共查询到20条相似文献,搜索用时 15 毫秒
1.
Tung-Ju Hsieh Tien-Jui Yen Te-Sheng Lin Hsun-Tang Chang Shu-Yun Huang Chun-Hua Hsu Lynn Farh Nei-Li Chan 《Nucleic acids research》2010,38(12):4173-4181
DNA gyrase is the only topoisomerase capable of introducing (−) supercoils into relaxed DNA. The C-terminal domain of the gyrase A subunit (GyrA-CTD) and the presence of a gyrase-specific ‘GyrA-box’ motif within this domain are essential for this unique (−) supercoiling activity by allowing gyrase to wrap DNA around itself. Here we report the crystal structure of Xanthomonas campestris GyrA-CTD and provide the first view of a canonical GyrA-box motif. This structure resembles the GyrA-box-disordered Escherichia coli GyrA-CTD, both adopting a non-planar β-pinwheel fold composed of six seemingly spirally arranged β-sheet blades. Interestingly, structural analysis revealed that the non-planar architecture mainly stems from the tilted packing seen between blades 1 and 2, with the packing geometry likely being defined by a conserved and unusual β-strand-bearing proline. Consequently, the GyrA-box-containing blade 1 is placed at an angled spatial position relative to the other DNA-binding blades, and an abrupt bend is introduced into the otherwise flat DNA-binding surface. Mutagenesis studies support that the proline-induced structural twist contributes directly to gyrase’s (−) supercoiling activity. To our knowledge, this is the first demonstration that a β-strand-bearing proline may impact protein function. Potential relevance of β-strand-bearing proline to disease phenylketonuria is also noted. 相似文献
2.
Sparks S Liu G Robbins KJ Lazo ND 《Biochemical and biophysical research communications》2012,422(4):551-555
Understanding how small molecules affect amyloid formation is of major biomedical and pharmaceutical importance due to the association of amyloid with incurable diseases including Alzheimer's, Parkinson's, and type II diabetes. Using solution state (1)H NMR, we demonstrate that curcumin, a planar biphenolic compound found in the Indian spice turmeric, delays the self-assembly of islet amyloid polypeptide to NMR-invisible assemblies. Accompanying circular dichroism studies show that curcumin disassembles α-helix in maturing assemblies of IAPP. The amount of α-helix disassembled correlates with predicted and experimentally determined helical content of IAPP obtained by others. Taken together, these results indicate that curcumin modulates IAPP self-assembly by unfolding α-helix on pathway to amyloid. The implications of this work in the elucidation of the mechanism for amyloid formation by IAPP in the presence of curcumin are discussed. 相似文献
3.
Arye Elfenbein John M. Rhodes Julia Meller Martin A. Schwartz Michiyuki Matsuda Michael Simons 《The Journal of cell biology》2009,186(1):75-83
Fibroblast growth factor 2 (FGF2) is a major regulator of developmental, pathological, and therapeutic angiogenesis. Its activity is partially mediated by binding to syndecan 4 (S4), a proteoglycan receptor. Angiogenesis requires polarized activation of the small guanosine triphosphatase Rac1, which involves localized dissociation from RhoGDI1 and association with the plasma membrane. Previous work has shown that genetic deletion of S4 or its adapter, synectin, leads to depolarized Rac activation, decreased endothelial migration, and other physiological defects. In this study, we show that Rac1 activation downstream of S4 is mediated by the RhoG activation pathway. RhoG is maintained in an inactive state by RhoGDI1, which is found in a ternary complex with synectin and S4. Binding of S4 to synectin increases the latter''s binding to RhoGDI1, which in turn enhances RhoGDI1''s affinity for RhoG. S4 clustering activates PKCα, which phosphorylates RhoGDI1 at Ser96. This phosphorylation triggers release of RhoG, leading to polarized activation of Rac1. Thus, FGF2-induced Rac1 activation depends on the suppression of RhoG by a previously uncharacterized ternary S4–synectin–RhoGDI1 protein complex and activation via PKCα. 相似文献
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Lee PY Kumagai Y Xu Y Li Y Barker T Liu C Sobel ES Takeuchi O Akira S Satoh M Reeves WH 《Journal of immunology (Baltimore, Md. : 1950)》2011,186(3):1747-1754
Exposure to naturally occurring hydrocarbon oils is associated with the development of chronic inflammation and a wide spectrum of pathological findings in humans and animal models. The mechanism underlying the unremitting inflammatory response to hydrocarbons remains largely unclear. The medium-length alkane 2,6,10,14 tetramethylpentadecane (also known as pristane) is a hydrocarbon that potently elicits chronic peritonitis characterized by persistent infiltration of neutrophils and monocytes. In this study, we reveal the essential role of IL-1α in sustaining the chronic recruitment of neutrophils following 2,6,10,14 tetramethylpentadecane treatment. IL-1α and IL-1R signaling promote the migration of neutrophils to the peritoneal cavity in a CXCR2-dependent manner. This mechanism is at least partially dependent on the production of the neutrophil chemoattractant CXCL5. Moreover, although chronic infiltration of inflammatory monocytes is dependent on a different pathway requiring TLR-7, type I IFN receptor, and CCR2, the adaptor molecules MyD88, IL-1R-associated kinase (IRAK)-4, IRAK-1, and IRAK-2 are shared in regulating the recruitment of both monocytes and neutrophils. Taken together, our findings uncover an IL-1α-dependent mechanism of neutrophil recruitment in hydrocarbon-induced peritonitis and illustrate the interactions of innate immune pathways in chronic inflammation. 相似文献
6.
Juan Pablo Petiti Silvina Gutiérrez Jorge Humberto Mukdsi Ana Lucía De Paul Alicia Inés Torres 《Journal of molecular histology》2009,40(5-6):417-425
The variations of the intracellular localization of the individual protein kinase C (PKC) isoforms are related with their different biological functions. In this study, we have investigated the precise intracellular translocation of endogenous PKCα and PKCε in PMA-stimulated normal and tumoral lactotroph cells by using confocal and immunogold electron microscopy, which was correlated with the rate of cell proliferation of both pituitary cell phenotypes. The present results showed that the short phorbol ester incubation stimulated the proliferation of normal and tumoral lactotroph cells, as determined by the measurement of the BrdU-labelling index. The translocation of PKCα to plasma and nuclear membranes induced by PMA was more marked than that observed for PKCε in normal and tumoral lactotroph cells. Our results showed that PKCs translocation to the plasma and nuclear membranes varied from isozyme to isozyme emphasizing that PKCα could be related with the mitogenic stimulus exerted by phorbol ester. These data support the notion that specific PKC isozymes may exert spatially defined effects by virtue of their directed translocation to distinct intracellular sites. 相似文献
7.
Wang WL Yeh SF Huang EY Lu YL Wang CF Huang CY Lin WJ 《Apoptosis : an international journal on programmed cell death》2007,12(10):1857-1871
Various pathways, including regulation of functions of the Bcl-2 family, are implicated in the survival promotion by PKCα, however the molecular mechanisms are still obscure. We have previously demonstrated that PKCα is selectively anchored to mitochondria by PICK1 in fibroblasts NIH 3T3. In this study, we show that over-expression of PICK1 in leukemia REH confers resistance to etoposide-induced apoptosis, which requires an interaction with PKCα as the non-interacting mutant PICK1 loses the pro-survival activity. The PKCα selective inhibitor Gö6976 also abolishes the anti-apoptotic effect indicating a requirement for PKC activity. Disruption of PICK1/PKCα interactions by inhibitory peptides significantly increases cellular susceptibility to etoposide. Similar effects are also observed in HL60 cells, which exhibit an intrinsic resistance to etoposide. Molecular analysis shows that the wild type PICK1, but not the non-interacting mutant, prevents the loss of mitochondrial membrane potential with a coincident increase in phosphorylation of the anti-apoptotic Bcl-2(Ser70) and a decrease in dimerization of the pro-apoptotic Bax. PICK1 may provide the spatial proximity for phosphorylation of Bcl-2(Ser70) by PKCα which then leads to a higher survival. Taken together, our results suggest that PICK1 may mediate the pro-survival activity of PKCα by serving as a molecular link between PKCα and mitochondria. 相似文献
8.
The 22 γ-protocadherins (γ-Pcdhs) potentially specify thousands of distinct homophilic adhesive interactions in the brain. Neonatal lethality of mice lacking the Pcdh-γ gene cluster has, however, precluded analysis of many brain regions. Here, we use a conditional Pcdh-γ allele to restrict mutation to the cerebral cortex and find that, in contrast to other central nervous system phenotypes, loss of γ-Pcdhs in cortical neurons does not affect their survival or result in reduced synaptic density. Instead, mutant cortical neurons exhibit severely reduced dendritic arborization. Mutant cortices have aberrantly high levels of protein kinase C (PKC) activity and of phosphorylated (inactive) myristoylated alanine-rich C-kinase substrate, a PKC target that promotes arborization. Dendrite complexity can be rescued in Pcdh-γ mutant neurons by inhibiting PKC, its upstream activator phospholipase C, or the γ-Pcdh binding partner focal adhesion kinase. Our results reveal a distinct role for the γ-Pcdhs in cortical development and identify?a signaling pathway through which they play this role. 相似文献
9.
Alessandro Scopelliti Christin Bauer Marcos Vidal 《Cell cycle (Georgetown, Tex.)》2016,15(12):1538-1544
Bursicon is the main regulator of post molting and post eclosion processes during arthropod development. The active Bursicon hormone is a heterodimer of Burs-α and Burs-β. However, adult midguts express Burs-α to regulate the intestinal stem cell niche. Here, we examined the potential expression and function of its heterodimeric partner, Burs-β in the adult midgut. Unexpectedly, our evidence suggests that Burs-β is not significantly expressed in the adult midgut. burs-β mutants displayed the characteristic developmental defects but showed wild type-like adult midguts, thus uncoupling the developmental and adult phenotypes seen in burs-α mutants. Gain of function data and ex vivo experiments using a cAMP biosensor, demonstrated that Burs-α is sufficient to drive stem cell quiescence and to activate dLGR2 in the adult midgut.
Our evidence suggests that the post developmental transactivation of dLGR2 in the adult midgut is mediated by Burs-α and that the β subunit of Bursicon is dispensable for these activities. 相似文献
10.
A neuron forms thousands of presynaptic nerve terminals on its axons, far removed from the cell body. The protein CSPα resides in presynaptic terminals, where it forms a chaperone complex with Hsc70 and SGT. Deletion of CSPα results in massive neurodegeneration that impairs survival in mice and flies. In CSPα-knockout mice, levels of presynaptic SNARE complexes and the SNARE protein SNAP-25 are reduced, suggesting that CSPα may chaperone SNARE proteins, which catalyse synaptic vesicle fusion. Here, we show that the CSPα-Hsc70-SGT complex binds directly to monomeric SNAP-25 to prevent its aggregation, enabling SNARE-complex formation. Deletion of CSPα produces an abnormal SNAP-25 conformer that inhibits SNARE-complex formation, and is subject to ubiquitylation and proteasomal degradation. Even in wild-type mouse terminals, SNAP-25 degradation is regulated by synaptic activity; this degradation is decreased by CSPα overexpression, and enhanced by CSPα deletion. Thus, SNAP-25 function is maintained during rapid SNARE cycles by equilibrium between CSPα-dependent chaperoning and ubiquitin-dependent degradation, revealing unique protein quality-control machinery within the presynaptic compartment. 相似文献
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Marika Sj?qvist Daniel Antfolk Saima Ferraris Vilma Rraklli Cecilia Haga Christian Antila Anders Mutvei Susumu Y Imanishi Johan Holmberg Shaobo Jin John E Eriksson Urban Lendahl Cecilia Sahlgren 《Cell research》2014,24(4):433-450
Activation of Notch signaling requires intracellular routing of the receptor, but the mechanisms controlling the distinct steps in the routing process is poorly understood. We identify PKCζ as a key regulator of Notch receptor intracellular routing. When PKCζ was inhibited in the developing chick central nervous system and in cultured myoblasts, Notch-stimulated cells were allowed to undergo differentiation. PKCζ phosphorylates membrane-tethered forms of Notch and regulates two distinct routing steps, depending on the Notch activation state. When Notch is activated, PKCζ promotes re-localization of Notch from late endosomes to the nucleus and enhances production of the Notch intracellular domain, which leads to increased Notch activity. In the non-activated state, PKCζ instead facilitates Notch receptor internalization, accompanied with increased ubiquitylation and interaction with the endosomal sorting protein Hrs. Collectively, these data identify PKCζ as a key regulator of Notch trafficking and demonstrate that distinct steps in intracellular routing are differentially modulated depending on Notch signaling status. 相似文献
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Pérez-Lara A Egea-Jiménez AL Ausili A Corbalán-García S Gómez-Fernández JC 《Biochimica et biophysica acta》2012,1821(11):1434-1442
Protein kinase Cα (PKCα) is activated by its translocation to the membrane. Activity assays show the importance of PIP(2) in determining the specific activity of this enzyme. A FRET stopped flow fluorescence study was carried out to monitor the rapid kinetics of protein binding to model membranes containing POPC/POPS/DOG and eventually PIP(2). The results best fitted a binding mechanism in which protein bound to the membrane following a two-phase mechanism with a first bimolecular reaction followed by a slow unimolecular reaction. In the absence of PIP(2), the rapid protein binding rate was especially dependent on POPS concentration. Formation of the slow high affinity complex during the second phase seems to involve specific interactions with POPS and DOG since it is only sensitive to changes within relatively low concentration ranges of these lipids. Both the association and dissociation rate constants fell in the presence of PIP(2). We propose a model in which PKCα binds to the membranes via a two-step mechanism consisting of the rapid membrane initial recruitment of PKCα driven by interactions with POPS and/or PIP(2) although interactions with DOG are involved too. PKCα searches on the lipid bilayer in two dimensions to establish interactions with its specific ligands. 相似文献
14.
Amit S. AdhikariBhairab N. Singh K. Sridhar RaoCh Mohan Rao 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2011,1813(8):1532-1542
αB-crystallin, a member of the small heat shock protein family, has been implicated in various biological functions including response to heat shock, differentiation and apoptosis, the mechanisms of which have not been well understood. Myoblasts, the precursor cells in muscle regeneration, when subjected to growth factor deprivation differentiate to form myotubes or undergo apoptosis. During differentiation, myoblasts express elevated levels of αB-crystallin as well as TNF-α but the connecting link between these proteins in cell signaling is not clearly understood. We have therefore investigated the role of αB-crystallin in TNF-α induced regulation of NF-κB.We demonstrate that in response to TNF-α treatment, αB-crystallin associates with IKKβ and activate its kinase activity, facilitating the degradation of phosphorylated I-kBα, a prime step in NF-κB activation. Reducing the level of αB-crystallin using the RNAi approach reduces the translocation of p65, further confirming the role of αB-crystallin in NF-κB activation. Our study shows that the ability of αB-crystallin to activate NF-κB depends on its phosphorylation status. The present study shows that αB-crystallin-dependent NF-κB activation protects myoblasts from TNF-α induced cytoxicity by enhancing the expression of the anti-apoptotic protein, Bcl 2. Thus, our study identifies yet another mechanism by which αB-crystallin exerts its anti-apoptotic activity. 相似文献
15.
This study investigated if the nephroprotective effect of Curcumin in streptozotocin-induced type 1 diabetes mellitus (DM) in rats involves downregulation/inhibition of p66Shc and examined the underlying mechanisms. Rats were divided into 4 groups (n = 12/group) as control, control + Curcumin (100 mg/kg), T1DM, and T1DM + Curcumin. Curcumin was administered orally to control or diabetic rats for 12 weeks daily. As compared to diabetic rats, Curcumin didn't affect either plasma glucose or insulin levels but significantly reduced serum levels of urea, blood urea nitrogen, and creatinine, and concurrently reduced albumin/protein urea and increased creatinine clearance. It also prevented the damage in renal tubules and mitochondria, mesangial cell expansion, the thickness of the basement membrane. Mechanistically, Curcumin reduced mRNA and protein levels of collagen I/III and transforming growth factor- β-1 (TGF-β1), reduced inflammatory cytokines levels, improved markers of mitochondrial function, and suppressed the release of cytochrome-c and the activation of caspase-3. In the kidneys of both control and diabetic rats, Curcumin reduced the levels of reactive oxygen species (ROS), increased mRNA levels of manganese superoxide dismutase (MnSOD) and gamma-glutamyl ligase, increased glutathione (GSH) and protein levels of Bcl-2 and MnSOD, and increased the nuclear levels of nuclear factor2 (Nrf2) and FOXO-3a. Besides, Curcumin reduced the nuclear activity of the nuclear factor-kappa B (NF-κB), downregulated protein kinase CβII (PKCβII), NADPH oxidase, and p66Shc, and decreased the activation of p66Shc. In conclusion, Curcumin prevents kidney damage in diabetic rats by activating Nrf2, inhibiting Nf-κB, suppressing NADPH oxidase, and downregulating/inhibiting PKCβII/p66Shc axis. 相似文献
16.
A novel function for the binder of Arl two (BART) molecule in pancreatic cancer cells is reported. BART inhibits invasiveness of pancreatic cancer cells through binding to a Ca(2+)-dependent, phosphorylated, guanosine triphosphatase (GTPase) membrane fusion protein, annexin7 (ANX7). A tumor suppressor function for ANX7 was previously reported based on its prognostic role in human cancers and the cancer-prone mouse phenotype ANX7(+/-). Further investigation demonstrated that the BART-ANX7 complex is transported toward cell protrusions in migrating cells when BART supports the binding of ANX7 to the protein kinase C (PKC) isoform PKCα. Recent evidence has suggested that phosphorylation of ANX7 by PKC significantly potentiates ANX7-induced fusion of phospholipid vesicles; however, the current data suggest that the BART-ANX7 complex reduces PKCα activity. Knocking down endogenous BART and ANX7 increases activity of PKCα, and specific inhibitors of PKCα significantly abrogate invasiveness induced by BART and ANX7 knockdown. These results imply that BART contributes to regulating PKCα activity through binding to ANX7, thereby affecting the invasiveness of pancreatic cancer cells. Thus, it is possible that BART and ANX7 can distinctly regulate the downstream signaling of PKCα that is potentially relevant to cell invasion by acting as anti-invasive molecules. 相似文献
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Nonalcoholic fatty liver disease (NAFLD) is a highly prevalent complication of obesity, yet cellular mechanisms that lead to its development are not well defined. Previously, we have documented hepatic steatosis in mice carrying a mutation in the Sec61a1 gene. Here we examined the mechanism behind NAFLD in Sec61a1 mutant mice. Livers of mutant mice exhibited upregulation of Pparg and its target genes Cd36, Cidec, and Lpl, correlating with increased uptake of fatty acid. Interestingly, these mice also displayed activation of the heat shock response (HSR), with elevated levels of heat shock protein (Hsp) 70, Hsp90, and heat shock factor 1. In cell lines, inhibition of Hsp90 function reduced Pparγ signaling and protein levels. Conversely, overexpression of Hsp90 increased Pparγ signaling and protein levels by reducing degradation. This may occur via a physical interaction as Hsp90 and Pparγ coimmunoprecipitated in vivo. Furthermore, inhibition of Hsp90 in Sec61a1 mutant hepatocytes also reduced Pparγ protein levels and signaling. Finally, overexpression of Hsp90 in liver cell lines increased neutral lipid accumulation, and this accumulation was blocked by Hsp90 inhibition. Our results show that the HSR and Hsp90 play an important role in the development of NAFLD, opening new avenues for the prevention and treatment of this highly prevalent disease. 相似文献
19.
Swant J Goodwin JS North A Ali AA Gamble-George J Chirwa S Khoshbouei H 《The Journal of biological chemistry》2011,286(51):43933-43943
Dysregulation of dopamine (DA) homeostasis is implicated in neurodegenerative diseases, drug addiction, and neuropsychiatric disorders. The neuronal plasma membrane dopamine transporter (DAT) is essential for the maintenance of DA homeostasis in the brain. α-Synuclein is a 140-amino acid protein that forms a stable complex with DAT and is linked to the pathogenesis of neurodegenerative disease. To elucidate the potential functional consequences of DAT/α-synuclein interaction, we explored α-synuclein modulation of DAT activity in midbrain dopaminergic neurons obtained from TH::RFP mice, immortalized DA neurons, and a heterologous system expressing DAT. We used dual pipette whole cell patch clamp recording to measure the DAT-mediated current before and after dialysis of recombinant α-synuclein into immortalized DA neurons. Our data suggest that intracellular α-synuclein induces a Na+ independent but Cl--sensitive inward current in DAT-expressing cells. This current is blocked by DAT blocker GBR12935 and is absent when heat-inactivated α-synuclein is dialyzed into these cells. The functional consequence of this interaction on DAT activity was further examined with real-time monitoring of transport function using a fluorescent substrate of DAT, 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP+). Overexpression of α-synuclein in DAT-positive immortalized DA neurons and CHO cells expressing DAT decreased the magnitude and rate of DAT-mediated substrate uptake without a decrease in the initial binding of the substrate at the plasma membrane. Taken together our findings are consistent with the interpretation that DAT/α-synuclein interaction at the cell surface results in a DAT-dependent, Na+-insensitive, Cl-sensitive inward current with a decrease in substrate uptake, suggesting that DAT/α-synuclein interaction can modulate dopamine transmission and thus neuronal function. 相似文献
20.
《Molecular cell》2014,53(5):791-805
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