PTBP1 controls miRNA loading on target RNAs: lessons from the CyCoNP lncRNA

Abstract

The concerted action of regulatory RNA and RNA binding proteins (RBPs) provides cells with highly versatile and transient tools to fine-tune gene expression in a broad variety of cellular systems (Unfried and Ulitsky, Nat Cell Biol 24: 608–615 [2022]; Hentze et al., Nat Rev Mol Cell Biol 19: 327–341 [2018]; Suzuki et al., Nat Genet 50: 657–661 [2018]). In this work, we explore the function of a specific interaction between PTBP1 and the cytoplasmic long noncoding RNA (lncRNA) CyCoNP, highly expressed in neural progenitors (Desideri et al., NAR 52: 9936–9952 [2024]), in which the RBP regulates the abundance of the lncRNA by a miRNA-mediated mechanism. PTBP1 is a well-known splicing regulator, although limited and peculiar examples of its involvement in other cellular processes, such as IRES-dependent translation and miRNA recognition of target RNAs, have been described (Dorn et al., Cell Death Dis 14: 6429 [2023]; Kim et al., Nat Commun 12: 5057 [2021]). We have recently characterized CyCoNP lncRNA as a regulator of NCAM1, which acts through a mechanism that involves direct RNA–RNA interaction with NCAM1 mRNA, balancing the availability and the localization of miR-4492 in its vicinity (Desideri et al., NAR 52: 9936–9952 [2024]). Here we expand the repertoire of molecular players acting in this circuitry by describing a direct interaction between PTBP1 and CyCoNP lncRNA. Through endogenous RNA purification, protein immunoprecipitation, and exploiting CyCoNP mutant constructs, we found that PTBP1, when interacting with CyCoNP, hampers miR-4492 binding to the lncRNA and in turn impedes its regulation on NCAM1 mRNA. This work aims to expand the biochemical characterization of regulatory networks relying on RBPs and their cognate target RNAs, highlighting the relevance of the analysis of the subcellular environment for each case of study.