LETR1 is a lymphatic endothelial-specific lncRNA governing cell proliferation and migration through KLF4 and SEMA3C

Abstract

Recent studies have revealed the importance of long noncoding RNAs (lncRNAs) as tissue-specific regulators of gene expression. There is ample evidence that distinct types of vasculature undergo tight transcriptional control to preserve their structure, identity, and functions. We determine a comprehensive map of lineage-specific lncRNAs in human dermal lymphatic and blood vascular endothelial cells (LECs and BECs), combining RNA-Seq and CAGE-Seq. Subsequent antisense oligonucleotide-knockdown transcriptomic profiling of two LEC- and two BEC-specific lncRNAs identifies LETR1 as a critical gatekeeper of the global LEC transcriptome. Deep RNA-DNA, RNA-protein interaction studies, and phenotype rescue analyses reveal that LETR1 is a nuclear trans-acting lncRNA modulating, via key epigenetic factors, the expression of essential target genes, including KLF4 and SEMA3C, governing the growth and migratory ability of LECs. Together, our study provides several lines of evidence supporting the intriguing concept that every cell type expresses precise lncRNA signatures to control lineage-specific regulatory programs.

The choice of negative control antisense oligonucleotides dramatically impacts downstream analysis depending on the cellular background

“…This study complements our previous findings, where we analyzed the functionality of human lncRNAs in vascular biology by performing ASO-mediated knockdown of 2 LEC- and 2 BEC-specific lncRNAs followed by CAGE-Seq [ 18 ]. Here, we presented the early control steps in which we carefully characterized the transcriptional impact on LECs and BECs of two commercially available negative control ASOs.…”

“…Next, we investigated whether the effects of ASO transfection on the general transcriptome of LECs or BECs were consistent between the two negative control ASOs by comparing them to the untransfected reference CAGE-Seq samples (Additional file 2 , refer to the original study [ 18 ] for further details). For the comparison, we considered only genes displaying a | log2FC| > 1 and an FDR corrected P -value < 0.05.…”

“…Based on these results, we therefore decided to exclude negative control B CAGE-Seq libraries from further analyses and only use negative control A to investigate the lncRNA candidate knockdown effects on the transcriptome of either LECs or BECs (Fig. 1 b, refer to the original study [ 18 ] for further details).…”

“…To explore the functional role of lncRNAs in LECs or BECs, we performed antisense oligonucleotide-mediated knockdown (ASOKD) of four lncRNA candidates, previously identified as LEC- or BEC-specific lncRNAs, followed by CAGE-Seq [ 18 ]. Here, we present the early quality control steps adopted in the analysis pipeline prior to determining the transcriptional changes after lncRNA target KD in either LECs or BECs.…”

The Vital Roles of LINC00662 in Human Cancers

“…Accumulating research has revealed that many lncRNAs play vital roles in cancer tumorigenicity ( Yari et al, 2019 ; Feng et al, 2020 ) and progression ( Zhu et al, 2017 ; Gong et al, 2020 ), are aberrantly expressed in a variety of cancers, and participate in many biological functions, such as cell proliferation ( Ducoli et al, 2021 ; Shen et al, 2021 ), epithelial-to-mesenchymal transition (EMT) ( Grelet et al, 2017 ; Zhao et al, 2017 ), invasion ( Schmidt et al, 2016 ; Jin et al, 2019 ), migration ( Liu L. et al, 2019 ; Zhao et al, 2018 ), angiogenesis ( Zhao et al, 2020 ; Li et al, 2021 ), and differentiation ( Luo et al, 2019 ; Tong et al, 2020 ). Moreover, lncRNAs can also function as competing endogenous RNAs (ceRNAs) ( Wang et al, 2017 ; Zhou et al, 2019 ), transcription factors ( Long et al, 2017 ; Ducoli et al, 2021 ), and in post-transcriptional processing ( Zhang et al, 2018 ; Botti et al, 2019 ). All these findings suggest that lncRNAs may be able to serve as potential cancer biomarkers, ameliorating some of the current problems caused by cancer.…”