ライフサイエンスコーパス: NELF

1 NELF and DSIF act together to inhibit transcription elon

2 NELF and DSIF collaborate to inhibit elongation by RNA p

3 NELF antisense experiments indicate that a reduction in

4 NELF causes Pol II to pause in the promoter-proximal reg

5 NELF colocalizes with RNAP II, and its level increases f

6 NELF depletion also delayed the dissociation of HSF from

7 NELF interacts with Pcf11, a transcription termination f

8 NELF is expressed in PNS and CNS tissues during embryoni

9 NELF-associated pausing of Pol II might be an obligatory

10 NELF-B is a BRCA1-interacting protein and subunit (with

11 NELF-B is predicted to form a HEAT repeat fold, also bin

12 NELF-E mRNA, encoding a regulator of pol II-mediated tra

13 NELF-mediated stalling of RNAPII also attenuates transcr

14 nsisting of eIF4E-Homologous Protein (4EHP), NELF-E, the 40S ribosome, and eIF3 subunits.

15 addition, we identify NCoR1-GPS2-HDAC3 as a NELF-interacting corepressor complex that is associated

16 bition of transcription, we did not detect a NELF-RNA contact when the nascent transcript was between

17 We identify a NELF core subcomplex formed by conserved regions in subu

18 Although NELF is essential for embryonic development, its role in

19 Although NELF is involved in promoter proximal pausing, depletion

20 F4 regulatory network consisting of 4EHP and NELF-E that impacts proteostasis during normal developme

21 Using a new competition assay, NELF-A and NELF-B are each shown to act independently as competitiv

22 of GR-regulated transactivation, NELF-A and NELF-B, relative to other factors in the overall gene in

23 equired for full activity of both NELF-A and NELF-B.

24 by conserved regions in subunits NELF-A and NELF-C, and resolve its crystal structure.

25 d throughout their lengths, while NELF-A and NELF-E contain nonconserved regions inserted between con

26 mediated phosphorylation of Spt5, NELF-A and NELF-E results in the dissociation of NELF from Pol II,

27 1, whereas SEC recruits P-TEFb to NELF-A and NELF-E via Paf1c and Med26, respectively.

28 Pol II through interaction with CBC-ARS2 and NELF/DSIF, and promotes 3' end processing by enhancing r

29 The amino acid sequences of NELF-B and NELF-D are highly conserved throughout their lengths, wh

30 s extensive colocalization of the NELF-B and NELF-D subunits at hundreds of interbands.

31 of mutagenized libraries of GFP-binding and NELF-E-binding aptamers to their respective targets and

32 ELF, reverse the negative effect of DSIF and NELF and simultaneously facilitate the action of TFIIF.

33 gation, including components of the DSIF and NELF complexes.

34 ough a capping-independent block of DSIF and NELF loading.

35 However, the mechanism by which DSIF and NELF participate in setting up the paused Pol II remains

36 Our results show that DSIF and NELF require a nascent transcript longer than 18 nt to s

37 EFb reverses the negative effect of DSIF and NELF through a mechanism dependent on its kinase activit

38 DSIF and NELF were associated with engaged Pol II and, in additio

39 Although DSIF and NELF were both required for inhibition of transcription,

40 The association of DSIF and NELF with initiated RNA Polymerase II (Pol II) is the ge

41 at TFIIF functionally competes with DSIF and NELF, and this competition is dependent on the relative

42 g the elongation complex containing DSIF and NELF, reverse the negative effect of DSIF and NELF and s

43 mbryos containing reduced levels of DSIF and NELF.

44 egion of genes and is stabilized by DSIF and NELF.

45 vidually for subunits of Mediator, DSIF, and NELF that that do not interact with DNA directly, but ra

46 and EGR3 showed similar decreased TFII-I and NELF-A INR binding and increased RNAPII SerP2 in the gen

47 We found that integrator and NELF, an RNA polymerase II pausing protein, were associa

48 l link between PARP-1, ADP-ribosylation, and NELF.

49 ay induced by negative factors Spt5/Spt4 and NELF, which is overcome by the positive factor P-TEFb (C

50 complex, along with pausing factors SPT5 and NELF-A, at the intragenic CTCF-cohesin binding sites.

51 p distinct from those controlled by Spt5 and NELF.

52 d that IFN-induced recruitment of P-TEFb and NELF/DSIF was under the control of BRD4.

53 on the relative concentrations of TFIIF and NELF.

54 single-stranded nucleic acids in vitro, and NELF-C associates with RNA in vivo.

55 ze pausing, but not all kinds of pausing are NELF-mediated.

56 COBRA1 (also known as NELF-B) is a BRCA1-binding protein that regulates RNA po

57 ive transcription elongation factors such as NELF, DSIF, and factor 2.

58 Using a new competition assay, NELF-A and NELF-B are each shown to act independently as

59 APII) past transcription end sites (TESs) at NELF depletion-induced genes.

60 al for antiviral immunity in insects because NELF and P-TEFb are required to restrict viral replicati

61 However, the role of DSIF in pausing beyond NELF recruitment remains unclear.

62 4) show that upon activation, eRNAs can bind NELF and are necessary for its transient removal from pr

63 pposite face of the NELF-AC subcomplex binds NELF-B.

64 cific IgG and IgA antibody responses in both NELF and blood against both vaccine homologous and varia

65 d that is required for full activity of both NELF-A and NELF-B.

66 s raises the possibility that RNA binding by NELF is not necessary in promoter-proximal pausing.

67 ular senescence-enriched regulon impacted by NELF and SPT6.

68 KSHV OriLytL-K7 lytic genes is inhibited by NELF during latency, but can also be promptly reactivate

69 results reveal an earlier stage, mediated by NELF, when repression occurs at the HIV LTR.

70 rget genes were, like Hsp70, up-regulated by NELF-depletion, whereas the majority of target genes sho

71 transcriptional activator, HSF, might cause NELF to dissociate from the elongation complex.

72 Cohesin, NELF, and Spt5 pausing and elongation factor knockdown e

73 and the negative elongation factor complex (NELF).

74 ymerase II (Pol II) and the pausing complex, NELF and DSIF, are detected near the transcription start

75 Consequently, NELF knockdown resulted in significant reduction in DNA

76 Cultured NELF-B UtcKO stromal cells exhibited perturbances in ext

77 Decreasing NELF also correlated with displacement of a positioned n

78 Decreasing NELF expression overcomes RNAP II pausing to enhance HIV

79 Depleting NELF increased processive HIV transcription and replicat

80 hromatin immunoprecipitation analyses detect NELF at the promoters of the hsp70 and beta1-tubulin gen

81 Drosophila NELF has four subunits similar to subunits of human NELF

82 Here, further characterization of Drosophila NELF is provided.

83 We analyzed the interactions among DSIF, NELF, and a reconstituted Drosophila Pol II elongation c

84 iator), pausing, and elongation (e.g., DSIF, NELF, PAF, and P-TEFb).

85 lectron microscopy structures of Pol II-DSIF-NELF complexes with NELF in two distinct conformations c

86 DSIF/NELF inhibits early transcript elongation until it is co

87 nascent RNA, TFIIS inhibition may help DSIF/NELF negatively regulate productive transcription.

88 These two activities of DSIF/NELF appear to be mechanistically distinct.

89 rt a previously undescribed activity of DSIF/NELF, namely inhibition of the transcript cleavage facto

90 mics of interactions between HIV-1 Tat, DSIF/NELF, and the transcription complexes actively engaged i

91 Our results demonstrate that DSIF/NELF associates with RNA pol II complexes during early t

92 FP and negative elongation factor subunit E (NELF-E) proteins with their corresponding canonical and

93 A common motif in each NELF was identified that is required for full activity o

94 Knockdown of either NELF or DSIF results in an increase in the levels of uaR

95 egatively regulate transcription elongation, NELF, Spt5, and Pcf11.

96 Stable knockdown of endogenous NELF-B has the opposite effects on an exogenous gene.

97 Reduction of the endogenous NELF proteins in breast cancer cells using small interfe

98 These findings establish NELF/ELOA-dependent mechanisms regulating transcriptiona

99 As expected, NELF and DSIF increased pausing, and P-TEFb promoted pau

100 ecruitment of the negative elongation factor NELF at start sites.

101 th the polymerase negative elongation factor NELF.

102 that includes the negative elongation factor NELF.

103 blation of the primary pause-inducing factor NELF does not increase expression of lineage markers, bu

104 knockout of the Pol II pause-inducing factor NELF in immature germ cells blocks differentiation to sp

105 rough knockdown of the pause-inducing factor NELF leads to broadly attenuated immune gene activation.

106 The pausing factor NELF is known to induce and stabilize pausing, but not a

107 y the complex of negative elongation factor (NELF) and 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazo

108 Negative elongation factor (NELF) and 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazo

109 Negative elongation factor (NELF) and DRB sensitivity inducing factor (DSIF) contrib

110 IF together with negative elongation factor (NELF) associates with RNA polymerase II during early elo

111 occupancy of the negative elongation factor (NELF) at the p21(CIP1) promoter, although the level of b

112 ed by the host's negative elongation factor (NELF) at the promoter regions of OriLytL-K7 lytic genes

113 alization of the negative elongation factor (NELF) complex and facilitated by PU.1.

114 t release of the negative elongation factor (NELF) complex and productive elongation.

115 depletion of the Negative Elongation Factor (NELF) complex increases both flu transcription and inter

116 The four-subunit negative elongation factor (NELF) complex mediates RNA polymerase II (Pol II) pausin

117 a decoy for the negative elongation factor (NELF) complex upon induction of immediate early genes (I

118 mediated by the negative elongation factor (NELF) complex, allows cells to coordinate and appropriat

119 -E) of the human negative elongation factor (NELF) complex, which participates in RNA polymerase II p

120 e pause-inducing negative elongation factor (NELF) complex.

121 ed member of the negative elongation factor (NELF) complex.

122 The principal negative elongation factor (NELF) contains four polypeptides and requires for activi

123 hanisms by which negative elongation factor (NELF) establishes and maintains HIV latency.

124 ause-stabilizing negative elongation factor (NELF) in macrophages did not affect baseline Pol II occu

125 The role of the negative elongation factor (NELF) in maintaining HIV latency was investigated follow

126 Negative elongation factor (NELF) induces RNAP II promoter proximal pausing and limi

127 he negative transcription elongation factor (NELF) inhibits basal transcription from the long termina

128 Negative elongation factor (NELF) is a critical transcriptional regulator that stabi

129 The human negative elongation factor (NELF) is a four-subunit protein complex that inhibits th

130 The Negative Elongation Factor (NELF) is a transcription regulatory complex that induces

131 thway, including negative elongation factor (NELF) that pauses RNA polymerase II (Pol II) and positiv

132 ns, depletion of negative elongation factor (NELF), a key factor in setting up paused Pol II, reduced

133 tes and inhibits negative elongation factor (NELF), a protein complex that regulates promoter-proxima

134 actor (DSIF) and negative elongation factor (NELF), act as negative transcription elongation factors

135 ere we show that negative elongation factor (NELF), an RNA polymerase II (Pol II) pausing factor, coo

136 ng complex (CBC)-Negative elongation factor (NELF), components of CBs, interferes with CBs' associati

137 nit of the human negative elongation factor (NELF), directly binds to ERalpha and represses ERalpha-m

138 e release of the negative elongation factor (NELF), thus stabilizing Pol II promoter-proximal pausing

139 actors (BRD4 and negative elongation factor (NELF)-E) and to define their sites and mechanisms of act

140 s (DSIF) and the negative elongation factor (NELF).

141 actor (DSIF) and Negative Elongation Factor (NELF).

142 d the inhibitory negative elongation factor (NELF).

143 es the 4-subunit negative elongation factor (NELF).

144 ttributed to the negative elongation factor (NELF).

145 deletion in the nasal embryonic LHRH factor (NELF) gene in pedigree 1 and an additional heterozygous

146 l factor termed nasal embryonic LHRH factor (NELF) that was discovered in a differential screen of mi

147 The pausing factors NELF and DSIF are associated with these antisense polyme

148 vity of TTF2, and influenced pausing factors NELF and DSIF, but did not affect the function of TFIIS

149 factors we describe are the pausing factors--NELF (negative elongation factor) and DSIF (DRB sensitiv

150 Nasal epithelial lining fluid (NELF) and plasma were collected before and after vaccina

151 Following NELF-E knockdown or tumor necrosis factor alpha (TNF-alp

152 This work thus uncovers a key role for NELF-mediated pausing in establishing the responsiveness

153 r associates with 39% of the genes that have NELF.

154 Our results define how NELF can support pausing, reactivation, and elongation b

155 discern the functional architecture of human NELF through X-ray crystallography, protein crosslinking

156 obtain a genome-wide understanding of human NELF-mediated transcriptional regulation in vivo, we car

157 s four subunits similar to subunits of human NELF.

158 triggering an interferon response, identify NELF as an important interface with the flu life cycle,

159 Thus, our studies implicate NELF functioning in early elongation complexes distinct

160 Importantly, NELF condensation is required for cellular viability und

161 ion of hsp70 results in a marked decrease in NELF at the hsp70 promoter.

162 nse experiments indicate that a reduction in NELF expression decreases olfactory axon outgrowth and t

163 of negative elongation factors that include NELF and DSIF.

164 Overall, our study indicates NELF-mediated pausing is essential to coordinate endomet

165 Ablation of individual NELF subunits destabilizes the NELF complex and causes c

166 mall interfering RNA knockdown of individual NELF subunits.

167 Instead, NELF depletion increased the utilization of downstream t

168 iously observed in fission yeast (which lack NELF).

169 ression and DSB repair in germ cells lacking NELF.

170 Indeed, ESCs lacking NELF have dramatically attenuated FGF/ERK activity, rend

171 Like NELF-associated genes, most BEAF-associated genes are hi

172 Likewise, NELF and DSIF prior to stimulation were hardly detectabl

173 ption initiation and by assisting in loading NELF onto Pol II after initiation.

174 ponents: GR < Cdk9 < BRD4 </= induced gene < NELF-E.

175 Mechanistically, NELF is associated with TCF1 and recruited preferentiall

176 rticoid receptor (GR), reporter, TIF2, NCoR, NELF-A, sSMRT, and STAMP) using our recently developed c

177 In contrast, no NELF or polymerase is detectible near the IP-10 promoter

178 shock induction, DSIF and polymerase but not NELF were strongly recruited to chromosomal puffs harbor

179 wn of the transcription factor SPT5, but not NELF-E, also gives rise to a specific inhibition of HSV-

180 gion (IDR) in NELFA is necessary for nuclear NELF condensation and can be functionally replaced by th

181 We now report new activities of NELF-B and other NELF complex subunits, which are to att

182 tion for genetic and biochemical analysis of NELF in Drosophila.

183 However, novel actions of BRD4 and of NELF-E in GR-controlled gene induction have been uncover

184 A shRNA-based knockdown assay of NELF revealed that it negatively regulates the passage o

185 TFII-I knockdown decreased the binding of NELF-A, a component of the promoter-proximal pausing com

186 sis to quantify the formation of clusters of NELF proteins in living, stressed cells.

187 est a diverse transcriptional consequence of NELF-mediated RNAPII pausing in the human genome.

188 cute and long-term molecular consequences of NELF loss and reveals a role for Pol II pausing in the p

189 microarray analysis of S2 cells depleted of NELF and discovered that NELF RNAi affects many rapidly

190 Instead, depletion of NELF delayed the time taken for these genes to shut off

191 Consequently, depletion of NELF expression induced transition of stalled RNAPII int

192 Here, we show that upon rapid depletion of NELF, RNA Pol II fails to be released into gene bodies,

193 Upon depletion of NELF-A, -C, or -E, the vast majority of NELF-regulated g

194 sequestered in the cytoplasm and deprived of NELF nuclear function still support cell proliferation a

195 Genetic disruption of NELF in macrophages enhanced transcription of AP-1-encod

196 -A and NELF-E results in the dissociation of NELF from Pol II, thereby transiting transcription from

197 orylating DSIF that leads to dissociation of NELF.

198 highly activated by the combined effects of NELF-E depletion and activation of initiation by TNF-alp

199 strict checkpoint for Cdk9, the evolution of NELF was likely critical to enable increased regulation

200 Elevated expression of NELF-E and KAT2B is associated with poorer prognosis in

201 A positively charged face of NELF-AC is involved in RNA binding, whereas the opposite

202 architecture and three RNA-binding faces of NELF.

203 t an important and rate-limiting function of NELF in anti-tumor immunity.

204 ne, eiger, displayed all of the hallmarks of NELF-dependent polymerase stalling.

205 Immunodepletion of NELF also impairs promoter proximal pausing on the hsp70

206 Immunodepletion of NELF or DSIF from a nuclear extract desensitizes transcr

207 1 in gastric cancer cells was independent of NELF-E.

208 mics analysis revealed that the knockdown of NELF-E or 4EHP commonly reduced several subunits of the

209 lated with and probably results from loss of NELF association and function.

210 Accordingly, loss of NELF-E leads to impaired SLUG binding on chromatin.

211 n of NELF-A, -C, or -E, the vast majority of NELF-regulated genes were down-regulated.

212 ners as a function of EMT and a co-option of NELF-E with the key EMT transcription factor SLUG.

213 3 domain of Spt5 mediates the recruitment of NELF to the elongation complex.

214 Moreover, reduction of NELF-E, 4EHP, RpS12, eIF3l, or eIF3h suppressed the expr

215 nism of P-TEFb recruitment and regulation of NELF/DSIF during transcription is not fully understood.

216 ronal enhancers impairs transient release of NELF from the specific target promoters during transcrip

217 E) further reveals a significant rewiring of NELF-E-associated chromatin partners as a function of EM

218 establish a previously unappreciated role of NELF in constraining transcription of inflammation inhib

219 The amino acid sequences of NELF-B and NELF-D are highly conserved throughout their

220 To determine the full spectrum of NELF target genes in Drosophila, we performed a microarr

221 response to the initial decidual stimulus of NELF-B UtcKO was similar to that of control mice; howeve

222 ferase, KAT2B, as a key functional target of NELF-E-SLUG.

223 Surprisingly, only one-third of NELF target genes were, like Hsp70, up-regulated by NELF

224 or mutation of the ADP-ribosylation sites on NELF-E promotes Pol II pausing, providing a clear functi

225 Depletion of one NELF subunit in salivary glands using RNA interference a

226 Critically, only NELF-mediated pausing establishes a strict requirement f

227 used small interfering to suppress NELF-B or NELF-E subunits in cultured human endometrial stromal ce

228 ow report new activities of NELF-B and other NELF complex subunits, which are to attenuate glucocorti

229 als that recruitment of COBRA1 and the other NELF subunits to endogenous ERalpha-responsive promoters

230 The paused NELF state supports Pol II stalling, whereas the poised

231 -B subunit in the mouse uterus using PgrCre (NELF-B UtcKO).

232 the expression of EMT markers, phenocopying NELF ablation.

233 ependent kinase 9 (CDK9)-that phosphorylates NELF and the carboxyl terminal domain of Pol II-and enri

234 Further, the poised NELF state can accommodate TFIIS binding to Pol II, allo

235 supports Pol II stalling, whereas the poised NELF state enables transcription elongation as it does n

236 nexpectedly, independent of CDK9, a presumed NELF kinase.

237 orchestrates efficient pausing by recruiting NELF to promoters before transcription initiation and by

238 nd Spt5, establishes the pause by recruiting NELF to the elongation complex.

239 nascent transcript and subsequently recruits NELF.

240 Condensate formation requires NELF dephosphorylation and SUMOylation induced by stress

241 P-TEFb-mediated phosphorylation of Spt5, NELF-A and NELF-E results in the dissociation of NELF fr

242 o binds RNA in vivo, and anchors the subunit NELF-E, which is confirmed to bind RNA in vivo.

243 plex formed by conserved regions in subunits NELF-A and NELF-C, and resolve its crystal structure.

244 Here we summarize NELF's roles in promoter-proximal pausing, transcription

245 ility, we used small interfering to suppress NELF-B or NELF-E subunits in cultured human endometrial

246 Surprisingly, NELF associates with almost one-half of the most highly

247 Long-term NELF or SPT6 depletion causes reversible growth arrest f

248 e LTR showed that pol II was paused and that NELF depletion released pol II.

249 lity, leading to the prevailing concept that NELF-mediated Pol II pausing is essential for cell proli

250 These results demonstrate that NELF plays a role as a common guidance molecule for olfa

251 omatin immunoprecipitation demonstrated that NELF, a negative transcription elongation factor, was as

252 2 cells depleted of NELF and discovered that NELF RNAi affects many rapidly inducible genes involved

253 We find that NELF depletion results in global loss of cap-binding com

254 Here, we find that NELF-depleted Drosophila melanogaster cells functionally

255 The finding that NELF also associates with the promoter regions of wingle

256 Lastly, our study indicates that NELF regulates alternative transcription initiation of B

257 most highly expressed genes, indicating that NELF is not necessarily a repressor of gene expression.

258 We propose that NELF and DSIF cause polymerase to pause in the promoter

259 We propose that NELF-mediated pausing allows Pol II to direct CBP-mediat

260 Here we report that NELF rapidly forms nuclear condensates upon stress in hu

261 ChIP assays reveal that NELF-B diminishes GR recruitment to promoter regions of

262 ray chip [ChIP-chip] analysis) revealed that NELF is concentrated at the 5' ends of 2,111 genes in Dr

263 analyses and myofiber cultures revealed that NELF works at a specific stage of regeneration whereby i

264 n vivo protein-DNA cross-linking showed that NELF and DSIF associate with the promoter region before

265 -stem-cell-specific deletion, we showed that NELF is required for efficient muscle regeneration and s

266 Chromatin immunoprecipitation showed that NELF knockdown led to dissociation of RNAPII from the pr

267 ted the cross-linking result and showed that NELF, DSIF, and RNA polymerase IIa colocalize at the hsp

268 Mounting evidence suggests that NELF association with RNA Pol II serves as checkpoint fo

269 The NELF-AC subcomplex binds single-stranded nucleic acids i

270 We deleted the NELF-B subunit in the mouse uterus using PgrCre (NELF-B

271 of individual NELF subunits destabilizes the NELF complex and causes cell lethality, leading to the p

272 ase pausing, in contrast to depletion of the NELF (negative elongation factor) pausing complex.

273 ction in a manner that is independent of the NELF complex.

274 found to bind to NELF-E, a component of the NELF complex.

275 nctions with TEAD to regulate binding of the NELF negative elongation factor and block SMAD2,3 induct

276 tic relationship between the presence of the NELF pausing factor and positioning of the +1 nucleosome

277 NA binding, whereas the opposite face of the NELF-AC subcomplex binds NELF-B.

278 osomes shows extensive colocalization of the NELF-B and NELF-D subunits at hundreds of interbands.

279 g small hairpin RNA (shRNA) knockdown of the NELF-E subunit, a condition that induced high levels of

280 ions additively affected the affinity of the NELF-E-binding aptamer, whose interaction depended mainl

281 n together, we uncover a crucial role of the NELF-E-SLUG-KAT2B epigenetic axis in breast cancer carci

282 nogaster cells functionally recapitulate the NELF-independent pausing we previously observed in fissi

283 Finally, we observe that the NELF-A tentacle interacts with the RPB2 protrusion and i

284 get gene transcription and interact with the NELF complex.

285 BRD4 is also found to bind to NELF-E, a component of the NELF complex.

286 and Tat-SF1, whereas SEC recruits P-TEFb to NELF-A and NELF-E via Paf1c and Med26, respectively.

287 modulators of GR-regulated transactivation, NELF-A and NELF-B, relative to other factors in the over

288 Unexpectedly, NELF-E modifies GR induction in a manner that is indepen

289 f the heat shock response remain intact upon NELF loss.

290 enes showed decreased expression levels upon NELF RNAi.

291 use is influenced by the timing between when NELF loads onto Pol II and how fast Pol II escapes the p

292 rved transcription elongation factor whereas NELF is mostly restricted to the metazoan lineage.

293 We propose a model in which NELF recruits Pcf11 and NCoR1-GPS2-HDAC3 to paused RNAP

294 ly conserved throughout their lengths, while NELF-A and NELF-E contain nonconserved regions inserted

295 hat haploinsufficiency of SLBP and/or WHSC2 (NELF-A) contributes to several novel cellular phenotypes

296 se pause-release and a transient genome-wide NELF dissociation from chromatin, unexpectedly, independ

297 determine if paused Pol II colocalized with NELF.

298 tructures of Pol II-DSIF-NELF complexes with NELF in two distinct conformations corresponding to paus

299 Forty-six of 56 genes with NELF were found to have paused Pol II.

300 BRCA1-interacting protein and subunit (with NELF-A, -C/D, and -E) of the human negative elongation f