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71.
A novel hybrid seed system for plants 总被引:1,自引:0,他引:1
Gils M Marillonnet S Werner S Grützner R Giritch A Engler C Schachschneider R Klimyuk V Gleba Y 《Plant biotechnology journal》2008,6(3):226-235
A two-component hybrid seed system has been developed that is broadly applicable and provides for effective generation and maintenance of the male-sterile parent, hybrid seed production and full restoration of fertility in the hybrid seed. The technology is based on the functional interaction of two loci that are inserted in the same position on two homologous chromosomes, and thus are 'linked in repulsion', and that jointly code for male sterility and herbicide resistance, both traits being expressed in heterozygous plants only. The localization to the same locus on a chromosome is achieved by the genetic transformation of plants with a construct containing both genetic elements (loci), and subsequent derivatization from the primary pro-locus of the two precursor lines using site-specific deletions. The functional interaction of the two loci is achieved through intein-based trans -splicing of two pairs of complementary protein fragments that provide for male sterility and herbicide resistance. Unlike the hybrid seed systems that are currently in use, the technology relies on the genetic modification of just one parent, and is therefore much simpler to develop and use. Arabidopsis has been used for the proof of principle presented here, but the essential elements of the technology are generic and have been shown to work in many crop species. 相似文献
72.
Immunogenicity of HLA-A1-restricted peptides derived from S100A4 (metastasin 1) in melanoma patients
Valeska Hofmeister-Mueller Claudia S. Vetter-Kauczok Ramona Ullrich Katharina Meder Eugene Lukanidin Eva-Bettina Broecker Per thor Straten Mads Hald Andersen David Schrama Juergen C. Becker 《Cancer immunology, immunotherapy : CII》2009,58(8):1265-1273
S100A4 (metastasin 1) belongs to the S100 family of Ca2+ binding proteins. While not present in most differentiated adult tissues, S100A4 is upregulated in the micromilieu of tumors.
It is primarily expressed by tumor-associated macrophages, fibroblasts, and tumor endothelial cells. Due to its strong induction
in tumors S100A4 is a promising target for cancer immunotherapy. By reverse immunology, using epitope prediction programs,
we identified 3 HLA-A1-restricted peptide epitopes (S100A4 A1-1, A1-2, and A1-3) which are subject to human T cell responses
as detected in peripheral blood of melanoma patients by means of IFN-γ ELISPOT and cytotoxicity assays. In addition, IFN-γ
responses to S100A4 A1-2 can not only be induced by stimulation of T cells with peptide-loaded DC but also by stimulation
with S100A4 protein-loaded DC, indicating that this epitope is indeed generated by processing of the endogenously expressed
protein. In addition, S100A4 A1-2 reactive T cells demonstrate lysis of HLA-A1+ fibroblasts in comparison to HLA-A1− fibroblasts. In summary, this HLA-A1-restricted peptide epitope is a candidate for immunotherapeutical approaches targeting
S100A4-expressing cells in the tumor stroma. 相似文献
73.
Dorothee Dormann Anja Capell Aaron M. Carlson Sunita S. Shankaran† Ramona Rodde Manuela Neumann‡ Elisabeth Kremmer§ Takashi Matsuwaki¶ Keitaro Yamanouchi¶ Masugi Nishihara¶ Christian Haass 《Journal of neurochemistry》2009,110(3):1082-1094
Neuronal and glial deposition of misfolded, proteolytically processed, polyubiquitinated and abnormally phosphorylated C-terminal fragments (CTFs) of the TAR DNA binding protein-43 (TDP-43) is a pathological hallmark of frontotemporal lobar degeneration with ubiquitin positive inclusions (FTLD-U) and certain cases of amyotrophic lateral sclerosis. We demonstrate that TDP-43 can be proteolytically processed by caspases upon induction of apoptosis to a major 35 kDa and a minor 25 kDa CTF. These fragments are initially soluble, but over time they accumulate as insoluble and pathologically phosphorylated derivatives. However, proteolytic processing appears not to be absolutely required for the deposition of insoluble TDP-43 species, since a caspase resistant mutant of TDP-43 is also converted into insoluble species. Phosphorylation at S409/410 apparently occurs late during the conversion of soluble to insoluble TDP-43, suggesting that phosphorylation is not a prerequisite for aggregation. Loss of function of the progranulin (PGRN) gene causes FTLD-U with TDP-43 positive inclusions and has been suggested to lead to caspase activation and subsequent TDP-43 processing. However, siRNA-mediated knockdown of PGRN in cell culture as well as a PGRN gene knockout in mice failed to cause the formation of the disease characterizing CTFs of TDP-43. Our findings therefore suggest that caspase-mediated processing generates CTFs of similar biochemical properties as those occurring in nuclear and cytoplasmic deposits of FTLD-U patients independent of PGRN levels. 相似文献
74.
It is of fundamental importance to understand the physiological differences leading to salt resistance and to get access to the molecular mechanisms underlying this physiological response. The aim of this work was to investigate the effects of short‐term salt exposure on the proteome of maize chloroplasts in the initial phase of salt stress (up to 4 h). It could be shown that sodium ions accumulate quickly and excessively in chloroplasts in the initial phase of moderate salt stress. A change in the chloroplast protein pattern was observed without a change in water potential of the leaves. 2‐DE revealed that 12 salt‐responsive chloroplast proteins increased while eight chloroplast proteins decreased. Some of the maize chloroplast proteins such as CF1e and a Ca2+‐sensing receptor show a rather transient response for the first 4 h of salt exposure. The enhanced abundance of the ferredoxin NADPH reductase, the 23 kDa polypeptide of the photosystem II, and the FtsH‐like protein might reflect mechanism to attenuate the detrimental effects of Na+ on the photosynthetic machinery. The observed transient increase and subsequent decrease of selected proteins may exhibit a counterbalancing effect of target proteins in this context. Intriguingly, several subunits of the CF1–CF0 complex are unequally affected, whereas others do not respond at all. 相似文献
75.
Kenneth M. D. Rutherford Sheena K. Robson Ramona D. Donald Susan Jarvis Dale A. Sandercock E. Marian Scott Andrea M. Nolan Alistair B. Lawrence 《Biology letters》2009,5(4):452-454
Pre-natal stress (PNS) or undernutrition can have numerous effects on an individual''s biology throughout their lifetime. Some of these effects may be adaptive by allowing individuals to tailor their phenotype to environmental conditions. Here we investigated, in the domestic pig Sus scrofa, whether one possible consequence of a predicted adverse environment could be altered pain perception. The behavioural response of piglets to the surgical amputation (‘docking’) of their tail or a sham procedure was measured for 1 min in piglets born to mothers who either experienced mid-gestation social stress or were left undisturbed throughout pregnancy. A behavioural pain score was found to predict the docked status of piglets with high discriminant accuracy. Piglets exposed to PNS had a significantly higher pain score than controls, and for each litter of tail-docked piglets, the average pain score was correlated with mid-gestation maternal cortisol levels. The data presented here provide evidence that the experience of stress in utero can result in a heightened acute response to injury in early life. Speculatively, this may represent an adaptive alteration occurring as a consequence of a pre-natal ‘early warning’ of environmental adversity. 相似文献
76.
Ramona Schmiedel E. Bartholomeus Kuettner Antje Keim Norbert Sträter Thomas Greiner-Stöffele 《DNA Repair》2009,8(2):219-231
The major AP endonuclease in Escherichia coli Exonuclease III (ExoIII) is frequently used in gene technology due to its strong exonucleolytic activity. A thermostabilized variant of ExoIII or a homologous enzyme from thermophilic organisms could be most useful for further applications. For this purpose we characterized a nuclease from the hyperthermophilic archaeon Archaeoglobus fulgidus (Af_Exo), which shares 33% overall sequence identity and 55% similarity to ExoIII. The gene coding for this thermostable enzyme was cloned and expressed in E. coli. The purified protein shows a strong Mg2+-dependent nicking activity at AP-sites, nicking of undamaged double-stranded (ds) DNA and a weak exonucleolytic activity. A V217G variant of the enzyme was crystallized with decamer ds-DNA molecule, and the three-dimensional structure was determined to 1.7 Å resolution. Besides our goal to find or produce a thermostable exonuclease, the structural and catalytic data of Af_Exo and a series of mutant proteins, based on the crystal structure, provide new insight into the mechanism of abasic site recognition and repair. Each of the hydrophobic residues Phe 200, Trp 215 and Val 217, forming a binding pocket for the abasic deoxyribose in Af_Exo, were mutated to glycine or serine. By expanding the size of the binding pocket the unspecific endonucleolytic activity is increased. Thus, size and flexibility of the mostly hydrophobic binding pocket have a significant influence on AP-site specificity. We suggest that its tight fitting to the flipped-out deoxyribose allows for a preferred competent binding of abasic sites. In a larger or more flexible pocket however, intact nucleotides more easily bind in a catalytically competent conformation, resulting in loss of specificity. Moreover, with mutations of Phe 200 and Trp 215 we induced a strong exonucleolytic activity on undamaged DNA. 相似文献
77.
DEAD box RNA helicases use the energy of ATP hydrolysis to unwind double-stranded RNA regions or to disrupt RNA/protein complexes. A minimal RNA helicase comprises nine conserved motifs distributed over two RecA-like domains. The N-terminal domain contains all motifs involved in nucleotide binding, namely the Q-motif, the DEAD box, and the P-loop, as well as the SAT motif, which has been implicated in the coordination of ATP hydrolysis and RNA unwinding. We present here the crystal structure of the N-terminal domain of the Thermus thermophilus RNA helicase Hera in complex with adenosine monophosphate (AMP). Upon binding of AMP the P-loop adopts a partially collapsed or half-open conformation that is still connected to the DEAD box motif, and the DEAD box in turn is linked to the SAT motif via hydrogen bonds. This network of interactions communicates changes in the P-loop conformation to distant parts of the helicase. The affinity of AMP is comparable to that of ADP and ATP, substantiating that the binding energy from additional phosphate moieties is directly converted into conformational changes of the entire helicase. Importantly, the N-terminal Hera domain forms a dimer in the crystal similar to that seen in another thermophilic prokaryote. It is possible that this mode of dimerization represents the prototypic architecture in RNA helicases of thermophilic origin. 相似文献
78.
Baum P Schmid R Ittrich C Rust W Fundel-Clemens K Siewert S Baur M Mara L Gruenbaum L Heckel A Eils R Kontermann RE Roth GJ Gantner F Schnapp A Park JE Weith A Quast K Mennerich D 《PloS one》2010,5(12):e14272
A phenocopy is defined as an environmentally induced phenotype of one individual which is identical to the genotype-determined phenotype of another individual. The phenocopy phenomenon has been translated to the drug discovery process as phenotypes produced by the treatment of biological systems with new chemical entities (NCE) may resemble environmentally induced phenotypic modifications. Various new chemical entities exerting inhibition of the kinase activity of Transforming Growth Factor β Receptor I (TGF-βR1) were qualified by high-throughput RNA expression profiling. This chemical genomics approach resulted in a precise time-dependent insight to the TGF-β biology and allowed furthermore a comprehensive analysis of each NCE's off-target effects. The evaluation of off-target effects by the phenocopy approach allows a more accurate and integrated view on optimized compounds, supplementing classical biological evaluation parameters such as potency and selectivity. It has therefore the potential to become a novel method for ranking compounds during various drug discovery phases. 相似文献
79.
Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues 下载免费PDF全文
80.