glucose homeostasis. Deficiencies in G6Pase and G6PT cause GSD-1a and GSD-1b, respectively. Both manifest phenotypic G6Pase deficiency, which is characterized by growth retardation, hypoglycemia, hepatomegaly, nephromegaly, hyperlipidemia, hyperuricemia, and lactic acidemia. GSD-1b patients also suffer from infectious complications due to chronic neutropenia and functional deficiencies of neutrophils and monocytes.

Type 1 Glycogen Storage Disease and HNF1_-Deficiency
Hiraiwa, Pan, Chou in collaboration with T.E. Akiyama and F.J. Gonzales (NCI)
We observed that the clinical manifestations (e.g., growth retardation, hepatomegaly, hyperlipidemia, and renal dysfunction) of GSD-1 patients are shared by Hnf1a-/- mice deficient in a transcriptional activator, hepatocyte nuclear factor 1a (HNF1a). Hnf1a-/- mice also develop noninsulin-dependent diabetes mellitus (NIDDM) caused by defective insulin secretion. It has been speculated that overexpression of G6Pase might contribute to the pathophysiology of NIDDM. We therefore sought to determine whether there is a molecular link between HNF1a deficiency and function of the G6Pase system. Transactivation studies revealed that HNF1a is required for transcription of the G6PT gene. Hepatic G6PT mRNA levels and microsomal G6P transport activity are also markedly reduced in Hnf1a-/- mice compared with Hnf1a+/+ and Hnf1a+/- littermates. On the other hand, hepatic G6Pase mRNA expression and activity are up-regulated in Hnf1a-/- mice, consistent with observations that G6Pase expression is increased in diabetic animals. Taken together, the results strongly suggest that metabolic abnormalities in Hnf1a-null mice are caused, in part, by G6PT deficiency and by perturbations of the G6Pase system, establishing for the first time a molecular link between the common phenotypes of GSD-1 and Hnf1a-/- mice.

Glucocorticoids Activate Transcription of the Glucose-6-Phosphate Transporter Gene
Hiraiwa
G6P plays a pivotal role in intermediate metabolism. Changes in G6P levels in cells affect not only glucose metabolism but also glycogen biosynthesis and lipid biosynthesis. Therefore, it is of vital importance to understand the regulation of G6PT gene expression. We therefore delineated the role of glucocorticoids in transcription of the G6PT gene. The glucocorticoids are involved in the regulation of a wide range of physiological process, including glucose metabolism. We showed that the basal G6PT promoter is contained within nucleotides -369 to -1 upstream of the translation state site, which contains three activation elements. We demonstrated that glucocorticoids activate G6PT gene transcription and that the glucocorticoid action is mediated through a glucocorticoid response element within activation element-2 of the promoter. Taken together, the results suggest that glucocorticoids play a pivotal role in regulating the G6PT gene.

Structure and Function Analysis of Mutations in Glucose-6-Phosphatase
Shieh, Terzioglu, Hisayusi, Hiraiwa, Pan, Chen, Chou
To date, 75 G6Pase mutations have been identified in GSD-1a patients on the basis of the mutations’ absence from the normal population and/or on their cosegregation with the disease phenotype. The mutations include 48 missense, nine nonsense, 15 insertion/deletion, and three splicing mutations. Interestingly, 64 percent of the candidate mutations are missense mutations that result in single amino acid substitutions. However, only 19 missense mutations have been functionally characterized. Characterization of these mutations will provide valuable information on functionally important residues of the protein. Using site-directed mutagenesis and transient expression assays, we have characterized all 48 missense mutations. The database of residual activity retained by these mutants will provide a reference in evaluating genotype-phenotype relationships and the minimal G6Pase activity required to correct the GSD-1a phenotype.
 

PUBLICATIONS

  1. Chen L-Y, Lin B, Pan C-J, Hiraiwa H, Chou JY. Structural requirements for the stability and microsomal transport activity of the human glucose-6-phosphate transporter. J Biol Chem 2000;275:34280-34286.
  2. Chou JY. Glucose-6-phosphatase. In: Creighton TE, ed. Encyclopedia of molecular medicine. New York: John Wiley, in press.
  3. Chou JY. Glucose-6-phosphatase: molecular biology and genetic deficiency. Recent Res Dev Biochem 2000;2:159-169.
  4. Chou JY. Glucose-6-phosphate transporter. In: Creighton TE, ed. Encyclopedia of molecular medicine. New York: John Wiley, in press.
  5. Chou JY. Methionine adenosyltransferase. In: Creighton TE, ed. Encyclopedia of molecular medicine. New York: John Wiley, in press.
  6. Chou JY. The molecular basis of type 1 glycogen storage diseases. Curr Mol Med 2000;1:25-44.
  7. Foster JD, Wiedemann JM, Pan C-J, Chou JY, Nordlie RC. Discriminant responses of the catalytic unit and glucose 6-phosphate transporter components of the hepatic glucose-6-phosphatase system in Ehrlich asites-tumor-bearing mice. Arch Biochem Biophys 2001;393:117-122.
  8. Hiraiwa H, Chou JY. Glucocorticoids activate transcription of the gene for glucose-6-phosphate transporter, deficient in glycogen storage Disease type 1b. DNA Cell Bio. 2001;20:447-453.
  9. Hiraiwa H, Pan C-J, Lin B, Akiyama TE, Gonzalez FJ, Chou JY. A molecular link between the common phenotypes of type 1 glycogen storage disease and HNF1_-null mice. J Biol Chem 2001;276:7963-7967.
  10. Lin B, Pan C-J, Chou JY. Human variant glucose-6-phosphate transporter is active in microsomal transport. Hum Genet 2001;107:526-529.
  11. Mandal AK, Zhang Z, Chou JY, Mukherjee AB. Pancreatic phospholipase A2 via its receptor regulates expression of key enzymes of phospholipid and sphingolipid metabolism. FASEB J 2001;15:1834-1836.
  12. Mandal AK, Zhang Z, Chou JY, Zimonjic D, Keck CL, Popescu N, Mukherjee AB. Molecular characterization of murine pancreatic phospholipase A2. DNA Cell Biol 2001;20:149-157.
  13. Phornphutkul C, Anikster Y, Huizing M, Braun P, Brodie C, Chou JY, Gahl WA. The promoter of a lysosomal membrane transporter gene, CTNS, binds Sp-1, shares sequences with promoter of an adjacent gene (CARKL), and causes cystinosis if mutated in a critical region. Am J Hum Genet 2001;69:712-721.