Staufen2 functions in Staufen1-mediated mRNA decay by binding to itself and its paralog and promoting UPF1 helicase but not ATPase activity

Abstract

Staufen (STAU)1-mediated mRNA decay (SMD) is a posttranscriptional regulatory mechanism in mammals that degrades mRNAs harboring a STAU1-binding site (SBS) in their 3′-untranslated regions (3′ UTRs). We show that SMD involves not only STAU1 but also its paralog STAU2. STAU2, like STAU1, is a double-stranded RNA-binding protein that interacts directly with the ATP-dependent RNA helicase up-frameshift 1 (UPF1) to reduce the half-life of SMD targets that form an SBS by either intramolecular or intermolecular base-pairing. Compared with STAU1, STAU2 binds ∼10-fold more UPF1 and ∼two-to fivefold more of those SBS-containing mRNAs that were tested, and it comparably promotes UPF1 helicase activity, which is critical for SMD. STAU1-or STAU2-mediated augmentation of UPF1 helicase activity is not accompanied by enhanced ATP hydrolysis but does depend on ATP binding and a basal level of UPF1 ATPase activity. Studies of STAU2 demonstrate it changes the conformation of RNA-bound UPF1. These findings, and evidence for STAU1−STAU1, STAU2−STAU2, and STAU1−STAU2 formation in vitro and in cells, are consistent with results from tethering assays: the decrease in mRNA abundance brought about by tethering siRNA-resistant STAU2 or STAU1 to an mRNA 3′ UTR is inhibited by downregulating the abundance of cellular STAU2, STAU1, or UPF1. It follows that the efficiency of SMD in different cell types reflects the cumulative abundance of STAU1 and STAU2. We propose that STAU paralogs contribute to SMD by "greasing the wheels" of RNA-bound UPF1 so as to enhance its unwinding capacity per molecule of ATP hydrolyzed.protein-protein interactions | protein-RNA interactions M ammalian cells contain two Staufen (STAU) paralogs, STAU1 and STAU2. Each derives from a separate gene that produces multiple protein isoforms via alternative pre-mRNA splicing and/or polyadenylation. Every STAU1 and STAU2 isoform contains multiple conserved dsRNA-binding domains (RBDs), of which dsRBD3 and dsRBD4 constitute the major if not sole active dsRNA-binding sites (1, 2).Although STAU1 and probably STAU2 are expressed ubiquitously, STAU2 is most abundant in heart and brain (1, 3). Both paralogs are present in ribonucleoprotein particles (RNPs) within neuronal cell bodies and dendrites (1, 4-7). Both are also involved in the microtubule-dependent transport of RNAs to dendrites of polarized neurons (5, 8), presumably via the tubulinbinding domain (2). Moreover, both paralogs play a crucial role in the formation and maintenance of the dendritic spines of hippocampal neurons and are required for synaptic plasticity and memory (7, 9-11), although the detection of paralog-specific RNA granules in the distal dendrites of rat hippocampal neurons has been used to argue for paralog-distinct functions (1). Additional support for paralog-specific roles derives from the demonstration using rat hippocampal slice cultures that STAU1 but not STAU2 functions in the late phase of forskolin-induced longterm potentiation (10-12), whereas STAU2 but not STAU1 is involved in metabotropic ...

A post-translational regulatory switch on UPF1 controls targeted mRNA degradation

“…S4I). Hence, ATP hydrolysis, rather than simply ATP binding, leads to dissociation of UPF1 from RNA, in keeping with the hypothesis that UPF1 is a helicase that translocates along RNA in a 59-to-39 direction (Bhattacharya et al 2000;Chamieh et al 2008;Chakrabarti et al 2011;Shigeoka et al 2012;Park et al 2013). We suggest that the large decrease in FRET observed in the presence of ATP is due to movement of UPF1 away from the donor fluorophore attached to RNA and/or dissociation of UPF1 from the 39 end of donor-labeled RNA (see Supplemental Fig.…”

Staufen1 senses overall transcript secondary structure to regulate translation

“…Which activity is observed depends on the cellular context, the identity of the bound RNA and the location of the binding site on the target RNA. Many of Staufen's previously documented activities parallel those of the EJC [7][8][9][10][11]19,20,33,[35][36][37][38] . To better understand EJC function, we recently determined the complete EJC RNA-binding landscape in HEK293 cells 24 .…”

“…Like many RNA-binding factors, the Drosophila and mammalian Staufen proteins have been implicated in multiple post-transcriptional processes including alternative splicing 13 , RNA localization 4,6,[30][31][32] , translational activation 7 and translation-dependent mRNA decay [8][9][10][11]14,20,33,34 . Which activity is observed depends on the cellular context, the identity of the bound RNA and the location of the binding site on the target RNA.…”

“…In flies, 3′ UTR-bound Staufen is required for proper localization and translational control of bicoid and prospero mRNAs during oogenesis 4,5 . In mammals, Stau1 has been implicated in mRNA transport to neuronal dendrites 6 , regulation of translation via physical interaction with the ribosome 7 , a form of translation-dependent mRNA degradation known as Staufen-mediated decay (SMD) [8][9][10][11] , regulation of stress-granule homeostasis 12 , alternative splicing, nuclear export and translation of a gene containing 3′-UTR CUG-repeat expansions 13 . Although Stau1 is not essential for mammalian development, neurons lacking Stau1 have dendritic spinemorphogenesis defects in vitro, and knockout mice have locomotor-activity deficits 14 .…”

The Pumilio-domain protein PUF6 contributes to SIDER2 retroposon-mediated mRNA decay in Leishmania

“…"Tethering" approaches have been used successfully to isolate proteins that activate mRNA turnover in human cells (Clement and Lykke-Andersen 2008;Park et al 2013;Yokoshi et al 2014) but also in parasites ). Here, we report for the first time in trypanosomatid protozoa, the use of the MS2 coat protein tethering system to capture RNA-binding proteins (RBPs) regulating mRNA decay.…”