Cloning of Novel Variants of HMGA1a to Understand the Role of Arginine Residues in DNA Binding
College
College of Arts and Sciences
Department
Biology
Faculty Mentor
Takita Sumter, Ph.D.
Abstract
Cancer initiation and progression occurs through a series of key molecular steps that lead to aberrations in tumor suppressor, oncoprotein, and signaling functions within the cell. While a number of pathways have been implicated in cancer progression, many mechanistic studies of genes altered as an adenoma progresses to a carcinoma result in upregulation of high mobility group A1 (hmga1). Mice bearing the hmga1 transgene develop aggressive lymphoid malignancies, and hmga1 overexpression leads to increased drug resistance and self-renewal capacity in a variety of cancers. The gene encodes three products as a result of alternative splicing – HMGA1a, HMGA1b, and HMGA1c – all of which preferentially bind DNA sequences rich in adenine (A) and thymine (T). While the specific function of these regions is not clearly understood, both DNA-protein and protein-protein interactions depend on the presence of three Arg-Gly-Arg (RGR) motifs. To elucidate the role of highly conserved arginine residues in HMGA1 function, we created single and double mutants of arginines in the first RGR motifs. Because the conversion of Arg at position 25 to alanine (R25A) and lysine (R25K) dramatically decreased DNA binding in vitro, we engineered mutations encoding both single arginine to glutamate (R25E) mutations and double mutations of arginine to glutamate and alanine (R25E27A). Mutations were generated using Quik-change mutagenesis approaches; specific conditions for gene amplification were optimized and verified by automated sequencing. From this, we hope to gain insights into the molecular networks involved in cancer initiation and progression.
Previously Presented/Performed?
Summer Undergraduate Research Experience (SURE) Symposium and Poster Session, Winthrop University, June and September 2016
Grant Support?
Supported by a grant from the National Institutes of Health IDeA Networks for Biomedical Research Excellence (NIH-INBRE), with prior support from a National Science Foundation Research Initiation Grant and an NIH Academic Research Enhancement Award.
Start Date
21-4-2017 1:00 PM
Cloning of Novel Variants of HMGA1a to Understand the Role of Arginine Residues in DNA Binding
DiGiorgio Campus Center, Room 220
Cancer initiation and progression occurs through a series of key molecular steps that lead to aberrations in tumor suppressor, oncoprotein, and signaling functions within the cell. While a number of pathways have been implicated in cancer progression, many mechanistic studies of genes altered as an adenoma progresses to a carcinoma result in upregulation of high mobility group A1 (hmga1). Mice bearing the hmga1 transgene develop aggressive lymphoid malignancies, and hmga1 overexpression leads to increased drug resistance and self-renewal capacity in a variety of cancers. The gene encodes three products as a result of alternative splicing – HMGA1a, HMGA1b, and HMGA1c – all of which preferentially bind DNA sequences rich in adenine (A) and thymine (T). While the specific function of these regions is not clearly understood, both DNA-protein and protein-protein interactions depend on the presence of three Arg-Gly-Arg (RGR) motifs. To elucidate the role of highly conserved arginine residues in HMGA1 function, we created single and double mutants of arginines in the first RGR motifs. Because the conversion of Arg at position 25 to alanine (R25A) and lysine (R25K) dramatically decreased DNA binding in vitro, we engineered mutations encoding both single arginine to glutamate (R25E) mutations and double mutations of arginine to glutamate and alanine (R25E27A). Mutations were generated using Quik-change mutagenesis approaches; specific conditions for gene amplification were optimized and verified by automated sequencing. From this, we hope to gain insights into the molecular networks involved in cancer initiation and progression.
