Many gene expression factors contain repetitive phosphorylation sites for single kinases, but the functional significance is poorly understood. Here we present evidence for hyperphosphorylation as a mechanism allowing UPF1, the central factor in nonsense-mediated decay (NMD), to increasingly attract downstream machinery with time of residence on target mRNAs. Indeed, slowing NMD by inhibiting late-acting factors triggers UPF1 hyperphosphorylation, which in turn enhances affinity for factors linking UPF1 to decay machinery. Mutational analyses reveal multiple phosphorylation sites contributing to different extents to UPF1 activity with no single site being essential. Moreover, the ability of UPF1 to undergo hyperphosphorylation becomes increasingly important for NMD when downstream factors are depleted. This hyperphosphorylation-dependent feedback mechanism may serve as a molecular clock ensuring timely degradation of target mRNAs while preventing degradation of non-targets, which, given the prevalence of repetitive phosphorylation among central gene regulatory factors, may represent an important general principle in gene expression.

Recent studies have shown that a subset of nucleoporins (Nups) can detach from the nuclear pore complex and move into the nuclear interior to regulate transcription. One such dynamic Nup, called Nup98, has been implicated in gene activation in healthy cells and has been shown to drive leukemogenesis when mutated in patients with acute myeloid leukemia (AML). Here we show that in hematopoietic cells, Nup98 binds predominantly to transcription start sites to recruit the Wdr82-Set1A/COMPASS (complex of proteins associated with Set1) complex, which is required for deposition of the histone 3 Lys4 trimethyl (H3K4me3)-activating mark. Depletion of Nup98 or Wdr82 abolishes Set1A recruitment to chromatin and subsequently ablates H3K4me3 at adjacent promoters. Furthermore, expression of a Nup98 fusion protein implicated in aggressive AML causes mislocalization of H3K4me3 at abnormal regions and up-regulation of associated genes. Our findings establish a function of Nup98 in hematopoietic gene activation and provide mechanistic insight into which Nup98 leukemic fusion proteins promote AML.

Nuclear pore complexes (NPCs) emerged as nuclear transport channels in eukaryotic cells ∼1.5 billion years ago. While the primary role of NPCs is to regulate nucleo-cytoplasmic transport, recent research suggests that certain NPC proteins have additionally acquired the role of affecting gene expression at the nuclear periphery and in the nucleoplasm in metazoans. Here we identify a widely expressed variant of the transmembrane nucleoporin (Nup) Pom121 (named sPom121, for "soluble Pom121") that arose by genomic rearrangement before the divergence of hominoids. sPom121 lacks the nuclear membrane-anchoring domain and thus does not localize to the NPC. Instead, sPom121 colocalizes and interacts with nucleoplasmic Nup98, a previously identified transcriptional regulator, at gene promoters to control transcription of its target genes in human cells. Interestingly, sPom121 transcripts appear independently in several mammalian species, suggesting convergent innovation of Nup-mediated transcription regulation during mammalian evolution. Our findings implicate alternate transcription initiation as a mechanism to increase the functional diversity of NPC components.