Defining unique structural features in the MAFA and MAFB transcription factors that control Insulin gene activity

Jeeyeon Cha; Xin Tong; Katie C Coate; Min Guo; Jin-Hua Liu; Garrett Reynolds; Emily M Walker; Richard A Stein; Hassane Mchaourab; Roland Stein

doi:10.1016/j.jbc.2024.107938

Defining unique structural features in the MAFA and MAFB transcription factors that control Insulin gene activity

J Biol Chem. 2024 Oct 28;300(12):107938. doi: 10.1016/j.jbc.2024.107938. Online ahead of print.

Authors

Affiliations

¹ Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA; Division of Diabetes, Endocrinology, and Metabolism, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
² Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA.
³ Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA; Division of Diabetes, Endocrinology, and Metabolism, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA; VA Tennessee Valley Healthcare System, Nashville, Tennessee, USA.
⁴ Division of Metabolism, Endocrinology & Diabetes, Departments of Molecular & Integrative Physiology and Internal Medicine, University of Michigan, Ann Arbor, Michigan, USA.
⁵ Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA; Center for Applied Artificial Intelligence in Protein Dynamics, Vanderbilt University, Nashville, Tennessee, USA.
⁶ Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA. Electronic address: roland.stein@vanderbilt.edu.

Abstract

MAFA and MAFB are related basic-leucine-zipper domain-containing transcription factors which have important overlapping and distinct regulatory roles in a variety of cellular contexts, including hormone production in pancreatic islet cells. Here, we first examined how mutating conserved MAF protein-DNA contact sites obtained from X-ray crystal structure analysis impacted their DNA-binding and Insulin enhancer-driven activity. While most of these interactions were essential and their disruption severely compromised activity, we identified that regions outside of these contact sites also contributed to transcriptional activity. AlphaFold 2 (https://alphafold.ebi.ac.uk), an artificial intelligence-based structural prediction program, was used to determine if there were also differences in the three-dimensional organization of the non-DNA binding/dimerization sequences of MAFA and MAFB. This analysis was conducted on the WT proteins as well as the pathogenic MAFA^Ser64Phe and MAFB^Ser70Ala transactivation domain mutants, with differences revealed between MAFA^WT and MAFB^WT as well as between MAFA^Ser64Phe and MAFA^WT, but not between MAFB^Ser70Ala and MAFB^WT. Moreover, dissimilarities between these proteins were also observed in their ability to cooperatively stimulate Insulin enhancer-driven activity in the presence of other islet-enriched transcription factors. Analysis of MAFA and MAFB chimeras disclosed that these properties were influenced by their unique C-terminal region structural differences predicted by AlphaFold 2. Our findings have revealed key structural features of these closely related proteins that impact their ability to regulate gene expression.

Keywords: AlphaFold 2; Insulin; MAFA; MAFB; gene expression; islet biology; transactivation; transcription factors.