Cha Jeeyeon, Tong Xin, Coate Katie C, Guo Min, Liu Jin-Hua, Reynolds Garrett, Walker Emily M, Stein Richard A, Mchaourab Hassane, Stein Roland
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.
Department of Molecular Physiology & Biophysics, Vanderbilt University, Nashville, Tennessee, USA.
J Biol Chem. 2024 Dec;300(12):107938. doi: 10.1016/j.jbc.2024.107938. Epub 2024 Oct 28.
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 and MAFB transactivation domain mutants, with differences revealed between MAFA and MAFB as well as between MAFA and MAFA, but not between MAFB and MAFB. 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.
MAFA和MAFB是相关的含碱性亮氨酸拉链结构域的转录因子，它们在多种细胞环境中具有重要的重叠和不同的调节作用，包括胰岛细胞中的激素产生。在这里，我们首先研究了从X射线晶体结构分析中获得的保守MAF蛋白-DNA接触位点的突变如何影响它们的DNA结合和胰岛素增强子驱动的活性。虽然这些相互作用大多是必不可少的，并且它们的破坏严重损害了活性，但我们发现这些接触位点之外的区域也有助于转录活性。基于人工智能的结构预测程序AlphaFold 2（https://alphafold.ebi.ac.uk）被用于确定MAFA和MAFB的非DNA结合/二聚化序列的三维组织是否也存在差异。对野生型蛋白以及致病性MAFA和MAFB反式激活结构域突变体进行了这项分析，结果显示MAFA和MAFB之间以及MAFA和MAFA之间存在差异，但MAFB和MAFB之间没有差异。此外，在存在其他富含胰岛的转录因子的情况下，这些蛋白在协同刺激胰岛素增强子驱动的活性的能力方面也存在差异。对MAFA和MAFB嵌合体的分析表明，这些特性受AlphaFold 2预测的它们独特的C末端区域结构差异的影响。我们的研究结果揭示了这些密切相关的蛋白影响其调节基因表达能力的关键结构特征。
