The MYC-regulated lncRNA LNROP (ENSG00000254887) enables MYC-driven cell proliferation by controlling the expression of OCT2

Abstract

MYC controls most of the non-coding genome. Several long noncoding transcripts were originally identified in the human B cell line P496-3 and then shown to be required for MYC-driven proliferation of Burkitt lymphoma-derived RAMOS cells. In this study, we used RAMOS cells exclusively as a representative of the human B cell lineage. One of the MYC-controlled lncRNAs required for RAMOS cell proliferation is ENSG00000254887 which we will term LNROP (long non-coding regulator of POU2F2). In the genome, LNROP is located in close proximity of POU2F2, the gene encoding OCT2. OCT2 is a transcription factor with important roles in sustaining the proliferation of human B cells. Here we show that LNROP is a nuclear RNA and a direct target of MYC. Downregulation of LNROP attenuates the expression of OCT2. This effect of LNROP on the expression of OCT2 is unidirectional as downregulation of OCT2 does not alter the expression of LNROP. Our data suggest that LNROP is a cis-acting regulator of OCT2. To illustrate the downstream reach of LNROP, we chose a prominent target of OCT2, the tyrosine phosphatase SHP-1. Downregulation of OCT2 elevates the expression of SHP-1. Our data suggest the following path of interactions: LNROP enables the proliferation of B cells by positively and unidirectionally regulating the growth-stimulatory transcription factor OCT2. In actively proliferating B cells, OCT2 attenuates the expression and anti-proliferative activity of SHP-1.

S-Phase induced RNAs control ORC1 engagement to H2A.Z and firing of early DNA replication origins

“…Gain of function was applied using the CRISPR activator (CRISPRa, CRISPR-VP64) system [33]. To this purpose the gRNA-2 was delivered in stably expressing dCas9-VP64 HEK 293, wherein c-MYC is not actively replicating as in K562 [34, 35]. This led to robust up-regulation of c-MYC ANCORs lasting up to five weeks, paralleled by increase in c-MYC expression.…”

Long Non-Coding RNAs as “MYC Facilitators”

“…LNROP (long non-coding regulator of POU2F2) is a transcript that was identified in a CRISPRi screen of MYC-regulated lncRNAs essential for B cell proliferation [ 84 ]. In the genome, it is situated in an antisense orientation to POU2F2, which is the gene that encodes the transcription factor OCT2 (octamer-binding protein 2).…”

“…(3) They are required for sustained MYC-driven cell proliferation. (4) They are not immediately essential for cell survival [ 46 , 58 , 84 ]. A summary of these characteristics is presented in Figure 1 .…”

“…In the human cell lines P496-3 and Ramos, more than 500 MYC facilitators have been identified [ 82 , 84 ]. Although both cell lines are nominally human B cells, their populations of facilitator lncRNAs, though overlapping, are not identical.…”

“…The salient point of this hypothesis is that the regulatory activity of MYC gains precision in promoter targeting through complex formation with a lncRNA. In the human cell lines P496-3 and Ramos, more than 500 MYC facilitators have been identified [82,84]. Although both cell lines are nominally human B cells, their populations of facilitator lncRNAs, though overlapping, are not identical.…”