Donald F. Steiner, M.D. - US grants

[unreadable] DESCRIPTION (provided by applicant): The major objective of this research program is to gain new insights into the molecular and cell biological mechanisms underlying the production and actions of insulin and related peptide hormones of the islets of Langerhans, including their genetic basis, evolutionary origins, and disorders in these processes, which may contribute to the pathophysiology of diabetes and/or other diseases. New projects and extensions of previous work are proposed as follows: (1) Studies on prohormone convertases PC1/PC3 (SPC3) and PC2 (SPC2), including further characterization of neuropeptide processing defects in mice with targeted disruptions of the PC1/PC3 and PC2 genes individually or in various crosses, (2) Studies on the role of the neuroendocrine protein proSAAS in normal proinsulin processing and on the mechanisms underlying ACTH hypersecretion in 7B2 null mice, (3) Studies on convertase subdomain structure and on transcriptional regulation of the PC 1/PC3 and PC2 genes, (4) Studies on prohormone structure in relation to their processing by convertases, (5) New studies on the identification and characterization of unique beta and alpha cell proteins and their roles in glucagon and insulin production and secretion, (6) Further studies on evolution of insulin and insulin/IGF receptor signaling systems, and (7) New studies to characterize the temporal course of changes in hepatic gene expression in diabetic animals in response to insulin. These studies should all contribute to improving our understanding of the genetic, evolutionary, and molecular biological mechanisms underlying islet hormone production and action and should contribute to a better understanding of the pathophysiology of diabetes.

DESCRIPTION (Adapted from Applicant's Abstract): The major objective of this research program is to gain new insights into the molecular and cell biological mechanisms underlying the production and actions of insulin and related peptide hormones of the islets of Langerhans, including their genetic basis, evolutionary origins, and disorders in these processes which may contribute to the pathophysiology of diabetes and/or other diseases. New projects and extensions of previous work are proposed as follows: 1) Studies on prohormone convertases PC2 (SPC2) and PC3 (SPC3), including their biosynthesis, maturation and actions on prohormone processing in mice with targeted disruptions in their genes; 2) Effects of the SPC2 gene null mutation on the development and morphology of the islets of Langerhans; 3) studies on the regulation and tissue-specific transcription of the genes encoding SPC2 and SPC3; 4) Studies on the structure and functional significance of conserved subdomains of the convertases; 5) Studies on the processing of ProIGF-I and its significance in normal and transformed cell lines; 6) Studies on the effects of carboxypeptidase E (CPE) deficiency on islet prohormone processing; 7) Studies on the insulin receptor with emphasis on the binding ectodomain, and the use of receptor fusion proteins for studies of its structure and biosynthesis; 8) Further studies on the evolution of insulin and the IGFs and their receptor proteins in lower vertebrates, with emphasis on the divergence of metabolic and growth regulatory functions of a common ancestral insulin-like peptide (ILP) in protochordates.

molecular biology; diabetes mellitus genetics; cell biology; biomedical facility; diabetes mellitus; pancreatic islets; nucleic acid sequence; restriction fragment length polymorphism; complementary DNA; computer assisted sequence analysis; molecular cloning; Xenopus oocyte; in situ hybridization; southern blotting; laboratory rat; Escherichia coli; linkage mapping; northern blottings; tissue /cell culture;

This proposal has long-term goals related to (a) identifying the structural attributes of insulin-like growth factor I (IGF-I) which determine its potency for interactions with type I IGF receptors, and its consequent overall biological effects, (b) evaluating the development of rules that might apply to the separate determinants of affinity and selectivity for receptor interactions within structurally related groups of ligands and receptors, and (c) considering the potential for development of IGF-I analogs that might be of therapeutic or experimental value. Specific aims for the project are summarized as follows: (a) To prepare hybrid IGF-I/insulin analogs with selected structural changes and amino acid replacements by chemical peptide synthesis and semisynthesis. (b) To evaluate the potencies of these analogs with regard to their abilities to bind to the type I IGF and insulin receptors through binding competition studies and direct analysis of radiolabeled ligand binding. (c) To consider the separate issues of receptor site affinity and selectivity in relation to the IGF-I and insulin systems through comparisons of analog interactions with both receptors. (d) To address aspects of potential differences in the solution structures of analogs by spectroscopic methods and chemical modification. (e) To evaluate the consequences of structural changes in these analogs in relation to their biological activities in vitro, by use of assays for 3H-thymidine incorporation into DNA and for additional metabolic responses. (f) To evaluate the potential biological implications of binding site heterogeneity in ligand interactions with the type I IGF receptor through the kinetic analysis of related binding studies. (g) To approach an understanding of type I IGF receptor heterogeneity at the molecular level through use of chemical and affinity methods. As aberrations in IGF-I physiology can be expected to result under different sets of circumstances in either inappropriately low or inappropriately high rates of tissue growth, and as the IGF-I system could be useful in by-passing the insulin system in states of insulin resistance, it is not difficult to envision the potential health-related importance of analogs of IGF-I that might (a) be selective for one or the other of its actions, (b) be even more potent than the natural hormone, (c) inhibit the action endogenous hormone by binding to receptor without inducing cellular responses, or (d) act through multiple receptor systems to produce desired metabolic effects. This proposal represents a study of the IGF-I system at a fundamental level, at this stage, and the application of the methods of chemical peptide synthesis and semisynthesis to the elucidation of relevant structure-function relationships.

This proposal seeks the continuation of the University of Chicago Diabetes Research and Training Center. The Center is an interdepartmental and cooperative endeavor involving more than 70 investigators. The aims of the Center are: 1) to foster diabetes and endocrinology research in a supportive, integrative, collaborative and multidisciplinary environment; 2) to enhance the clinical and basic research capabilities of established diabetes investigators; 3) to encourage investigators not involved in diabetes research to become interested in pursuing problems related to diabetes and endocrinology; 4) to develop and implement programs for the training of both health care professionals and diabetic patients in the diagnosis and management of diabetes; 5) to facilitate the transfer of newer research findings to the clinical arena where they may be formally considered in terms of the pathogenesis, diagnosis and treatment of the disease; and 6) to inform others in both professional and lay settings of the accomplishments, opportunities and advancements in diabetes research and training. This application contains proposals for three separate components; the Basic Biomedical Research component, the Administrative Core Component and the Demonstration and Education component. The Basic Biomedical Research component consists of four Core Laboratories to foster diabetes research, and a Pilot and Feasibility Program to foster the participation of younger and established investigators in research related to diabetes. The Administrative Core Component is responsible for overseeing the operation of the Center as a whole. The Demonstration and Education Component consists of an Education Resource Core and a Demonstration Unit. Taken together, these objectives and components define The University of Chicago DRTC.

DESCRIPTION (provided by applicant): The major objective of this research program is to gain new insights into the molecular and cell biological mechanisms underlying the production and actions of insulin and related peptide hormones of the islets of Langerhans and the neuroendocrine system, including their genetic basis, evolutionary origins, and disorders in these processes, which may contribute to the pathophysiology of diabetes and/or other diseases. New projects and extensions of previous work are proposed as follows: (1) New studies on mice with gene disruptions of the neuroendocrine prohormone convertases, including PC5/6 isoform A, to assess its role in processing important hormonal and neuropeptide precursors in brain, hypothalamus, islets and gastrointestinal tract and including, studies on the processing of ghrelin and related CNS/hypothalamic peptides involved in regulation of food intake and energy metabolism, (2) New studies on the role of prohormone structure in cleavage site selectivity of individual neuroendocrine PC family members, (3) Studies on the fate of PC1/3 and PC2 in vivo in neuroendocrine cells after completion of processing of secretory granule contents, i.e., retention, secretion and/or degradation, (4) Further studies on the evolution of the insulin family of hormones and the detailed structural basis of insulin's interaction with and activation of the insulin receptor. These studies should all contribute to improving our understanding of the genetic, cellular and molecular biological, and evolutionary mechanisms underlying islet hormone production and action and should contribute to a better understanding of the pathophysiology of diabetes.