Department of Biochemistry, Stanford University School of Medicine, Stanford, CA, USA.
Biophysics Program, Stanford University School of Medicine, Stanford, CA, USA.
Nat Commun. 2022 Aug 4;13(1):4522. doi: 10.1038/s41467-022-31968-z.
Genomic methods have been valuable for identifying RNA-binding proteins (RBPs) and the genes, pathways, and processes they regulate. Nevertheless, standard motif descriptions cannot be used to predict all RNA targets or test quantitative models for cellular interactions and regulation. We present a complete thermodynamic model for RNA binding to the S. cerevisiae Pumilio protein PUF4 derived from direct binding data for 6180 RNAs measured using the RNA on a massively parallel array (RNA-MaP) platform. The PUF4 model is highly similar to that of the related RBPs, human PUM2 and PUM1, with one marked exception: a single favorable site of base flipping for PUF4, such that PUF4 preferentially binds to a non-contiguous series of residues. These results are foundational for developing and testing cellular models of RNA-RBP interactions and function, for engineering RBPs, for understanding the biophysical nature of RBP binding and the evolutionary landscape of RNAs and RBPs.
基因组学方法在鉴定 RNA 结合蛋白 (RBPs) 及其调控的基因、途径和过程方面非常有价值。然而，标准的基序描述不能用于预测所有的 RNA 靶标，也不能用于测试细胞相互作用和调控的定量模型。我们提出了一个完整的热力学模型，用于酵母 Pumilio 蛋白 PUF4 与 6180 个 RNA 的直接结合数据，这些 RNA 是使用 RNA 大规模平行阵列 (RNA-MaP) 平台测量的。PUF4 模型与相关的 RBPs（人类 PUM2 和 PUM1）非常相似，只有一个明显的例外：一个有利于碱基翻转的单一有利位点，使得 PUF4 优先结合非连续的一系列残基。这些结果为开发和测试 RNA-RBP 相互作用和功能的细胞模型、工程化 RBPs、理解 RBP 结合的生物物理性质以及 RNA 和 RBPs 的进化景观奠定了基础。
