mRNA decapping is a critical step in eukaryotic cytoplasmic mRNA turnover. Cytoplasmic mRNA decapping is catalyzed by Dcp2 in conjunction with its co-activator Dcp1, and is stimulated by decapping enhancer proteins. mRNAs associated with the decapping machinery can assemble into cytoplasmic mRNP granules called processing bodies (PBs). Evidence suggests that PB-associated mRNPs are translationally repressed and can be degraded or stored for subsequent translation. However, whether mRNP assembly into a PB is important for translational repression, decapping or decay has remained controversial. Here we discuss the regulation of decapping machinery recruitment to specific mRNPs and how their assembly into PBs is governed by the relative rates of translational repression, mRNP multimerization and mRNA decay.

In mammalian cells, mRNAs with AU-rich elements (AREs) are targeted for translational silencing and rapid degradation. Here we present evidence that in human cells the proteins Tristetraprolin (TTP) and BRF-1 deliver ARE-mRNAs to processing bodies (PBs), cytoplasmic assemblies of mRNAs, and associated factors that promote translational silencing and mRNA decay. First, depletion of endogenous TTP and BRF proteins, or overexpression of dominant-negative mutant TTP proteins, impairs the localization of reporter ARE-mRNAs in PBs. Second, TTP and BRF-1 localize tethered mRNAs to PBs. Third, TTP can nucleate PB formation on untranslated mRNAs even when other mRNAs are trapped in polysomes by cycloheximide treatment. ARE-mRNA localization in PBs is mediated by the TTP N-and C-terminal domains and occurs downstream from mRNA polysome release, which in itself is not sufficient for mRNA PB localization. The accumulation of ARE-mRNAs in PBs is strongly enhanced when the mRNA decay machinery is rendered limiting by mRNA decay enzyme depletion or TTP/BRF-1 overexpression. Based on these observations, we propose that the PB functions as a reservoir that sequesters ARE-mRNAs from polysomes, thereby silencing ARE-mRNA function even when mRNA decay is delayed. This function of the PB can likely be extended to other mRNA silencing pathways, such as those mediated by microRNAs, premature termination codons, and mRNA deadenylation.[Keywords: mRNA turnover; AU-rich elements; TTP; BRF-1; processing bodies] Supplemental material is available at http://www.genesdev.org.

Summary Cellular mRNAs exist in messenger ribonucleoprotein (mRNP) complexes, which undergo transitions during the lifetime of the mRNAs and direct posttranscriptional gene regulation. A final posttranscriptional step in gene expression is the turnover of the mRNP, which involves degradation of the mRNA and recycling of associated proteins. How tightly associated protein components are released from degrading mRNPs is unknown. Here we demonstrate that the ATPase activity of the RNA helicase Upf1 allows disassembly of mRNPs undergoing nonsense-mediated mRNA decay (NMD). In the absence of Upf1 ATPase activity, partially degraded NMD mRNA intermediates accumulate in complex with NMD factors and concentrate in processing bodies. Thus, disassembly and completion of turnover of mRNPs undergoing NMD requires ATP hydrolysis by Upf1. This uncovers a previously unappreciated and potentially regulated step in mRNA decay, and raises the question of how other mRNA decay pathways release protein components of substrate mRNPs.

Faithful execution of developmental gene expression programs occurs at multiple levels and involves many different components such as transcription factors, histone-modification enzymes, and mRNA processing proteins. Recent evidence suggests that nucleoporins, well known components that control nucleo-cytoplasmic trafficking, have wide-ranging functions in developmental gene regulation that potentially extend beyond their role in nuclear transport. Whether the unexpected role of nuclear pore proteins in transcription regulation, which initially has been described in fungi and flies, also applies to human cells is unknown. Here we show at a genome-wide level that the nuclear pore protein NUP98 associates with developmentally regulated genes active during human embryonic stem cell differentiation. Overexpression of a dominant negative fragment of NUP98 levels decreases expression levels of NUP98-bound genes. In addition, we identify two modes of developmental gene regulation by NUP98 that are differentiated by the spatial localization of NUP98 target genes. Genes in the initial stage of developmental induction can associate with NUP98 that is embedded in the nuclear pores at the nuclear periphery. Alternatively, genes that are highly induced can interact with NUP98 in the nuclear interior, away from the nuclear pores. This work demonstrates for the first time that NUP98 dynamically associates with the human genome during differentiation, revealing a role of a nuclear pore protein in regulating developmental gene expression programs.