Department of Biochemistry, McGill University, Montréal, Canada.
Centre de recherche en biologie structurale, McGill University, Montréal, Canada.
Nucleic Acids Res. 2022 Sep 9;50(16):9534-9547. doi: 10.1093/nar/gkac696.
La-related proteins (LARPs) comprise a family of RNA-binding proteins involved in a wide range of posttranscriptional regulatory activities. LARPs share a unique tandem of two RNA-binding domains, La motif (LaM) and RNA recognition motif (RRM), together referred to as a La-module, but vary in member-specific regions. Prior structural studies of La-modules reveal they are pliable platforms for RNA recognition in diverse contexts. Here, we characterize the La-module of LARP1, which plays an important role in regulating synthesis of ribosomal proteins in response to mTOR signaling and mRNA stabilization. LARP1 has been well characterized functionally but no structural information exists for its La-module. We show that unlike other LARPs, the La-module in LARP1 does not contain an RRM domain. The LaM alone is sufficient for binding poly(A) RNA with submicromolar affinity and specificity. Multiple high-resolution crystal structures of the LARP1 LaM domain in complex with poly(A) show that it is highly specific for the RNA 3'-end, and identify LaM residues Q333, Y336 and F348 as the most critical for binding. Use of a quantitative mRNA stabilization assay and poly(A) tail-sequencing demonstrate functional relevance of LARP1 RNA binding in cells and provide novel insight into its poly(A) 3' protection activity.
LARP 相关蛋白（LARPs）是一组参与广泛的转录后调控活动的 RNA 结合蛋白。LARPs 共享独特的串联 RNA 结合结构域，La 基序（LaM）和 RNA 识别基序（RRM），合称为 La 结构域，但成员特异性区域不同。先前对 La 结构域的结构研究表明，它们是在不同环境中识别 RNA 的柔韧平台。在这里，我们对 LARP1 的 La 结构域进行了表征，LARP1 在调节核糖体蛋白的合成方面发挥着重要作用，其合成反应受 mTOR 信号和 mRNA 稳定的调控。LARP1 的功能已得到很好的描述，但它的 La 结构域没有结构信息。我们发现，与其他 LARPs 不同，LARP1 的 La 结构域不包含 RRM 结构域。单独的 LaM 足以以亚微摩尔亲和力和特异性结合 poly(A) RNA。与 poly(A) 复合的 LARP1 LaM 结构域的多个高分辨率晶体结构表明，它对 RNA 3'-末端具有高度特异性，并确定 LaM 残基 Q333、Y336 和 F348 是结合的最关键残基。使用定量的 mRNA 稳定测定和 poly(A) 尾测序表明，LARP1 在细胞中的 RNA 结合具有功能相关性，并为其 poly(A) 3' 保护活性提供了新的见解。
