ライフサイエンスコーパス: Claspin

1 ese potential phosphorylation sites in human Claspin.

2 reduction in the Chk1-activating potency of Claspin.

3 R (Xatr) and Xenopus Rad17 (Xrad17), but not Claspin.

4 e purification and characterization of human Claspin.

5 d17, the Rad9-Hus1-Rad1 (9-1-1) complex, and Claspin.

6 and presentation of Chk1 to this complex by Claspin.

7 eine and by immunodepletion of either ATR or Claspin.

8 sitive checkpoint pathway containing ATR and Claspin.

9 romises the ability of Chk1 to interact with Claspin.

10 ts kinase-domain and of its partnership with Claspin.

11 n dramatically diminished phosphorylation of Claspin.

12 quence repeats in the Chk1-binding domain of Claspin.

13 by accelerating the proteolytic turnover of claspin.

14 and interaction of MutLalpha with TopBP1 and Claspin.

15 Chk1, but not p53, is strongly stimulated by Claspin.

16 kinase Chk1 depends on the mediator protein Claspin.

17 ndependent of checkpoint mediators Tipin and Claspin.

18 s by preventing the interaction of Chk1 with Claspin, a Chk1 adaptor protein that is required for Chk

19 CLASPIN, a critical player in replication fork stabiliza

20 We have identified Claspin, a novel protein that binds to Xenopus Chk1 (Xch

21 recently identified N-terminal DBD of human Claspin, a presumptive homolog of yeast Mrc1 proteins.

22 s ATR-to-Chk1 signaling by promoting loss of Claspin, a protein that assists ATR to phosphorylate Chk

23 rylation sites in the Chk1-binding domain of Claspin abolish its ability to mediate ATR phosphorylati

24 Under such adaptation-defective conditions, Claspin accumulates on chromatin at high levels, and Chk

25 Without Claspin, activated ATR-ATRIP phosphorylated Chk1 weakly

26 adducts stimulates ATR kinase activity, and Claspin acts synergistically with damaged DNA to increas

27 Claspin also induced significant autophosphorylation of

28 We show that Claspin also participates in the detection of chromosoma

29 increases the amount and phosphorylation of Claspin, an activator of Chk1 phosphorylation.

30 This process induces the stabilization of Claspin, an activator of the DNA-damage checkpoint, and

31 ve despite DNA damage, and protein levels of claspin, an adaptor of ataxia telangiectasia-mutated and

32 k1 kinase also requires its association with Claspin, an adaptor protein essential for Chk1 protein s

33 gulation of Chk1 and its interacting partner Claspin, an adaptor protein that is required for the pho

34 We show that BRCA1 ubiquitylates claspin, an essential coactivator of the CHK1 checkpoint

35 horylation of the checkpoint adaptor protein Claspin and activation of the Chk1 effector kinase, both

36 romatin with DSBs, whereas depletion of both Claspin and BRCA1 completely abolishes this activation.

37 e induces the formation of a complex between Claspin and BRCA1, a second regulator of Chk1 activation

38 RK function and previously unknown roles for Claspin and Chk1 as negative regulators of DNA replicati

39 e data indicate that the interaction between Claspin and Chk1 is complex.

40 where And-1 promotes the interaction between Claspin and Chk1, thereby stimulating efficient Chk1 act

41 In addition, purified Claspin and DDK are capable of a direct in vitro interac

42 ), are essential for the interaction between Claspin and DDK.

43 USP20 in turn deubiquitinates and stabilizes Claspin and enhances the activation of ATR-Chk1 signalin

44 phosphorylations promote binding of Chk1 to Claspin and ensuing activation of Chk1 by ATR.

45 MutSalpha-Claspin and MutLalpha-Claspin interactions were not demo

46 und that phosphorylated Rad17 interacts with Claspin and regulates its phosphorylation.

47 y, phosphorylation of Rad1 is independent of Claspin and the Rad9 carboxy terminus, both of which are

48 gulation of the DNA-damage mediator proteins Claspin and TopBP1, impaired DNA-damage-induced dissocia

49 d interaction of MutSalpha with ATR, TopBP1, Claspin, and Chk1 and interaction of MutLalpha with TopB

50 hat several, including ATRIP, RPA70, TopBP1, Claspin, and CINP, are required for efficient HSV-1 repl

51 eractions with the mediator proteins TopBP1, Claspin, and Timeless (Tim).

52 , including several mediators, such as Mdc1, Claspin, and TopBP1.

53 G domain mutation have been used to identify Claspin as a new target of BRCA1 E3 ligase activity in r

54 oupled with mass spectrometry, we identified Claspin as a novel Cdh1-interacting protein and further

55 Our findings identify claspin as an in vivo substrate for the BRCA1 E3 ligase

56 Our results identify Claspin as the most upstream molecule in the signaling p

57 We show here that Claspin associates with chromatin in a regulated manner

58 Claspin associates with replication forks upon origin un

59 Timeless, a Claspin-associating protein, is also required for effici

60 Claspin, ATR, and Rad17 all bind to chromatin independen

61 Claspin, ATR, and Rad17 may collaborate in checkpoint re

62 More importantly, down-regulation of Claspin augments the premature chromatin condensation in

63 show that, during this checkpoint response, Claspin becomes phosphorylated on threonine 906 (T906),

64 In Xenopus, phosphorylated Claspin binds to xChk1 and regulates xChk1 activation in

65 t it functions to promote phosphorylation of Claspin bound Chk1 by the ataxia-telangiectasia and Rad-

66 unction requires the DNA replication protein Claspin but not ATR.

67 A1 is required for the activation of the ATR-Claspin-Chk1 and ATR-H2AX pathways following UV treatmen

68 t for Claspin function and the regulation of Claspin-Chk1 interaction in human cells.

69 e phosphorylation sites on Claspin inhibited Claspin-Chk1 interaction in vivo, impaired Chk1 activati

70 The ATR-Claspin-Chk1 pathway is critical for turning on the cell

71 -dependent function of Rad17 in an ATR-Rad17-Claspin-Chk1-signaling cascade that responds to specific

72 ctivation of Cdh1 leads to activation of the Claspin/Chk1 pathway.

73 E(2) also inhibits Claspin:Chk1 protein association via AKT phosphorylation

74 s that DNA-PKcs is required to maintain Chk1-Claspin complex stability and transcriptional regulation

75 eraction stabilizes Timeless-Tipin and Tipin-Claspin complexes on RPA-coated ssDNA and in doing so pr

76 We also find that a fragment of Claspin containing the Chk1-binding domain together with

77 We show that Claspin contains a replication fork-interacting domain (

78 We also find that Claspin contains a small Chk1-activating domain (residue

79 The activated form of Claspin contains two repeated phosphopeptide motifs that

80 position on Chk1 of the phosphate group from Claspin corresponds to the location of activation-loop p

81 tiple components of the SCF(beta-TrCP)-based claspin degradation machinery were found deregulated in

82 on of components of the SCF(beta-TrCP)-based claspin degradation machinery.

83 s and promotes mitotic entry by accelerating claspin degradation through a mechanism that involves de

84 say conditions, nonetheless has some role in Claspin-dependent activation.

85 Interestingly, Claspin-depleted cells retained significant levels of Ch

86 However, Claspin-depleted egg extracts that have been reconstitut

87 Furthermore, Claspin-depleted extracts are unable to arrest the cell

88 We report that Claspin-depleted HeLa and HCT116 cells display levels of

89 rmination events with a frequency similar to CLASPIN depletion, resulting in excessive endogenous DNA

90 no difference in the expression level of the claspin deubiquitinating enzyme USP7 was detected.

91 In this process, Claspin dissociates from chromatin, and Chk1 undergoes i

92 Overall, these results indicate that Claspin docks with a phosphate-binding site in the catal

93 ated by Chk1, suggesting that Chk1 regulates Claspin during checkpoint response.

94 amage, we further explored the exact role of Claspin during Chk1 activation following replication str

95 CK1gamma1 phosphorylates the CKAD of Claspin efficiently in vitro, and depletion of CK1gamma1

96 Importantly, restoration of CLASPIN expression in USP9X-depleted cells partially sup

97 ology, we have shown that down-regulation of Claspin expression inhibits Chk1 activation in response

98 In this study, we show that Claspin expression is downregulated by the proteasome-me

99 stability and transcriptional regulation of Claspin expression.

100 f breast cancer cells inhibited survivin and claspin expression.

101 Expression of Claspin fluctuates in a cell cycle-dependent manner, but

102 Immunodepletion of Claspin from egg extracts abolishes both the phosphoryla

103 Significantly, removal of Claspin from egg extracts only partially abrogates the a

104 in repeats in human Claspin is important for Claspin function and the regulation of Claspin-Chk1 inte

105 317 and Ser345, raising the possibility that Claspin function during normal fork progression may exte

106 Based on these findings, we propose that Claspin functions at late stages of Chk1 activation foll

107 ermore, we show that c-Rel directly controls Claspin gene transcription.

108 Unexpectedly, we found that Claspin has a second, positive role in control of the ce

109 Since Claspin has also been shown to participate in Chk1 activ

110 These results suggest that Claspin has properties of both a tumor suppressor and an

111 9X regulated the expression and stability of CLASPIN in an S-phase-specific manner.

112 e data imply a potentially critical role for Claspin in replication checkpoint control in mammalian c

113 tudied the respective roles of ATR-ATRIP and Claspin in the activation of Chk1.

114 sent study we investigated the role of human Claspin in the DNA damage/replication checkpoint in mamm

115 The function of Claspin in this DSB-triggered pathway depends on phospho

116 this study, we found that DDK phosphorylates Claspin in vitro and forms a nuclear complex containing

117 a nuclear complex containing Cdc7, Drf1, and Claspin in Xenopus egg extracts.

118 We found that mutant forms of Claspin incapable of interacting with DDK are still able

119 Mutation of these phosphorylation sites on Claspin inhibited Claspin-Chk1 interaction in vivo, impa

120 ics approach and identified USP9X as a novel CLASPIN-interacting protein.

121 MutSalpha-Claspin and MutLalpha-Claspin interactions were not demonstrable with purified

122 In Xenopus, Claspin interacts with Chk1 after DNA damage through a r

123 Moreover, Claspin interacts with the checkpoint proteins ATR and R

124 of global replication fork progression, and Claspin interacts with the replication machinery and mig

125 We observed that human Claspin is a cell cycle regulated protein that peaks at

126 Claspin is a checkpoint protein involved in ATR (ataxia

127 Claspin is a Chk1-interacting protein that participates

128 Claspin is a homolog of Mrc1, a checkpoint protein requi

129 Claspin is a newly identified protein that regulates Chk

130 acting protein and further demonstrated that Claspin is a novel Cdh1 ubiquitin substrate.

131 , albeit to a lesser extent, suggesting that Claspin is a universal requirement for high replication

132 horylation of Chk1 when the mediator protein Claspin is also tethered to the DNA with TopBP1.

133 Claspin is an essential protein for the ATR-dependent ac

134 Taken together, these findings indicate that Claspin is an essential upstream regulator of Xchk1.

135 The mediator protein Claspin is critical for the activation of the checkpoint

136 rom the DNA replication checkpoint response, Claspin is degraded in a betaTrCP-dependent manner.

137 Claspin is essential for the ATR-dependent activation of

138 The checkpoint mediator protein Claspin is essential for the ATR-dependent activation of

139 Claspin is essential for the ATR-dependent activation of

140 phosphorylation of Claspin repeats in human Claspin is important for Claspin function and the regula

141 The checkpoint mediator protein Claspin is indispensable for the ATR-dependent phosphory

142 Phosphorylation of Claspin is mediated by Plk1 and is essential for binding

143 Claspin is necessary for the ATR-dependent activation of

144 us, the SCFbetaTrCP-dependent degradation of Claspin is necessary for the efficient and timely termin

145 Claspin is not only necessary for the propagation of the

146 Chk1 activation after hydroxyurea treatment, Claspin is only required to sustain Chk1 activation.

147 We show that Xenopus Claspin is phosphorylated at the MBT at both DNA replica

148 In addition, we found that Thr-916 on Claspin is phosphorylated by Chk1, suggesting that Chk1

149 In vivo, Claspin is phosphorylated in a canonical DSGxxS degron s

150 In addition, Claspin is phosphorylated in response to replication str

151 We found that Claspin is phosphorylated in vivo at Thr-916 in response

152 vestigate, for the first time, whether human Claspin is required for high rates of replication fork p

153 udy, we have shown that the human homolog of Claspin is required for resistance to multiple forms of

154 Claspin is required for the ATR-dependent activation of

155 ain conserved in the yeast Mrc1 orthologs of Claspin is sufficient for its mediator activity.

156 Yeast Mrc1, ortholog of metazoan Claspin, is both a central component of normal DNA repli

157 e Saccharomyces cerevisiae ortholog of human Claspin, is facilitated by the SCFDia2 ubiquitin ligase

158 ctivation of ATR but also, through Tipin and Claspin, it plays an important role in the action of ATR

159 A1 E3 inactivation decreases chromatin-bound claspin levels and impairs homology-directed DNA repair

160 s and their upstream kinase IKK can regulate Claspin levels by controlling its mRNA expression.

161 hibition of Chk1 activity by UCN01 decreases Claspin levels in cells.

162 Conversely, overexpression of Chk1 increases Claspin levels.

163 ntial for Chk1 phosphorylation, indicating a Claspin-like but distinct role for APE2 in ATR-Chk1 sign

164 Claspin localizes in the nuclei, but it only associates

165 responds to the N/C ratio and indicate that Claspin may also respond to an independent timer to trig

166 These findings suggest that Claspin may be a component of the replication ensemble a

167 Phosphorylated Claspin may mimic an activating phosphorylation of Chk1

168 Claspin may then be one of the phosphoproteins through w

169 the deubiquitylating enzyme Usp28 to permit Claspin-mediated activation of Chk1 in response to DNA d

170 Here we have reconstituted a Claspin-mediated checkpoint system with purified human p

171 on RPA-coated ssDNA and in doing so promotes Claspin-mediated phosphorylation of Chk1 by ATR.

172 ngs suggest that the interaction of DDK with Claspin mediates a checkpoint-independent function of Cl

173 Claspin mediates the activation of CHK1 by ATR in respon

174 The S phase-specific adaptor protein Claspin mediates the checkpoint response to replication

175 In response to replication stress, Claspin mediates the phosphorylation and activation of C

176 During replicative stress, Claspin mediates the phosphorylation and consequent acti

177 Another protein, Claspin, mediates the activation of a cellular checkpoin

178 LT also bound the Claspin mediator protein, which normally facilitates the

179 Significantly, expression of a stable Claspin mutant unable to bind betaTrCP prolongs the acti

180 These dependencies suggest that binding of Claspin occurs around the time of initial DNA unwinding

181 interaction promotes the phosphorylation of Claspin on a nearby serine (S934) by Plx1.

182 se results indicate that stable retention of Claspin on chromatin is not necessary for activation of

183 Conversely, neither phosphorylation of Claspin on these sites nor the presence of BRCA1 is nece

184 lure to load the checkpoint mediator protein Claspin onto chromatin.

185 thesis or activate Chk1 in cells depleted of Claspin, or when Chk1 was depleted or subject to chemica

186 ositive role in control of the cell cycle as Claspin overexpression increased cell proliferation.

187 Further, in egg extracts, Claspin phosphorylation depends on a threshold N/C ratio

188 nsistent with a model in which ATR regulates Claspin phosphorylation in response to DNA damage and re

189 (Cbeta) or PP2A(Aalpha) had little effect on Claspin phosphorylation, but the amount of Claspin was r

190 These findings suggest that Claspin plays a role in monitoring DNA replication durin

191 isiae and show that Cmr1--together with Mrc1/Claspin, Pph3, the chaperonin containing TCP1 (CCT) and

192 the mechanisms involved in the regulation of Claspin protein levels have not been explored.

193 gic inhibition or knockdown of Plk1 restored claspin protein levels, Chk1 activation, and p53 stabili

194 mportantly, in response to DNA damage in G2, Claspin proteolysis is inhibited to allow the prompt ree

195 nt response, and introduce crucial roles for Claspin, Rad17 phosphorylation and the ubiquitin proteas

196 romatin enrichment of replication protein A, Claspin, Rad17-RFC, and Rad9-Rad1-Hus1 was not detected

197 ediates a checkpoint-independent function of Claspin related to DNA replication.

198 ts together indicate that phosphorylation of Claspin repeats in human Claspin is important for Claspi

199 We identified a conserved binding site on Claspin required for its interaction with DDK.

200 In vitro ubiquitylation of Claspin requires betaTrCP, Plk1, and an intact DSGxxS de

201 cate that the Plx1-dependent inactivation of Claspin results in termination of a DNA replication chec

202 e data indicate a role of Chk1 in regulating Claspin stability in the cell.

203 athway and that Chk1 is required to maintain Claspin stability.

204 BRCA1 and Claspin then function to activate the tumor suppressor C

205 knockdown of the fork stabilization protein Claspin, Timeless, or Tipin.

206 Thus, Chk1 and the Claspin-Timeless module of replication forks not only pa

207 Binding of Claspin to chromatin depends on the pre-replication comp

208 ation of this work is that stable binding of Claspin to chromatin may play a role in other functions

209 er BP1 or BP2 compromises optimal binding of Claspin to chromatin.

210 sence of BP1 has no effect on the ability of Claspin to mediate activation of Chk1.

211 The addition of Claspin to this reaction strongly stimulated the phospho

212 Binding of Claspin to Xchk1 is highly elevated in the presence of D

213 with this possibility, depletion of Chk1 and Claspin together doubled the percentage of very slow for

214 1, which does not form a stable complex with Claspin under our assay conditions, nonetheless has some

215 ave been reconstituted with these mutants of Claspin undergo DNA replication more slowly.

216 Claspin was found to control BRCA1 phosphorylation on se

217 Phosphorylation of Claspin was found to depend on the ataxia telangiectasia

218 n Claspin phosphorylation, but the amount of Claspin was reduced.

219 Expression of a nondegradable mutant of claspin was shown to inhibit mitotic entry in HPV-16 E7-

220 cause this region is also conserved in human Claspin, we investigated the regulation and function of

221 This loss of Claspin which ultimately facilitates cell proliferation

222 Once Chk1 is activated, it stabilizes Claspin, which in turn helps to maintain Chk1 activation

223 Claspin, which is stabilized by Chk1, regulates the bind

224 eckpoint kinase requires the adaptor protein Claspin, which recruits Chk1 for phosphorylation by ATR.

225 nvolves the Chk1-activating domain (CKAD) of Claspin, which undergoes phosphorylation on multiple con

226 (Cdh1) reactivation in DNA-damaged G2 cells, Claspin, which we show to be an APC/C(Cdh1) substrate in

227 colin-treated extracts containing mutants of Claspin with alanine substitutions at positions 906 or 9

228 s suggest that only transient interaction of Claspin with replication forks potentiates its Chk1-acti

229 to ssDNA and facilitates the association of Claspin with ssDNA.

230 lved in Chk1 activation, it is possible that Claspin works as an adaptor molecule bringing these mole