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

The concerted action of regulatory RNA and RNA binding proteins (RBPs) provide cells with highly versatile and transient tools to fine tune gene expression in a broad variety of cellular systems (Unfried and Ulitsky 2022, Hentze et al. 2018, Suzuki et al. 2018). In this work, we explore the function of a specific interaction between PTBP1 and the cytoplasmic long non-coding RNA (lncRNA) CyCoNP, highly expressed in neural progenitors (Desideri et al. 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. 2023, Kim et al. 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. 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.