FUS binds the CTD of RNA polymerase II and regulates its phosphorylation at Ser2

Howard Hughes Medical Institute.
Genes & development (Impact Factor: 10.8). 12/2012; 26(24):2690-5. DOI: 10.1101/gad.204602.112
Source: PubMed


Mutations in the RNA-binding protein FUS (fused in sarcoma)/TLS have been shown to cause the neurodegenerative disease amyotrophic lateral sclerosis (ALS), but the normal role of FUS is incompletely understood. We found that FUS binds the C-terminal domain (CTD) of RNA polymerase II (RNAP2) and prevents inappropriate hyperphosphorylation of Ser2 in the RNAP2 CTD at thousands of human genes. The loss of FUS leads to RNAP2 accumulation at the transcription start site and a shift in mRNA isoform expression toward early polyadenylation sites. Thus, in addition to its role in alternative RNA splicing, FUS has a general function in orchestrating CTD phosphorylation during RNAP2 transcription.

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    • "that FUS can affect CTD phosphorylation in vitro (Schwartz et al. 2012). Thus, nascent RNA would stimulate fibrous assembly of FUS, which in turn would attenuate CTD phosphorylation and suppress transcriptional activity in vivo. "
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    ABSTRACT: More than half of all human genes produce prematurely terminated polyadenylated short mRNAs. However, the underlying mechanisms remain largely elusive. CLIP-seq (cross-linking immunoprecipitation [CLIP] combined with deep sequencing) of FUS (fused in sarcoma) in neuronal cells showed that FUS is frequently clustered around an alternative polyadenylation (APA) site of nascent RNA. ChIP-seq (chromatin immunoprecipitation [ChIP] combined with deep sequencing) of RNA polymerase II (RNAP II) demonstrated that FUS stalls RNAP II and prematurely terminates transcription. When an APA site is located upstream of an FUS cluster, FUS enhances polyadenylation by recruiting CPSF160 and up-regulates the alternative short transcript. In contrast, when an APA site is located downstream from an FUS cluster, polyadenylation is not activated, and the RNAP II-suppressing effect of FUS leads to down-regulation of the alternative short transcript. CAGE-seq (cap analysis of gene expression [CAGE] combined with deep sequencing) and PolyA-seq (a strand-specific and quantitative method for high-throughput sequencing of 3' ends of polyadenylated transcripts) revealed that position-specific regulation of mRNA lengths by FUS is operational in two-thirds of transcripts in neuronal cells, with enrichment in genes involved in synaptic activities. © 2015 Masuda et al.; Published by Cold Spring Harbor Laboratory Press.
    Genes & development 05/2015; 29(10):1045-57. DOI:10.1101/gad.255737.114 · 10.80 Impact Factor
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    • "Based on these experiments, we propose that the absence of ELAV leads to inefficient Ubx RNA processing and retention of RNA at the site of transcription, with a consequential reduction in Ubx mRNA release from DNA and lower Ubx protein formation, suggesting that ELAV-dependent Ubx RNA processing could ‘fine-tune’ the levels of expression of Ubx within the nervous system. This model is consistent with our data and with previous reports on other systems (including human beta-globin, glyceraldehyde 3-phosphate dehydrogenase, ribosomal protein L3, DNA damage-inducible transcript 3) indicating that non-canonical RNA processing reactions usually prevent mRNA flow to the cytoplasm by tethering processed mRNAs to DNA around transcription sites (Custodio et al., 1999, 2007; Schwartz et al., 2012). "
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    ABSTRACT: The regulated head-to-tail expression of Hox genes provides a coordinate system for the activation of specific programmes of cell differentiation according to axial level. Recent work indicates that Hox expression can be regulated via RNA processing but the underlying mechanisms and biological significance of this form of regulation remain poorly understood. Here we explore these issues within the developing Drosophila central nervous system (CNS). We show that the pan-neural RNA-binding protein (RBP) ELAV (Hu antigen) regulates the RNA processing patterns of the Hox gene Ultrabithorax (Ubx) within the embryonic CNS. Using a combination of biochemical, genetic and imaging approaches we demonstrate that ELAV binds to discrete elements within Ubx RNAs and that its genetic removal reduces Ubx protein expression in the CNS leading to the respecification of cellular subroutines under Ubx control, thus defining for the first time a specific cellular role of ELAV within the developing CNS. Artificial provision of ELAV in glial cells (a cell type that lacks ELAV) promotes Ubx expression, suggesting that ELAV-dependent regulation might contribute to cell type-specific Hox expression patterns within the CNS. Finally, we note that expression of abdominal A and Abdominal B is reduced in elav mutant embryos, whereas other Hox genes (Antennapedia) are not affected. Based on these results and the evolutionary conservation of ELAV and Hox genes we propose that the modulation of Hox RNA processing by ELAV serves to adapt the morphogenesis of the CNS to axial level by regulating Hox expression and consequently activating local programmes of neural differentiation.
    Development 05/2014; 141(10):2046-56. DOI:10.1242/dev.101519 · 6.46 Impact Factor
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    • "Most recently, FUS was shown to directly bind NEAT1 (16), providing a basis for physical association of the protein with paraspeckles. Interestingly, FUS shares many similarities with paraspeckle proteins, namely RNA/DNA binding capacity, involvement in chromosomal translocations leading to malignancies (24,25), interaction with C-terminal domain of RNA polymerase II (26,27) and redistribution to the perinucleolar region upon transcription inhibition (15,28). Although paraspeckles are absent in neurons under basal conditions, their formation at the early stages of ALS, triggered by increased synthesis of NEAT1, was recently demonstrated (16), suggesting participation of paraspeckles in response to neuronal stress or damage. "
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    ABSTRACT: Paraspeckles are nuclear bodies formed by a set of specialised proteins assembled on the long non-coding RNA NEAT1; they have a role in nuclear retention of hyperedited transcripts and are associated with response to cellular stress. Fused in sarcoma (FUS) protein, linked to a number of neurodegenerative disorders, is an essential paraspeckle component. We have shown that its recruitment to these nuclear structures is mediated by the N-terminal region and requires prion-like activity. FUS interacts with p54nrb/NONO, a major constituent of paraspeckles, in an RNA-dependent manner and responds in the same way as other paraspeckle proteins to alterations in cellular homeostasis such as changes in transcription rates or levels of protein methylation. FUS also regulates NEAT1 levels and paraspeckle formation in cultured cells, and FUS deficiency leads to loss of paraspeckles. Pathological gain-of-function FUS mutations might be expected to affect paraspeckle function in human diseases because mislocalised ALS-linked FUS variants sequester other paraspeckle proteins into aggregates formed in cultured cells and into neuronal inclusions in a transgenic mouse model of FUSopathy. Furthermore, we detected abundant p54nrb/NONO-positive inclusions in motor neurons of patients with familial forms of ALS caused by FUS mutations, but not in other ALS cases. Our results suggest that both loss and gain of FUS function can trigger disruption of paraspeckle assembly, which may impair protective responses in neurons and thereby contribute to the pathogenesis of FUSopathies.
    Human Molecular Genetics 12/2013; 23(9). DOI:10.1093/hmg/ddt622 · 6.39 Impact Factor
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