Results 1 - 10 of 2544
Results 1 - 10 of 2544. Search took: 0.028 seconds
|Sort by: date | relevance|
[en] Highlights: • Tyrosine kinase Fer preferentially binds to highly curved membranes via its FX domain. • An algorithmic search identified an intrinsically disordered region at the carboxy-terminal half of the FX domain. • The intrinsically disordered sequence forms an amphipathic helix upon binding to curved membranes. • Tyrosine kinase activity of Fer is enhanced by the presence of highly curved membranes. • -- Abstract: . Tyrosine kinases are important enzymes that mediate signal transduction at the plasma membrane. While the significance of membrane localization of tyrosine kinases has been well evaluated, the role of membrane curvature in their regulation is unknown. Here, we demonstrate that an intrinsically disordered region in the tyrosine kinase Fer acts as a membrane curvature sensor that preferentially binds to highly curved membranes in vitro. This region forms an amphipathic α-helix upon interaction with curved membranes, aligning hydrophobic residues on one side of the helical structure. Further, the tyrosine kinase activity of Fer is significantly enhanced by the membrane in a manner dependent on curvature. We propose a model for the regulation of Fer based on an intramolecular interaction and the curvature-dependent membrane binding mediated by its intrinsically disordered region.
[en] Discoidin Domain Receptor 2 (DDR2) is a collagen-binding receptor tyrosine kinase that initiates delayed and sustained tyrosine phosphorylation signalling. To understand the molecular basis of this unique phosphorylation profile, here we utilise fluorescence microscopy to map the spatiotemporal localisation of DDR2 and tyrosine phosphorylated proteins upon stimulation with collagen. We show that cellular phosphorylated proteins are localised to the interface where DDR2 is in contact with collagen and not in the early endosomes or lysosomes. We find that DDR2 localisation is independent of integrin activation and the key DDR2 signalling effector SHC1. Structure-function analysis reveals that DDR2 mutants defective for collagen binding or kinase activity are unable to localise to the cell surface, demonstrating for the first time that both collagen binding and kinase functions are required for spatial localisation of DDR2. This study provides new insights into the underlying structural features that control DDR2 activation in space and time.
[en] Highlights: • C. pneumoniae infection promotes monocyte TEM through increasing VEC permeability. • C. pneumoniae infection stimulates VE-cadherin tyrosine phosphorylation by Src. • VE-cadherin phosphorylation is involved in VE-cadherin internalization. • VE-cadherin phosphorylation is involved in EC hyperpermeability caused by infection. • VE-cadherin phosphorylation is associated with the infection-induced monocyte TEM. Migration of monocytes into the subendothelial layer of the intima is one of the critical events in early atherosclerosis. Chlamydia pneumoniae (C. pneumoniae) infection has been shown to promote monocyte transendothelial migration (TEM). However, the exact mechanisms have not yet been fully clarified. In this study, we tested the hypothesis that C. pneumoniae infection increases vascular endothelial cell (VEC) permeability and subsequent monocyte TEM through stimulating the tyrosine phosphorylation of vascular endothelial-cadherin (VE-cadherin). Here, we demonstrated that C. pneumoniae infection promoted monocyte TEM in a TEM assay possibly by increasing the permeability of a VEC line EA.hy926 cell as assessed by measuring the passage of FITC-BSA across a VEC monolayer. Subsequently, Western blot analysis showed that C. pneumoniae infection induced VE-cadherin internalization. Our further data revealed that Src-mediated VE-cadherin phosphorylation at Tyr658 was involved in C. pneumoniae infection-induced internalization of VE-cadherin, VEC hyperpermeability and monocyte TEM. Taken together, our data indicate that C. pneumoniae infection promotes monocyte TEM by increasing VEC permeability via the tyrosine phosphorylation and internalization of VE-cadherin in VECs.
[en] Highlights: • MAPK phosphatase DUSP18 binds to ataxin-1 and suppresses JNK activation. • DUSP18 inhibits the SUMOylation of ataxin-1 without affecting phosphorylation. • DUSP18 blocks ataxin-1 aggregation and disrupt its association with Sp100. We previously reported that SUMOylation promotes the aggregation of ataxin-1 and JNK is involved in the process. Here we show that dual-specificity phosphatase 18 (DUSP18), a member of protein tyrosine phosphatases, exerts the opposite effects on ataxin-1. DUSP18 associated with ataxin-1 and suppressed JNK activated by ataxin-1. Interestingly DUSP18, but not the other DUSPs interacting with ataxin-1, caused the mobility shift of ataxin-1. De-phosphorylation by DUSP18 was initially suspected as a cause for such an effect; however, the phosphorylation of ataxin-1 was unchanged. Instead DUSP18 inhibited SUMOylation and reduced ataxin-1 aggregation. The catalytic mutant of DUSP18 failed to reduce the SUMOylation and aggregation of ataxin-1 indicating that the phosphatase activity is indispensable for the effects. Moreover, DUSP18 disrupted the co-localization of ataxin-1 with the PML component Sp100. These results together implicate that JNK and DUSP18 reciprocally modulate the SUMOylation, which plays a regulatory role in the aggregation of ataxin-1.
[en] Radioiodine labeled tyrosine analogues, such as L-3[I-123]iodo-α-methyltyrosine, have been used for the evaluation of cerebral amino acid transport. Furthermore, these could be used for the tumor grading. We added one methyl-group to the L-3-α-methyltyrosine, expecting the increased cellular membrane permeability. The aim of this study was to evaluate the feasibility of OMIMT as an agent for tumor image. After synthesis of o-methyl-L-α-methyltyrosine (OMAMT), OMAMT was labeled with I-131 using Iodogen method. Synthesis of [I-131] IMT was performed as previously described (C. Krummeich et al. J. Appl. Rad. Isot, 45: 929, 1994). Uptake and retention studies were performed using 9L gliosarcoma cells (106) at various time points upto 4 hr. OMIMT uptake was 2.5 times higher than IMT uptake at 60 min and same after 2hr. Female Fischer rats were implanted with the 9L gliosarcoma cell line into right thigh. The biodistribution (five rats per group) was evaluated (30 min, 2hr, 24hr) after iv injection of 3.7 MBq I-131 labeled OMIMT and IMT. Maximum accumulation in tumors occurred at 30 min for both OMIMT and IMT. The tumor uptake of OMIMT was significantly higher than that of IMT at early time point studied (3.74 vs 0.38 %ID/g at 30 min and 2.40 vs 0.24 %ID/g at 2 hr, respectively). However, The tumor uptakes of both radiolabels were similar at 24 hr (0.04 vs 0.05 %ID/g, respectively). The kidney was the major route of elimination and had the greater accumulation (23.98 vs 4.03 %ID/g at 30 min). Gamma camera images were obtained at 30 min , 2 hr and 24 hr. Tumor was visualized as early as at 30 min. These data suggested that [I-131]OMIMT might be useful as a tumor imaging agent and had advantage for the tumor image
[en] Metastatic disease is present at diagnosis in 30% of the patients with colorectal cancer (CRC), and approximately half of early-stage patients with CRC will eventually present with metastatic disease. Until recently, few chemotherapy options were available to treat metastatic colorectal carcinoma (MCRC). Fluorouracil (5-FU) with leucovorin (LV) modulation has a marginal but positive effect on survival in those patients. The recent incorporation of irinotecan (CPT -11) and oxaliplatin for the management of advanced CRC has generated further improvement in survival. The development of oral fluoropyrimidines, mimicking continuous infusion 5-FU, is convenient to use. Recently completed or ongoing clinical trials to study novel targeting agents have initiated a new era of drug development. Anti-angiogenesis drugs, tyrosine kinase inhibitors, and epidermal growth factor blockers are among the new generation of agents. (author)
[en] The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (ε- and ζ-SAIL Phe) and tyrosine (ε-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven that these SAIL amino acids provide dramatic spectral simplification and sensitivity enhancement for the aromatic ring NMR signals. In addition to these SAIL aromatic amino acids, we recently synthesized δ-SAIL Phe and δ-SAIL Tyr, which allow us to observe and assign δ-13C/1H signals very efficiently. Each of the various types of SAIL Phe and SAIL Tyr yields well-resolved resonances for the δ-, ε- or ζ-13C/1H signals, respectively, which can readily be assigned by simple and robust pulse sequences. Since the δ-, ε-, and ζ-proton signals of Phe/Tyr residues give rise to complementary NOE constraints, the concomitant use of various types of SAIL-Phe and SAIL-Tyr would generate more accurate protein structures, as compared to those obtained by using conventional uniformly 13C, 15N-double labeled proteins. We illustrated this with the case of an 18.2 kDa protein, Escherichia coli peptidyl-prolyl cis-trans isomerase b (EPPIb), and concluded that the combined use of ζ-SAIL Phe and ε-SAIL Tyr would be practically the best choice for protein structural determinations.
[en] The metastatic potential of malignant tumor has been shown to be correlated with the increased expression of tri- and tetra-antennary β1,6-N-acetylglucosamine (β1,6-GlcNAc) N-glycans. In this study, We found that GnT-V expression was negatively correlated with receptor protein tyrosine phosphatase type μ(RPTPμ) in human glioma tissues. To study whether RPTPμ is a novel substance of GnT-V which further affect RPTPμ′s downstream dephosphorylation function, we preform lentiviral infection with GnT-V gene to construct stably transfected GnT-V glial cell lines. We found RPTPμ undergone severer cleavage in GnT-V transfected glioma cells compare to Mock cells. RPTPμ intracellular domain fragments increased while β1,6-GlcNAc-branched N-glycans increased, in consistent with the decrease of RPTPμ′s catalytic activity. The results showed that abnormal glycosylation could decrease the phosphorylation activity of PTP μ, and affect PLCγ-PKC pathways. Both protease inhibitor Furin and N-glycan biosynthesis inhibitor swainsonine could decrease cell mobility in GnT-V-U87 transfectants and other glioma cell lines. All results above suggest increased post-translational modification of RPTPμ N-glycans by GnT-V attenuates its tyrosine phosphatase activity and promotes glioma cell migration through PLCγ-PKC pathways, and that the β1,6-GlcNAc-branched N-glycans of RPTPμ play a crucial role in glioma invasivity.
[en] Although anlotinib, a multi-targeted receptor tyrosine kinase inhibitor has been reported have antitumor effects in many preclinical and clinical trials, little is known about its effect on hepatocellular carcinoma (HCC). Here, we have shown the antitumor effects of anlotinib on HCC. Data indicated that anlotinib application significantly inhibited HCC cell viability, proliferation, colony formation, and prompted apoptosis in vitro. Furthermore, animal experiments also illustrated that anlotinib alleviated HCC progression. Mechanically, we demonstrated that anlotinib treatment downregulated the anti-apoptotic protein Bcl-2 and Survivin, but upregulated pro-apoptotic molecule Bax, which accounts for its therapeutic effect on HCC. Pathway analysis has shown decreased phosphorylation levels of Erk and Akt. Together, this study suggests that anlotinib may have a direct antitumor progression effect on HCC by inhibiting Bcl-2 and Survivin expression, promoting Bax expression via inactivating Erk and Akt pathways and could be a promising agent treating HCC.