Glucocorticoid receptor interacts with PNRC2 in a ligand-dependent manner to recruit UPF1 for rapid mRNA degradation

Abstract

Glucocorticoid receptor (GR), which was originally known to function as a nuclear receptor, plays a role in rapid mRNA degradation by acting as an RNA-binding protein. The mechanism by which this process occurs remains unknown. Here, we demonstrate that GR, preloaded onto the 5′UTR of a target mRNA, recruits UPF1 through proline-rich nuclear receptor coregulatory protein 2 (PNRC2) in a ligand-dependent manner, so as to elicit rapid mRNA degradation. We call this process GR-mediated mRNA decay (GMD). Although GMD, nonsense-mediated mRNA decay (NMD), and staufen-mediated mRNA decay (SMD) share upstream frameshift 1 (UPF1) and PNRC2, we find that GMD is mechanistically distinct from NMD and SMD. We also identify de novo cellular GMD substrates using microarray analysis. Intriguingly, GMD functions in the chemotaxis of human monocytes by targeting chemokine (C-C motif) ligand 2 (CCL2) mRNA. Thus, our data provide molecular evidence of a posttranscriptional role of the well-studied nuclear hormone receptor, GR, which is traditionally considered a transcription factor. glucocorticoid receptor | PNRC2 | UPF1 | glucocorticoid receptor-mediated mRNA decay | Nonsense-mediated mRNA decay A t the cellular level, glucocorticoid receptor (GR), which belongs to the nuclear receptor superfamily, functions as a transcription factor regulating various physiological processes (1-3). In the presence of a glucocorticoid, which diffuses through the plasma membrane into the cytoplasm, cytosolic GR binds to the glucocorticoid. The resulting glucocorticoid-GR complex is activated and then enters the nucleus. Once in the nucleus, GR dimerizes, binds to specific cis-acting elements, and recruits coregulatory proteins for transcriptional activation or repression (4, 5).The majority of the coregulatory proteins commonly contain a nuclear receptor box (NR box, also called an LXXLL motif), which is important for interactions between coregulatory proteins and nuclear receptors (4-6). The proline-rich nuclear receptor coregulatory protein (PNRC), however, is an exception because it interacts with nuclear receptors through an SH3-binding motif [SD(E)PPSPS] rather than an NR box (7,8). Two PNRC paralogs, PNRC1 and PNRC2, have been identified in mammalian cells (7,8). PNRC1 and PNRC2 are believed to play similar roles in nuclear receptor-mediated signaling because they interact with similar groups of nuclear receptors.Although PNRC2 is known to function as a coregulatory protein for nuclear receptors, it has a distinct function in mRNA decay pathways including nonsense-mediated mRNA decay (NMD), staufen (STAU)-mediated mRNA decay (SMD), and replication-dependent histone mRNA degradation (HMD) (9-13). NMD serves as a mechanism of both mRNA quality control and posttranscriptional regulation by selectively recognizing and degrading cellular transcripts that are abnormal or that contain a premature translation termination codon (PTC), as reviewed elsewhere (14-16). A key NMD factor, UPF1, is recruited to a terminating ribosome at a PTC. UPF1 then recr...

Identification and molecular characterization of cellular factors required for glucocorticoid receptor-mediated mRNA decay

“…All of these data indicate that, unlike NMD, GMD does not require translation termination factors and occurs independently of a translation event. In support of these conclusions, our previous data showed that artificial insertion of a strong stem-loop structure into the 5 ′ UTR of GMD reporter mRNA does not affect GMD efficiency, although translational efficiency of the reporter mRNA is drastically reduced (Cho et al 2015).…”

“…Because our previous data showed that down-regulation of SMG1 does not affect GMD (Cho et al 2015), it was quite unexpected that UPF1 R -4SA failed to restore GMD in our complementation experiments (Fig. 2C).…”

“…Although all of these pathways share PNRC2 and UPF1 as common factors, GMD is mechanistically different from NMD in several respects (Cho et al 2015). First, NMD is tightly coupled to translation because an elongating ribosome has to recognize a PTC on the mRNA; in contrast, our previous data using a GMD reporter mRNA harboring a strong stem-loop structure suggest that GMD could occur independently of translation.…”

“…Although it has long been appreciated that GR functions as a DNA-binding transcription factor, several recent reports demonstrated the ability of GR to bind a subset of RNAs (Dhawan et al 2007;Kino et al 2010;Ishmael et al 2011;Dhawan et al 2012;Cho et al 2015). Unlike GR binding to DNAs, GR associates with mRNAs even in the absence of a ligand.…”

“…PNRC2 provides a binding platform for upstream frameshift 1 (UPF1) and decapping enzyme 1a (DCP1A), the latter of which is a component of the decapping complex. Eventually, the resulting complex triggers rapid degradation of target mRNAs, termed "GR-mediated mRNA decay" (GMD) (Cho et al 2015).…”