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Statements

USP8 is phosphorylated. 98 / 98
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"Furthermore, we demonstrate that optimal Usp8 tyrosine phosphorylation is not necessary for binding of Usp8 to EGFR."
23333852
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"The binding of BAP-treated UBPY to 14-3-3ε was significantly reduced compared to that of untreated UBPY, indicating that the phosphorylation of UBPY is required for its 14-3-3 binding ( Fig. 2 C)."
17720156
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"The [ 32 P] labeling experiment also suggested the presence of other phosphorylation site(s) in UBPY because the phosphorylation of UBPY S680A was still faintly detected."
17720156
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"To investigate how Usp8 tyrosine phosphorylation is regulated, we tested whether TGFα stimulation of EGFR leads to efficient Usp8 tyrosine phosphorylation."
23333852
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"We therefore tested whether enhanced recycling of EGFR in response to TGFα stimulation leads to efficient Usp8 tyrosine phosphorylation."
23333852
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"Our data support the model whereby EGF stimulation of the EGFR leads to MIT-domain dependent recruitment of Usp8 towards the activated receptor-complex at the endosomes where also the SRC kinases are [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"To test whether Src-family tyrosine kinases may be responsible for Usp8 tyrosine phosphorylation upon stimulation of EGFR-ErbB2, we used the specific Src-family kinase inhibitors PP2 and Src Inhibitor[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"However, removal of the MIT domain did not completely abolish Usp8 tyrosine phosphorylation."
21044682
sparser
"Interestingly, overexpression of constitutively active mutant SRC (SRC Y527A) significantly increases USP8 tyrosine phosphorylation ."
28923280
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"Moreover, co-expression of constitutively active SRC Y527A with EGFR and either Usp8 WT or C748A substantially increased basal and ligand-induced Usp8 tyrosine phosphorylation ( Fig. 2 A–B)."
23333852
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"Moreover, activated EGFR-ErbB2 coprecipitates with tyrosine-phosphorylated Flag-tagged Usp8, demonstrating that Usp8 binds to this chimeric receptor."
21044682
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"Our studies indicate that both constitutive and ligand-induced Usp8 tyrosine phosphorylation signal can be increased by co-expression of constitutively active SRC Y527A together with Usp8 and EGFR ( F[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"Ligand-induced EGFR-ErbB2-mediated Usp8 tyrosine phosphorylation was lower than observed for EGFR."
21044682
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"Our results showed, however, that Usp8 tyrosine phosphorylation was not decreased in any of these mutants ( Fig. 6 C–D), demonstrating that these two tyrosines are not solely responsible for the Usp8 [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"Quantification of the Usp8 tyrosine phosphorylation signal of three independent experiments indicates that Usp8 wt and its C748A mutant were similarly tyrosine phosphorylated in cells with ErbB wild-t[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"We cannot rule out that removal of the MIT-domain results in a conformational effect resulting in reduced Usp8 tyrosine phosphorylation, but this is less likely since this mutant has been extensively [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"Our current findings further demonstrate that Usp8 binds to ErbB2 and that Usp8 tyrosine phosphorylation is dependent on ErbB2- ( Fig. 4 ) and Src- kinase ( Fig. 5 ) activities, thereby extending our [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"Interestingly, the Cbl binding site of EGFR-ErbB2 is not required for Usp8 tyrosine phosphorylation and coprecipitation ( Fig. 3 ), suggesting that Usp8 might be recruited to the EGFR-ErbB2 also via m[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
rlimsp
"USP8 is a deubiquitinase (DUB) and can be phosphorylated in its RSYSS motif, leading to association with 14-3-3 protein and subsequent impairment of its DUB activity [5]."
26285834
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"Since decreased USP8 tyrosine phosphorylation is associated with enhanced endosomal recycling of EGFR when cells are stimulated by TGFα, it is likely that USP8 phosphorylation may regulate its DUB activity."
23902637
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"To test whether Y12 could be a site for Usp8 tyrosine phosphorylation, we generated a Y12F mutant."
23333852
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"We further show that Usp8 tyrosine phosphorylation upon stimulation of EGFR-ErbB2 is (a) independent of Y1091, (b) dependent on Src- and ErbB2- kinase activity, (c) enhanced upon coexpression of catal[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"Our results demonstrate that the MIT domain is required for optimal tyrosine phosphorylation of Usp8, which is in line with previous results that the MIT domain is necessary for endosomal recruitment [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"To fully understand the underlying mechanism of USP8 activation, we searched for a Tyr kinase that is responsible for the phosphorylation of USP8."
29472535
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"Activation of different tyrosine kinase receptors suppresses ciliogenesis in retinal epithelial cells by stabilizing the trichoplein-Aurora A complex following phosphorylation of the deubiquitinase USP8 [ xref ]."
32630309
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"To test whether the kinase activity of ErbB2 is required for Usp8 tyrosine phosphorylation in EGFR-ErbB2 containing cells, we used the EGFR/ErbB2-kinase inhibitor PD153035."
21044682
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"This model is supported by our current findings that TGFα stimulation of EGFR and EGF stimulation of ERBB2 [52] , conditions that are associated with enhanced endosomal recycling and decreased MVB ta[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"To address the question of the functional significance of Usp8 tyrosine phosphorylation, it is crucial to elucidate the regulation of Usp8 tyrosine phosphorylation and to identify the Usp8 tyrosine re[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"However, no effect on total Usp8 tyrosine phosphorylation was observed upon mutation of Y12 ( Fig. 6 C–D), suggesting that Y12 is not solely responsible for Usp8 tyrosine phosphorylation."
23333852
sparser
"Although the MIT domain is necessary for endosomal recruitment [30] , we cannot rule out that Usp8 tyrosine phosphorylation occurs within the MIT domain."
23333852
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"Even though the MIT domain is essential for tyrosine phosphorylation of the truncated Usp8 construct 1–504 ( Fig. 4 ), removal of the MIT domain does not completely abolish tyrosine phosphorylation of[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"Our previous results have shown that Usp8 tyrosine phosphorylation upon EGF-induced stimulation is EGFR- and Src- tyrosine kinase dependent [41] ."
21044682
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"Using specific tyrosine kinase inhibitors and kinase-dead ERBB mutants, we previously demonstrated that the DUB Usp8 is a substrate for EGFR- and ERBB2-activated SRC-family tyrosine kinases and that U[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"To determine whether Usp8 tyrosine phosphorylation is critically dependent on ErbB receptor ubiquitination by Cbl, we analyzed Usp8 tyrosine phosphorylation after EGF stimulation of cells containing t[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"For example, phosphorylation of USP8 during interphase promotes sequestration by 14-3-3 proteins ( Mizuno et al., 2007 ), whereas loss of this phosphorylation during mitosis allows USP8 recruitment to[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21596315
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"USP8 is a deubiquitinase (DUB) and can be phosphorylated in its RSYSS motif, leading to association with 14-3-3 protein and subsequent impairment of its DUB activity [ xref ]."
26285834
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"However, mutation of three MIT domain tyrosine residues did not abolish Usp8 tyrosine phosphorylation."
23333852
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"The finding that we did detect phospho-serine but no phospho-tyrosine residues by MS analysis suggests that the stoichiometry of Usp8 tyrosine phosphorylation may be very low and therefore not detecta[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"In the present study we show that overexpression of constitutively active SRC enhances constitutive and ligand-induced Usp8 tyrosine phosphorylation."
23333852
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"Here, we demonstrate, in the context of a chimeric EGFR-ErbB2 receptor, that (i) EGF induces pY1091 Cbl binding site-dependent K63-polyubiquitination of EGFR-ErbB2, (ii) Cbl is tyrosine phosphorylated upon stimulation of EGFR-ErbB2 wt and Y1091F mutant receptor, (iii) EGF-induced activation of EGFR-ErbB2 induces Usp8 tyrosine phosphorylation, and (iv) ubiquitination of the EGFR-ErbB2 wt and Y1091F mutant is enhanced upon coexpression of catalytically inactive Usp8-C748A in the presence and absence of EGF."
21044682
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"We then analyzed the roles of EGFR-mediated phosphorylation of USP8 in ciliogenesis."
29472535
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"These data suggest that USP8 phosphorylation, possibly on Tyr residue(s), enhances its DUB activity."
29472535
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"As already shown in Fig. 5 A–B, deletion of the complete MIT-domain (Usp8 Δ140) resulted in substantially decreased Usp8 tyrosine phosphorylation ( Fig. 6 A–B)."
23333852
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"Nevertheless, our findings clearly demonstrate that optimal Usp8 tyrosine phosphorylation requires an intact Usp8 MIT domain."
21044682
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"Furthermore, we demonstrate that S680 is not involved in the regulation of Usp8 tyrosine phosphorylation."
23333852
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"In agreement with our previous observation, Usp8 tyrosine phosphorylation following stimulation of EGFR was completely blocked in the presence of PD153035 ( Fig. 4 A, top panel, and 4 C)."
21044682
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"With the additional MIT-deletion mutants Δ90 and Δ74, we demonstrate that the first 74 amino acids of the MIT-domain of Usp8 are not essential for proper MIT-domain function that results in Usp8 tyros[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"We also show that enhanced endosomal recycling of the EGFR induced by TGFα stimulation is associated with decreased Usp8 tyrosine phosphorylation."
23333852
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"Consistent with our previously published results [17] , ligand-induced Usp8 tyrosine phosphorylation was enhanced in Usp8 C748A compared to Usp8 WT ( Fig. 2 A–B )."
23333852
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"We further show that Usp8 tyrosine phosphorylation upon stimulation of EGFR-ErbB2 is (a) independent of Y1091, (b) dependent on Src- and EGFR-ErbB2-kinase activity, (c) enhanced upon coexpression of Usp8-C748A, and (d) partly dependent on the Microtubule Interacting and Transport (MIT) domain of Usp8."
21044682
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"Deletion of the N-terminal 90 amino acids (Usp8 Δ90) resulted in an almost comparable decrease in Usp8 tyrosine phosphorylation ( Fig. 6 )."
23333852
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"Our finding that the MIT domain of Usp8 is essential for optimal Usp8 tyrosine phosphorylation ( Fig. 6 ) is consistent with the model that Usp8 is tyrosine phosphorylated upon MIT-dependent recruitme[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
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"Mechanistically we have observed that this is due to neuregulin-induced and PI3-K/Akt-dependent phosphorylation of USP8, leading to increased stability of USP8, which in turn promotes the stabilization of Nrdp1, leading to ErbB3 degradation ( xref )."
18706412
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"In breast tumor cells, Akt activation led to USP8 (thr907) phosphorylation, which in turn led to increased DUB activity toward the E3 ligase Nrdp1 ( xref )."
20484045
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"We next examined whether the 14-3-3 binding to UBPY requires the phosphorylation of UBPY."
17720156
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"The cells also demonstrated the increased levels of USP8 phosphorylation and trichoplein expression, in accordance with the EGFR autophosphorylation."
29472535
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"It is plausible that after the 14-3-3 protein allosterically binds to phosphorylated USP8; USP8 undergoes a conformational change from an active state to an inhibitory state ( xref )."
34177595
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"EGF stimulation of EGFR-ErbB4 CYT-1 results in coprecipitation of USP8 with activated receptor and USP8 tyrosine phosphorylation."
23153581
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"We therefore hypothesize that tyrosine phosphorylation of Usp8 occurs in an ErbB receptor-Src-Usp8 trimolecular complex."
21044682
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"Although the intensity of the signals was clearly lower than observed for EGFR (results not shown), our results show that ligand-induced USP8 coprecipitation and USP8 tyrosine phosphorylation are incr[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23153581
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"These results show that EGF stimulation of EGFR-ERBB4 CYT-1 triggers USP8 tyrosine phosphorylation and demonstrate that USP8 is part of the ERBB4 signaling cascade."
23153581
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"However, the site of tyrosine phosphorylation and the functional significance of Usp8 tyrosine phosphorylation are as yet unknown."
23333852
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"To test this hypothesis, we first analyzed USP8 tyrosine phosphorylation upon EGF stimulation of EGFR-ErbB4 CYT-1."
23153581
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"The level of phosphorylated UBPY detected with this antibody was significantly lower in nocodazole-treated cells than in untreated cells ( Fig. 7 C)."
17720156
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"Although we cannot exclude the possibility that reduced Usp8 tyrosine phosphorylation upon removal of the MIT domain is due to conformational effects, we believe this explanation is less likely as thi[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"We therefore hypothesize that tyrosine phosphorylation of Usp8 could regulate the function of Usp8."
23333852
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"However, the tyrosine phosphorylation site of Usp8 and the precise role of Usp8 tyrosine phosphorylation remain to be elucidated."
23333852
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"To determine whether the MIT domain is also necessary for the enhanced tyrosine phosphorylation of Usp8 C748A, we tested the effect of ΔMIT Usp8 C748A on tyrosine phosphorylation."
23333852
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"Nevertheless, our findings demonstrate that optimal Usp8 tyrosine phosphorylation requires an intact Usp8 MIT domain, which is consistent with the proposed role of the MIT domain in recruitment of Usp[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"Moreover, EGF stimulation of EGFR induces USP8 tyrosine phosphorylation while, in contrast, TGF-α reduces tyrosine phosphorylation ."
28923280
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"EGFR activation causes USP8 to become phosphorylated and to associate with EGFR on endosomes where it dynamically regulates EGFR ubiquitination state during trafficking."
25721765
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"Nevertheless, these results strengthen our conclusion that an intact MIT domain is essential for optimal Usp8 tyrosine phosphorylation."
23333852
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"As shown in Fig. 2 , Usp8 becomes tyrosine phosphorylated upon treatment of EGFR-ErbB2 with EGF, although to a lesser extent than in EGFR stimulated cells."
21044682
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"Additionally, USP8 phosphorylation is also important for the regulation of ciliogenesis in the dividing cells."
32512887
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"The Y1103F mutation of the Cbl binding site did not affect USP8 tyrosine phosphorylation of the EGFR-ERBB4 CYT-1 receptor ( Fig. 6 A), as confirmed by quantitative analysis ( Fig. 6 B)."
23153581
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"Taken together, these results show that Y189, Y255, Y308, Y336, Y641 and Y679 are not solely responsible for the total Usp8 tyrosine phosphorylation detected upon EGF stimulation of EGFR, and therefor[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"USP8 phosphorylation leads to its activation and stabilization, which in turn stabilizes Nrdp1."
20868762
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"Co-expression of the ΔMIT Usp8 C748A mutant partly recovered the decrease in Usp8 tyrosine phosphorylation seen upon deletion of the MIT-domain and was comparable to Usp8 WT tyrosine phosphorylation, [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"As we have demonstrated before, deletion of the MIT-domain in Usp8 results in decreased EGF-induced Usp8 tyrosine phosphorylation [17] ."
23333852
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"In the context of a chimeric EGFR-ErbB2 receptor, we have shown that (i) EGF-induced K63-linked polyubiquitination of the ErbB2 cytoplasmic tail occurs efficiently in a pY1091-dependent manner, (ii) c[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"These results strengthen our previous findings that (i) USP8 is a target for Src-family tyrosine kinases [17,21] , (ii) ligand-induced recycling of ErbB receptors is associated with decreased interac[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"It can be hypothesized that Usp8 tyrosine phosphorylation regulates Usp8 activity by affecting the function of the MIT domain, the Rhod domain, the interaction with 14–3–3 or its enzymatic activity."
23333852
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"Our results demonstrate that the MIT-domain is necessary for ligand-induced tyrosine phosphorylation of Usp8 1-504."
23333852
sparser
"These results show that the MIT domain is essential for optimal Usp8 tyrosine phosphorylation."
23333852
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"However, the functional significance of Usp8 tyrosine phosphorylation is as yet unknown."
21044682
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"Removal of the MIT-domain abolished the tyrosine phosphorylation of Usp8 1–504."
23333852
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"However, site-directed mutagenesis of Y255 did not result in a detectable decrease in total Usp8 tyrosine phosphorylation ( Fig. 7 )."
23333852
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"We speculate that only a relatively small fraction of cellular Usp8 is recruited from the cytoplasm towards endosomes, resulting in a low stoichiometry of tyrosine phosphorylated Usp8."
23333852
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"Interestingly, removal of the MIT domain does not completely abolish Usp8 tyrosine phosphorylation, suggesting that Usp8 might be recruited to endosomal membranes also via alternative mechanisms (e.g.[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"Using Usp8 deletion constructs, our results demonstrate that Usp8 is efficiently tyrosine phosphorylated on at least one tyrosine residue in the N-terminal 504 amino acids of Usp8, as detected by immu[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"These results support our previous findings that ligand-induced recycling of ERBB receptors is associated with decreased interaction with and tyrosine phosphorylation of Usp8, and suggest that only th[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
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"Tyrosine phosphorylation of Usp8 was confirmed on a parallel Western blot ( Fig. 8 B)."
23333852
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"Stimulation of EGFR K721A and EGFR-ErbB2 K733A resulted in complete inhibition of Usp8 tyrosine phosphorylation and abrogated the coprecipitation of the autophosphorylated EGFR and EGFR-ErbB2 with Usp[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
21044682
sparser
"Alternatively, the remaining tyrosine phosphorylation of the Usp8-Δ140 mutant may also indicate that Usp8 is partly phosphorylated by activated Src-family kinases in the cytoplasm."
21044682
sparser
"MS analysis did not reveal any evidence for Usp8 tyrosine phosphorylation, although it clearly revealed the presence of several phosphorylated serine residues including pS153, pS392, pS680 and pS722 i[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"To further identify tyrosine residues that are involved in Usp8 tyrosine phosphorylation, we investigated several tyrosine residues within the Rhod (residues 185–310) and linker domain (residues 310–7[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
sparser
"The finding that site-directed mutagenesis did not reveal any evidence in favor of tyrosine phosphorylation may be due to the fact that we did not mutate all tyrosines in this region or to the possibi[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23333852
rlimsp
"We conclude that UBPY is catalytically inhibited in a phosphorylation-dependent manner by 14-3-3s during the interphase, and this regulation is cancelled in the M phase."
17720156