Takayuki Nojima

Takayuki Nojima
Kyushu University | Kyudai · Medical Institute of Bioregulation - MIB Hospital

PhD
RNA polymerase II, Transcription termination, RNA processing, LncRNA transcription, Nascent RNA

About

45
Publications
10,025
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2,361
Citations
Citations since 2017
33 Research Items
1818 Citations
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Introduction
Reveal nascent transcription coupled to RNA processing on protein coding and lncRNA genes.
Additional affiliations
February 2021 - present
Kyushu University
Position
  • Professor (Associate)
Description
  • Group leader
September 2010 - February 2021
University of Oxford
Position
  • Senior Researcher

Publications

Publications (45)
Article
Full-text available
Transcription is a highly dynamic process. Consequently, we have developed native elongating transcript sequencing technology for mammalian chromatin (mNET-seq), which generates single-nucleotide resolution, nascent transcription profiles. Nascent RNA was detected in the active site of RNA polymerase II (Pol II) along with associated RNA processing...
Article
Full-text available
Numerous long intervening noncoding RNAs (lincRNAs) are generated from the mammalian genome by RNA polymerase II (Pol II) transcription. Although multiple functions have been ascribed to lincRNAs, their synthesis and turnover remain poorly characterized. Here, we define systematic differences in transcription and RNA processing between protein-codi...
Article
Full-text available
The highly intronic nature of protein coding genes in mammals necessitates a co-transcriptional splicing mechanism as revealed by mNET-seq analysis. Immunoprecipitation of MNase-digested chromatin with antibodies against RNA polymerase II (Pol II) shows that active spliceosomes (both snRNA and proteins) are complexed to Pol II S5P CTD during elonga...
Article
Full-text available
Extensive tracts of the mammalian genome that lack protein-coding function are still transcribed into long noncoding RNA. While these lncRNAs are generally short lived, length restricted, and non-polyadenylated, how their expression is distinguished from protein-coding genes remains enigmatic. Surprisingly, depletion of the ubiquitous Pol-II-associ...
Article
Full-text available
Mammalian chromatin is the site of both RNA polymerase II (Pol II) transcription and coupled RNA processing. However, molecular details of such co-transcriptional mechanisms remain obscure, partly because of technical limitations in purifying authentic nascent transcripts. We present a new approach to characterize nascent RNA, called polymerase int...
Article
Full-text available
CDK9 is a kinase critical for the productive transcription of protein-coding genes by RNA polymerase II (pol II). As part of P-TEFb, CDK9 phosphorylates the carboxyl-terminal domain (CTD) of pol II and elongation factors, which allows pol II to elongate past the early elongation checkpoint (EEC) encountered soon after initiation. We show that, in a...
Article
Full-text available
Mammalian genomes express two principal gene categories through RNA polymerase II-mediated transcription: protein-coding transcription units and non-coding RNA transcription units. Non-coding RNAs are further divided into relatively abundant structural RNAs, such as small nuclear RNAs, and into a myriad of long non-coding RNAs (lncRNAs) of often lo...
Preprint
Full-text available
CDK9 is a critical kinase required for the productive transcription of protein-coding genes by RNA polymerase II (pol II) in higher eukaryotes. Phosphorylation of targets including the elongation factor SPT5 and the carboxyl-terminal domain (CTD) of RNA pol II allows the polymerase to pass an early elongation checkpoint (EEC), which is encountered...
Article
Transcription termination in eukaryotic cells involves the recognition of polyadenylation signals (PAS) that signal the site of pre-mRNA cleavage and polyadenylation. Most eukaryotic genes contain multiple PAS that are used by alternative polyadenylation (APA), a co-transcriptional process that increases transcriptomic diversity and modulates the f...
Article
Gene transcription occurs via a cycle of linked events, including initiation, promoter-proximal pausing, and elongation of RNA polymerase II (Pol II). A key question is how transcriptional enhancers influence these events to control gene expression. Here, we present an approach that evaluates the level and change in promoter-proximal transcription...
Preprint
Full-text available
Mammalian chromatin is the site of both RNA polymerase II (Pol II) transcription and coupled RNA processing. However, molecular details of such co-transcriptional mechanisms remain obscure, partly due to technical limitations in purifying authentic nascent transcripts. We present a new approach to purify and profile nascent RNA, called Polymerase I...
Article
Full-text available
Herpes Simplex virus (HSV) protein ICP27 is a multifunctional gene expression regulator, which assumes various roles during the course of the viral infection. Previously we identified ICP27 switches RNA isoforms expression of particular genes such as PML (promyelocytic leukemia). Although ICP27 protein contains several RNA binding domains, it remai...
Article
Full-text available
Cyclin-dependent kinase 12 (CDK12) phosphorylates the carboxyl-terminal domain (CTD) of RNA poly-merase II (pol II) but its roles in transcription beyond the expression of DNA damage response genes remain unclear. Here, we have used TT-seq and mNET-seq to monitor the direct effects of rapid CDK12 inhibition on transcription activity and CTD phospho...
Preprint
Full-text available
Gene transcription occurs via a cycle of linked events including initiation, promoter proximal pausing and elongation of RNA polymerase II (Pol II). A key question is how do transcriptional enhancers influence these events to control gene expression? Here we have used a new approach to quantify transcriptional initiation and pausing in vivo, while...
Article
Full-text available
Accurate regulation of mRNA termination is required for correct gene expression. Here, we describe a role for SCAF4 and SCAF8 as anti-terminators, suppressing the use of early, alternative polyadenylation (polyA) sites. The SCAF4/8 proteins bind the hyper-phosphorylated RNAPII C-terminal repeat domain (CTD) phosphorylated on both Ser2 and Ser5 and...
Article
Full-text available
The pervasive nature of RNA polymerase II (Pol II) transcription requires efficient termination. A key player in this process is the cleavage and polyadenylation (CPA) factor PCF11, which directly binds to the Pol II C-terminal domain and dismantles elongating Pol II from DNA in vitro. We demonstrate that PCF11-mediated termination is essential for...
Data
List of upregulated and downregulated genes following SPT6 depletion in nuclear poly(A)+ RNA-seq.
Data
Table S2. Intergenic DNA Replication Origins versus mNET-Seq Signals, Related to Figure 7 List of intergenic replication origins overlapping with basal or increased nascent transcription in control or SPT6 depletion conditions
Preprint
Full-text available
The pervasive nature of RNA polymerase II (Pol II) transcription requires efficient termination. A key player in this process is the cleavage and polyadenylation (CPA) factor PCF11, which directly binds to the Pol II C-terminal domain and dismantles elongating Pol II from DNA in vitro. We demonstrate that PCF11-mediated termination is essential for...
Data
Datasets containing enriched proteins identified in two replicates of IPs with antibodies to S2P, S5P and T4P CTD. Proteins are ranked according to fold change (FC) relative to mock IP.
Article
Full-text available
Mitochondria are descendants of endosymbiotic bacteria and retain essential prokaryotic features such as a compact circular genome. Consequently, in mammals, mitochondrial DNA is subjected to bidirectional transcription that generates overlapping transcripts, which are capable of forming long double-stranded RNA structures1,2. However, to our knowl...
Data
Excel spreadsheet of mNET-seq gene regions used for Meta-profile generation and quantitation results.
Article
Full-text available
Influenza virus intimately associates with host RNA polymerase II (Pol II) and mRNA processing machinery. Here, we use mammalian native elongating transcript sequencing (mNET-seq) to examine Pol II behavior during viral infection. We show that influenza virus executes a two-pronged attack on host transcription. First, viral infection causes decreas...
Article
Full-text available
The 5' splice site mutation (IVS20+6T>C) of the inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene in familial dysautonomia (FD) is at the sixth intronic nucleotide of the 5' splice site. It is known to weaken U1 snRNP recognition and result in aberrantly spliced mRNA product in neuronal t...
Data
Table S2. Supporting Gene Lists Used for Genomic Analyses and PCA, Related to Experimental Procedures Dataset 1 is a list of all genomic regions used for the analyses in Figures 1-4. Many overlapping genes and genes with low expression in HeLa cells were excluded (Supplemental Experimental Procedures). Dataset 2 is a list of all lincRNA genes used...
Article
Full-text available
The transcription cycle of RNA polymerase II (Pol II) correlates with changes to the phosphorylation state of its large subunit C-terminal domain (CTD). We recently developed Native Elongation Transcript sequencing using mammalian cells (mNET-seq), which generates single-nucleotide-resolution genome-wide profiles of nascent RNA and co-transcription...
Article
Full-text available
Significance Familial dysautonomia (FD) is caused by missplicing of the IκB kinase complex-associated protein (IKAP) gene, which results in the skipping of exon 20, especially in neurons. FD would be treatable if exon 20 inclusion were increased correctly to reestablish correct splicing. Here, we have established a dual-color splicing reporter that...
Article
Full-text available
Mammalian RNA polymerase II (Pol II) transcription termination is an essential step in protein-coding gene expression that is mediated by pre-mRNA processing activities and DNA-encoded terminator elements. Although much is known about the role of pre-mRNA processing in termination, our understanding of the characteristics and generality of terminat...
Chapter
Multicolor fluorescent reporters are useful tools to visualize patterns of alternative splicing (AS) in cultured cells and in living organisms at a single-cell resolution. The multicolor reporters have been utilized to search for cis-elements and trans-acting factors involved in the regulation of AS, and to screen for chemical compounds affecting t...
Article
Full-text available
Since alternative splicing of pre-mRNAs is essential for generating tissue-specific diversity in proteome, elucidating its regulatory mechanism is indispensable to understand developmental process or tissue-specific functions. We have been focusing on tissue-specific regulation of mutually exclusive selection of alternative exons because this impli...
Article
Full-text available
Viruses use alternative splicing to produce a broad series of proteins from small genomes by utilizing the cellular splicing machinery. Since viruses use cellular RNA binding proteins for viral RNA processing, it is presumable that the splicing of cellular pre-mRNAs is affected by viral infection. Here, we showed that herpes simplex virus type 2 (H...
Article
Eukaryotic gene expression is controlled in multi steps. Pre-messenger RNAs (pre-mRNAs) transcribed by RNA polymerase II (RNAPII) are capped at 5' end, spliced to remove introns and polyadenylated at 3' end. These processes are followed by other post-transcriptional steps that include mRNA export, translation and mRNA degradation. Recent studies ha...
Article
Full-text available
The removal of intervening sequences from transcripts is catalyzed by the spliceosome, a multicomponent complex that assembles on the newly synthesized pre-mRNA. Pre-mRNA translation in the cytoplasm leads to the generation of aberrant proteins that are potentially harmful. Therefore, tight control to prevent undesired pre-mRNA export from the nucl...
Article
Full-text available
RNA export factor (REF) is a component of the exon junction complex (EJC) that is deposited on mRNA in a splicing-dependent manner, and targets spliced mRNA for export. In this study, analysis of the RNA-binding protein complexes revealed that REF associates with beta-globin mRNA at the region other than the EJC deposition site. Comparison between...

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