Enhanced binding of guanylated poly(A) RNA by the LaM domain of LARP1

La-related proteins (LARPs) are a family of RNA-binding proteins that share a conserved La motif (LaM) domain. LARP1 plays a role in regulating ribosomal protein synthesis and stabilizing mRNAs and has a unique structure without an RNA binding RRM domain adjoining the LaM domain. In this study, we investigated the physical basis for LARP1 specificity for poly(A) sequences and observed an unexpected bias for sequences with single guanines. Multiple guanine substitutions did not increase the affinity, demonstrating preferential recognition of singly guanylated sequences. We also observed that the cyclic di-nucleotides in the cCAS/STING pathway, cyclic-di-GMP and 3‘,3’-cGAMP, bound with sub-micromolar affinity. Isothermal titration measurements were complemented by high-resolution crystal structures of the LARP1 LaM with six different RNA ligands, including two stereoisomers of a phosphorothioate linkage. The selectivity for singly substituted poly(A) sequences suggests LARP1 may play a role in the stabilizing effect of poly(A) tail guanylation.

La相关蛋白（La-related proteins, LARPs）是一类拥有保守La基序（La motif, LaM）结构域的RNA结合蛋白家族。LARP1可调控核糖体蛋白合成并稳定信使RNA（mRNA），其结构独具特色，不存在与LaM结构域邻接的RNA结合RRM（RNA recognition motif）结构域。本研究探究了LARP1对聚腺苷酸序列特异性识别的分子基础，意外观察到其对携带单鸟嘌呤的序列存在结合偏好。多个鸟嘌呤取代并未提升结合亲和力，证实LARP1优先识别单鸟苷酸化序列。此外，本研究还发现cCAS/STING通路中的环状二核苷酸——环二GMP（cyclic-di-GMP）与3',3'-cGAMP——以亚微摩尔亲和力结合。我们通过等温滴定量热实验结合高分辨率晶体结构解析，获取了LARP1的LaM结构域与六种不同RNA配体的复合物结构，其中包含两类硫代磷酸酯键立体异构体。对单取代聚腺苷酸序列的结合选择性提示，LARP1或在聚腺苷酸尾鸟苷酸化的稳定效应中发挥功能。