Self-balanced regulation by the long non-coding RNA Lockd on the cell cycle progression of cortical neural progenitor cells through counteracting cis and trans roles

Neural stem/progenitor cells (NSPCs) undergo active proliferation and exit the cell cycle upon precise regulation to produce differentiated progenies in order. Long non-coding RNAs (lncRNAs) have emerged as critical players in the developmental processes of NSPCs; however, relatively few have been shown to regulate the cell cycle in vivo directly. Here, we identified an NSPC-expressed lncRNA Lockd (lncRNA downstream of Cdkn1b) in the developing forebrain. Using in vivo loss of function models by premature termination of Lockd transcription via knockin polyadenylation signals or shRNA-mediated knockdown of Lockd (Lockd-KD), we show that Lockd is required for proper cell cycle progression of cortical NSPCs and the production of TBR2+ intermediate neural progenitor cells during cortical development. Interestingly, a comparison of genetic profiling in the two models reveals that Lockd promotes the expression of two counteracting cell cycle-related genes, Cdkn1b in cis and Ccnd1 in trans. Overexpression of Ccnd1 or Cdkn1b-KD can rescue the cellular phenotypes of reduced cycling progenitors in Lockd-KD. Our results imply that lncRNA could act through distinct cis and trans mechanisms to achieve a self-balanced function. Set 1: The control and Lockd-shRNA expressing plasmids were electroporated into NSPCs at E14.5, and GFP positive cells were sorted by flow cytometry at E16.5 (two biological replicates for control or Lockd-shRNA). Set 2: E13.5 cortices of wild type (WT) and PlyA-KI mice were also dissected and pooled for transcriptome profiling (two biological replicates for WT or PlyA-KI).