Abstract 2444: Do all 4 NCL RNA Binding Domains play a role in mRNA target specificity? An in silico study to decipher NCL-mRNA interactions

Abstract

Abstract RNA binding proteins (RBPs) are post-transcriptional regulators that associate with RNA molecules through one or more of their RNA-binding domains (RBDs). The dysregulation of RBPs, and the RNA metabolism pathways they intersect, are known drivers of tumorigenesis. Nucleolin (NCL) is a stress responsive RBP with four RBDs that directly impacts gene expression by altering rRNA processing, RNA localization, miRNA biogenesis, mRNA decay, and translation. NCL is known to interact with both the 3' and 5' untranslated regions (UTR) and coding regions of target mRNAs to either modulate their stability or translation via recognition of AU-rich or G-rich elements in the RNA. However, the consequences of NCL-mRNA interactions are target-specific as well as context and location dependent. While NCL interacts with both 3' and 5' UTR of TP53 mRNA and suppresses its translation, interactions of NCL with 5' UTR of an anti-apoptotic gene, BCL2, stabilizes this transcript. NCL-mediated enhanced biogenesis of tumorigenic mir-15a and mir-16, on the other hand, results in the translational repression of BCL2. Because NCL exerts its functions in such a complex manner, elucidation of its roles in tumorigenic pathways is paramount in the development of therapeutical applications targeting NCL-RNA interactions. Mechanisms driving NCL-mRNA interactions are currently unknown and this knowledge gap is further compounded by the incomplete nature of the current structural information for NCL and its mRNA targets. In this study, we aim to elucidate NCL RBD-mRNA interactions using computational tools. We investigated interactions of NCL with a specific subset of mRNAs that are implicated in breast cancer progression. We have previously generated and validated in silico models for all the four NCL-RBDs (1-4) in various combinations. Using several secondary and tertiary structure prediction programs along with structure refinement programs, we have generated 3D models of TP53, BCL2, CSF-1, and GADD45α mRNA UTR molecules. Our docking analyses suggests that RBD1-2 and RBD3-4 collaborate in most NCL-mRNA interactions using RNA binding motifs (RNP1 & RNP2) on NCL. We further identify additional contact points in the bridging linker regions between the RBDs that might be crucial for the structural stability of these complexes. This study provides details previously not reported in the literature and provides a comprehensive map of NCL RBD-mRNA interactions that have direct pathophysiological relevance in breast cancer. Altered levels of NCL expression are linked with poor prognosis in breast cancers and targeting NCL results in a better survival outcome. Understanding how NCL associates with its mRNA targets will reveal binding pockets and hence potential sites for drug interventions in addition to characterizing the complex nature of NCL-mRNA interactions/specificity in multiple cancer types. Citation Format: Avdar San, Anjana Saxena, Shaneen Singh. Do all 4 NCL RNA Binding Domains play a role in mRNA target specificity? An in silico study to decipher NCL-mRNA interactions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2444.