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991.
ADP-ribosylation factor 6 and a functional PIX/p95-APP1 complex are required for Rac1B-mediated neurite outgrowth 下载免费PDF全文
The mechanisms coordinating adhesion, actin organization, and membrane traffic during growth cone migration are poorly understood. Neuritogenesis and branching from retinal neurons are regulated by the Rac1B/Rac3 GTPase. We have identified a functional connection between ADP-ribosylation factor (Arf) 6 and p95-APP1 during the regulation of Rac1B-mediated neuritogenesis. P95-APP1 is an ADP-ribosylation factor GTPase-activating protein (ArfGAP) of the GIT family expressed in the developing nervous system. We show that Arf6 has a predominant role in neurite extension compared with Arf1 and Arf5. Cotransfection experiments indicate a specific and cooperative potentiation of neurite extension by Arf6 and the carboxy-terminal portion of p95-APP1. Localization studies in neurons expressing different p95-derived constructs show a codistribution of p95-APP1 with Arf6, but not Arf1. Moreover, p95-APP1-derived proteins with a mutated or deleted ArfGAP domain prevent Rac1B-induced neuritogenesis, leading to PIX-mediated accumulation at large Rab11-positive endocytic vesicles. Our data support a role of p95-APP1 as a specific regulator of Arf6 in the control of membrane trafficking during neuritogenesis. 相似文献
992.
Tobella LM Bunster M Pooley A Becerra J Godoy F Martínez MA 《Journal of industrial microbiology & biotechnology》2005,32(9):397-401
Poly-β-hydroxyalkanoates (PHA) polymer is synthesized by different bacterial species. There has been considerable interest
in the development and production of biodegradable polymers; however, the high cost of PHA production has restricted its applications.
Kraft cellulose industry effluents containing 2,4,6-trichlorophenol (10 or 20 μg ml−1) were used by the bacteria Sphingopyxis chilensis S37 and Wautersia sp. PZK to synthesize PHA. In this condition, S. chilensis S37 was able to grow and degrade 2,4,6-trichlorophenol (ca. 60%) and 80% of these cells accumulated PHA. Wautersia PZK completely degraded 2,4,6-TCP and more than 90% of the cells accumulated PHA in 72 h. The PHA detection was performed
by flow cytometry and polyester composition was characterized by gas chromatography-mass spectroscopy (GC-MS), indicating
that these polymers are made by 3-hydroxybutyric acid and 3-hydroxyhexadecanoic acid for S37 and PZK strains, respectively.
Results demonstrated that strains’ growth and PHA production and composition are not modified in cellulose effluents with
or without 2,4,6-TCP (10–20 μg ml−1). Therefore, our results indicate that S. chilensis S37 and Wautersia sp. PZK are able to degrade a toxic compound such as a 2,4,6-TCP and simultaneously produce a valuable biopolymer using low-value
substrates. 相似文献
993.
Fenoll LG García-Molina F Gilabert MA Varón R García-Ruiz PA Tudela J García-Cánovas F Rodríguez-López JN 《Biological chemistry》2005,386(4):351-360
The catalytic cycle of heme peroxidases involves three processes: the formation of compound I, its conversion to compound II and regeneration of the native enzyme. Each of the processes consists of a reversible binding stage followed by an irreversible transformation stage. Our group has proposed a continuous, sensitive and reliable chronometric method for measuring the steady-state rate of peroxidase activity. Furthermore, we have derived an analytical expression for the steady-state rate and simplified it, taking into consideration the experimental values of the rate constants of some stages previously determined by other authors in stopped-flow assays. We determined the value of the constant for the transformation of a series of phenols and anilines by compound II, and found that it involves a deprotonation step and an electron transfer step. Study of the solvent deuterium isotope effect on the oxidation of phenol revealed the non-rate-limiting character of the deprotonation step in a proton inventory study. Usage of the Marcus equation showed that the electronic transfer step is rate-limiting in both cases, while phenols and anilines were oxidised at different rates for the same potentials. This can be attributed to the shorter electron-tunnelling distance for electron transfer to the iron ion in the phenols than in the anilines. 相似文献
994.
Chena MA Elizondo S Rodríguez-Páez L Nogueda B Baeza I Wong C 《Journal of enzyme inhibition and medicinal chemistry》2005,20(2):189-197
The trypanocidal activity of N-isopropyl oxamate (NIPOx) and the ethyl ester of N-isopropyl oxamate (Et-NIPOx) were tested on cultured epimastigotes (in vitro) and on murine trypanosomiasis (in vivo) using five different T. cruzi strains. When benznidazole and nifurtimox, used for comparison, were tested we found that only three of these T. cruzi strains were affected, whereas the other two strains, Miguz and Compostela, were resistant to the in vitro and the in vivo trypanocidal activity of these substances. In addition, when NIPOx was tested on cultured epimastigotes and on mice parasitaemia, trypanocidal activity was not obtained on either of these T. cruzi strains. Our experiments strongly suggest that NIPOx does not penetrate intact epimastigotes due to the polarity of its carboxylate whereas Et-NIPOx, acting as a prodrug, exhibited in vitro and in vivo trypanocidal activity in the five tested T. cruzi strains. 相似文献
995.
A 320-kilobase artificial chromosome encoding the human HLA DR3-DQ2 MHC haplotype confers HLA restriction in transgenic mice 总被引:2,自引:0,他引:2
Chen Z Dudek N Wijburg O Strugnell R Brown L Deliyannis G Jackson D Koentgen F Gordon T McCluskey J 《Journal of immunology (Baltimore, Md. : 1950)》2002,168(6):3050-3056
MHC class II haplotypes control the specificity of Th immune responses and susceptibility to many autoimmune diseases. Understanding the role of HLA class II haplotypes in immunity is hampered by the lack of animal models expressing these genes as authentic cis-haplotypes. In this study we describe transgenic expression of the autoimmune prone HLA DR3-DQ2 haplotype from a yeast artificial chromosome (YAC) containing an intact similar320-kb region from HLA DRA to DQB2. In YAC-transgenic mice HLA DR and DQ gene products were expressed on B cells, macrophages, and dendritic cells, but not on T cells indicating cell-specific regulation. Positive selection of the CD4 compartment by human class II molecules was 67% efficient in YAC-homozygous mice lacking endogenous class II molecules (Abeta(null/null)) and expressing only murine CD4. A broad range of TCR Vbeta families was used in the peripheral T cell repertoire, which was also purged of Vbeta5-, Vbeta11-, and Vbeta12-bearing T cells by endogenous mouse mammary tumor virus-encoded superantigens. Expression of the HLA DR3-DQ2 haplotype on the Abeta(null/null) background was associated with normal CD8-dependent clearance of virus from influenza-infected mice and development of CD4-dependent protection from otherwise lethal infection with Salmonella typhimurium. HLA DR- and DQ-restricted T cell responses were also elicited following immunization with known T cell determinants presented by these molecules. These findings demonstrate the potential for human MHC class II haplotypes to function efficiently in transgenic mice and should provide valuable tools for developing humanized models of MHC-associated autoimmune diseases. 相似文献
996.
997.
Norambuena L Marchant L Berninsone P Hirschberg CB Silva H Orellana A 《The Journal of biological chemistry》2002,277(36):32923-32929
The synthesis of non-cellulosic polysaccharides and glycoproteins in the plant cell Golgi apparatus requires UDP-galactose as substrate. The topology of these reactions is not known, although the orientation of a plant galactosyltransferase involved in the biosynthesis of galactomannans in fenugreek is consistent with a requirement for UDP-galactose in the lumen of the Golgi cisternae. Here we provide evidence that sealed, right-side-out Golgi vesicles isolated from pea stems transport UDP-galactose into their lumen and transfer galactose, likely to polysaccharides and other acceptors. In addition, we identified and cloned AtUTr1, a gene from Arabidopsis thaliana that encodes a multitransmembrane hydrophobic protein similar to nucleotide sugar transporters. Northern analysis showed that AtUTr1 is indeed expressed in Arabidopsis. AtUTr1 is able to complement the phenotype of MDCK ricin-resistant cells; a mammalian cell line deficient in transport of UDP-galactose into the Golgi. In vitro assays using a Golgi-enriched vesicle fraction obtained from Saccharomyces cerevisiae expressing AtUTr1-MycHis is able to transport UDP-galactose but also UDP-glucose. AtUTr1- MycHis does not transport GDP-mannose, GDP-fucose, CMP-sialic acid, UDP-glucuronic acid, or UDP-xylose when expressed in S. cerevisiae. AtUTr1 is the first transporter described that is able to transport UDP-galactose and UDP-glucose. Thus AtUTr1 may play an important role in the synthesis of glycoconjugates in Arabidopsis that contain galactose and glucose. 相似文献
998.
The aminoacylase-1/metallopeptidase 20 (Acy1/M20) family is the largest metallopeptidase family. Several crystal structures feature a metal-binding and a dimerization-mediating domain, both arranged in an extended open conformation. We have recently shown [Lindner et al. (2003) J. Biol. Chem. 278, 44496-44504] that in human Acy1 the invariant residues Glu147 and His206 from the metal-binding and the dimerization domain, respectively, are recruited to the active site from opposite dimer subunits. We hypothesized that, to facilitate this, formation of the binary complex is associated with domain closure, which would also position additional residues in the functional active site of Acy1. These would include two partially conserved dimerization domain residues: an asparagine (Asn263) and an arginine (Arg276) from the same subunit as His206 and Glu147, respectively. In this paper, we investigate the significance of the three dimerization domain residues of human Acy1 His206, Asn263, and Arg276 and, additionally, the nearby Asp274 for catalysis using site-directed mutagenesis. Enzyme complementation assays confirm the putative subunit allocations of these residues, and steady-state kinetics support roles for all of them in catalysis but only involve the Arg276 in substrate-binding. The results are consistent with a model of the closed conformation for the structure of the related enzyme carboxypeptidase G2. This study demonstrates experimentally for the first time for a member of the Acy1/M20 family that several residues outside of the metal-binding domain are involved in binding and catalysis. 相似文献
999.
20 S RNA virus is a persistent positive strand RNA virus found in Saccharomyces cerevisiae. The viral genome encodes only its RNA polymerase, p91, and resides in the cytoplasm in the form of a ribonucleoprotein complex with p91. We succeeded in generating 20 S RNA virus in vivo by expressing, from a vector, genomic strands fused at the 3'-ends to the hepatitis delta virus antigenomic ribozyme. Using this launching system, we analyzed 3'-cis-signals present in the genomic strand for replication. The viral genome has five-nucleotide inverted repeats at both termini (5'-GGGGC... GCCCC-OH). The fifth G from the 3'-end was dispensable for replication, whereas the third and fourth Cs were essential. The 3'-terminal and penultimate Cs could be eliminated or modified to other nucleotides; however, the generated viruses recovered these terminal Cs. Furthermore, extra nucleotides added at the viral 3'-end were eliminated in the launched viruses. Therefore, 20 S RNA virus has a mechanism(s) to maintain the correct size and sequence of the viral 3'-end. This may contribute to its persistent infection in yeast. We also succeeded in generating 20 S RNA virus similarly from antigenomic strands provided active p91 was supplied from a second vector in trans. Again, a cluster of four Cs at the 3'-end in the antigenomic strand was essential for replication. In this work, we also present the first conclusive evidence that 20 S and 23 S RNA viruses are independent replicons. 相似文献
1000.
Ernsberger U Esposito L Partimo S Huber K Franke A Bixby JL Kalcheim C Unsicker K 《Cell and tissue research》2005,319(1):1-13
We have analyzed the distribution of neural crest-derived precursors and the expression of catecholaminergic and neuronal markers in developing adrenal tissue of chick embryos. Undifferentiated neural crest cells are found in presumptive adrenal regions from embryonic day 3 (E3) onward. An increasing proportion of cells expressing tyrosine hydroxylase (TH) mRNA indicates catecholaminergic differentiation of precursors not only in primary sympathetic ganglia, but also in presumptive adrenal regions. Whereas precursors and differentiating cells show mesenchymal distribution until E5, discrete adrenal anlagen form during E6. Even during E5, catecholaminergic cells with low or undetectable neurofilament M (NF-M) mRNA expression prevail in positions at which adrenal anlagen become distinct during E6. The predominance of TH-positive and NF-M-negative cells is maintained throughout embryogenesis in adrenal tissue. RNA encoding SCG10, a pan-neuronal marker like NF-M, is strongly expressed throughout adrenal anlagen during E6 but is found at reduced levels in chromaffin cells compared with neuronal cells at E15. Two additional neuronal markers, synaptotagmin 1 and neurexin 1, are expressed at low to undetectable levels in developing chromaffin cells throughout embryogenesis. The developmental regulation of neuronal markers shows at least three different patterns among the four mRNAs analyzed. Importantly, there is no generalized downregulation of neuronal markers in developing adrenal anlagen. Thus, our observations question the classical concept of chromaffin differentiation from a common sympathoadrenal progenitor expressing neuronal properties and suggest alternative models with changing instructive signals or separate progenitor populations for sympathetic neuronal and chromaffin endocrine cells.Chaya Kalcheim and Klaus Unsicker are supported by the Deutsche Forschungsgemeinschaft (SFB 488) 相似文献