90,774 research outputs found
KIF14 and citron kinase act together to promote efficient cytokinesis
No full textMultiple mitotic kinesins and microtubule-associated proteins (MAPs) act in concert to direct cytokinesis (Glotzer, M. 2005. Science. 307:1735-1739). In anaphase cells, many of these proteins associate with an antiparallel array of microtubules termed the central spindle. The MAP and microtubule-bundling protein PRC1 (protein-regulating cytokinesis 1) is one of the key molecules required for the integrity of this structure (Jiang, W., G. Jimenez, N.J. Wells, T.J. Hope, G.M. Wahl, T. Hunter, and R. Fukunaga. 1998. Mol. Cell. 2:877-885; Mollinari, C., J.P. Kleman, W. Jiang, G. Schoehn, T. Hunter, and R.L. Margolis. 2002. J. Cell Biol. 157:1175-1186). In this study, we identify an interaction between endogenous PRC1 and the previously uncharacterized kinesin KIF14 as well as other mitotic kinesins (MKlp1/CHO1, MKlp2, and KIF4) with known functions in cytokinesis (Hill, E., M. Clarke, and F.A. Barr. 2000. EMBO J. 19:5711-5719; Matuliene, J., and R. Kuriyama. 2002. Mol. Biol. Cell. 13:1832-1845; Kurasawa, Y., W.C. Earnshaw, Y. Mochizuki, N. Dohmae, and K. Todokoro. 2004. EMBO J. 23:3237-3248). We find that KIF14 targets to the central spindle via its interaction with PRC1 and has an essential function in cytokinesis. In KIF14-depleted cells, citron kinase but not other components of the central spindle and cleavage furrow fail to localize. Furthermore, the localization of KIF14 and citron kinase to the central spindle and midbody is codependent, and they form a complex depending on the activation state of citron kinase. Contrary to a previous study (Di Cunto, F., S. Imarisio, E. Hirsch, V. Broccoli, A. Bulfone, A. Migheli, C. Atzori, E. Turco, R. Triolo, G.P. Dotto, et al. 2000. Neuron. 28:115-127), we find a general requirement for citron kinase in human cell division. Together, these findings identify a novel pathway required for efficient cytokinesis
Drosophila citron kinase is required for the final steps of cytokinesis
No full textThe mechanisms underlying completion of cytokinesis are still poorly understood. Here, we show that the Drosophila orthologue of mammalian Citron kinases is essential for the final events of the cytokinetic process. Flies bearing mutations in the Drosophila citron kinase (dck) gene were defective in both neuroblast and spermatocyte cytokinesis. In both cell types, early cytokinetic events such as central spindle assembly and contractile ring formation were completely normal. Moreover, cytokinetic rings constricted normally, leading to complete furrow ingression. However late telophases of both cell types displayed persistent midbodies associated with disorganized F actin and anillin structures. Similar defects were observed in dck RNA interference (RNAi) telophases, which, in addition to abnormal F actin and anillin rings, also displayed aberrant membrane protrusions at the cleavage site. Together, these results indicate that mutations in the dck gene result in morphologically abnormal intercellular bridges and in delayed resolution of these structures, suggesting that the wild-type function of dck is required for abscission at the end of cytokinesis. The phenotype of Dck-depleted cells is different from those observed in most Drosophila cytokinesis mutants but extraordinarily similar to that caused by anillin RNAi, suggesting that Dck and anillin are in the same pathway for completion of cytokinesis
Antimicrobial Efficacy of Citron Essential Oil on Spoilage and Pathogenic Microorganisms in Fruit-Based Salads
No full textThe aim of this study was the evaluation of the effects of pure citral and citron essential oil on microbial
spoilage and growth and survival of pathogenic microorganisms during storage. They were added in the syrup
of industrial ready-to-eat fruit salads stored at 9 ◦C. Both citral (25 to 125 ppm) and citron essential oil (300, 600,
900 ppm) were able to prolong the microbial shelf life of the fruit-based salads. The essential oil gave excellent results,
avoiding the undesirable effects attributable to the cytotoxicity of citral. Citron essential oil doubled the time
needed for the wild microflora to reach concentrations able to produce a perceivable spoilage in condition of thermal
abuse (9 ◦C). The same essential oil had reduced effects on the survival of Gram-negative species Salmonella
Enteritidis and Escherichia coli, but showed a strong inhibition toward the Gram-positive pathogen Listeria monocytogenes
Role of YAP/TAZ downstream of common oncogenic drivers
No full textYAP and TAZ are transcriptional coactivators that play a central role in cancer initiation and growth. Despite being widespread activated in a number of human malignancies, YAP/TAZ aberrant triggering is rarely due to genetic alteration, but it is likely occurring as a consequence of mis-regulation of the complex network of signaling pathways that converge on these factors. We thus asked if recurrent oncogenic lesions, that are considered as tumorigenic drivers, might work though YAP/TAZ to induce cell transformation. We found that YAP/TAZ are not only activated upon oncogenic stimulation, but are required for the onset of malignant phenotypes induced by oncogenes in carcinogenesis models both in vitro and in vivo. Interestingly, we also found that the physical composition of the extracellular environment is crucial to allow oncogenes to foster tumor initiation and progression, by sustaining YAP/TAZ activity. Indeed, cells that are not able to properly organize F-actin cytoskeleton are refractory to oncogene-induced transformation. The finding that YAP/TAZ are the molecular nexus on which different tumorigenic drivers converge, and that oncogenes mediated transformation requires proficient transduction of mechanical signals to YAP/TAZ, might have relevant implications for the identification of a common molecular target to inhibit tumor onset
Biological properties and resistance reversal effect of Helichrysum i Diversity of citron (Citrus medica L.) and phylogenetic analysis of related citron hybrids using molecular markers and essential oil compositions.
No full textInternational audienceCitrus diversity is known to be essentially supported by three essential basic species, mandarin, pummelo and citron, which are at the origin of quite all the cultivated citrus. Based on studies of phenotype, the citron group (Citrus medica) is supposedly low in genetic diversity compared to mandarins and pummelos. To assess the diversity of the citron group we analyzed the molecular polymorphism of nuclear and cytoplasmic genetic markers and the variation in composition of essential oils from leaves and peels of 24 varieties of citron or apparent hybrids of citron. Our results demonstrate that the citron group has a relatively high allelic diversity, probably due to multiple introductions of different varieties. Self-fertilization was the mode of reproduction which most frequently led to the development of commercial varieties with specific aromas and flavors, such as the cultivar ‘Corsican’, a putative ‘Common Poncire’ self-crossed hybrid. However, some varieties appeared to be interspecific hybrids related to citron but only by the male parent. This varietal diversity was probably favored by spreading seeds to extend the cultivation of citron in Mediterranean countries. The chemical diversity does not always correspond to genetic diversity but nevertheless reveals specific profiles associated with some genotypes
Molecular cloning, expression and functional analyses of citron kinase in zebrafish embryos
No full textCitron kinase (CRIK) 屬於絲胺酸/酥胺酸磷酸激酶，並為已活化Rho的下游作用蛋白之一。若於體外培養之哺乳細胞中過度表現喪失功能之CRIK突變種會造成細胞分裂的缺陷，此結果顯示CRIK會影響細胞分裂，其於細胞分裂溝及神經組織之聚集更增加其調控細胞分裂及神經發育之可能性。在果蠅發育過程CRIK雖為各類組織細胞質分裂所必需，而CRIK基因剔除老鼠胚胎早期可發育並出生，但於出生後伴隨明顯神經細胞分裂缺陷和凋亡而死亡。CRIK基因可能在不同物種間有差異或有功能相近之同源基因存在，而導致其在不同物種之功能差異，為探討CRIK在發育過程的基因表現及其功能，我們在模式動物斑馬魚中以in silica cloning和RACE方式選殖出斑馬魚CRIK基因(zcrik)，比較分析其與其他物種CRIK蛋白質序列，利用RT-PCR偵測到成魚腦、眼睛、腎臟、胰臟、卵巢及睪丸組織中皆有zcrik的表現，在原位雜合試驗(in situ hybridization)中可見在發育24小時胚中zcrik表現多在細胞具有高度增殖的區域，如神經先驅細胞等，此現象與先前發表在老鼠表現模式相類似。為了進一步探討zcrik在斑馬魚發育的功能，我們顯微注射morpholino oligonucleotides (MO)至一胞期胚來抑制zcrik mRNA之轉譯，結果顯示zcrik-MO處理過之胚在發育六小時內死亡率較對照組高，並於早期細胞分裂時期有分裂不完整與不正常分裂情形發生，此分裂嚴重受干擾的胚亦在之後細胞遷移過程產生障礙，使得胚無法繼續正常發育而死亡，而存活之胚在受精後24小時觀察其外觀上有明顯無頭和神經管消失等缺陷發生，此結果和先前研究中指出CRIK對神經細胞分裂發育有影響相呼應，再則此等胚之缺陷與過度腹部化(ventralized)的胚型態相類似，由此推測z CRIK在胚發育時期不僅具有調節細胞分裂與神經發育的功能，對於斑馬魚的原腸胚發育過程及背腹部化亦有一定程度的影響。Citron kinase (CRIK) belongs to an evolutionarily conserved family of serine-threonine kinases and is a target molecule for activated Rho. CRIK has been suggested to be involved in regulating cytokinesis in mammalian cells due to the inhibition of cytokinesis by overexpressing dominant negative CRIK. It accumulates at the cleavage furrow and neuronal tissues that implies its roles in early development and neurogenesis. In Drosophila melanogaster, CRIK is essential for normal cell division in all tissue of Drosophila, while the knock out citron kinase allows early embryonic development, but shows defective neurogenesis in mice. To further study the role of CRIK in development, we utilized the recently developed vertebrate model, zebrafish. In this study, we first isolated the zebrafish citron kinase (zcrik) by in silica cloning and RACE experiments. Secondly, the zcrik expression patterns were examined using in situ hybridization and RT-PCR. We found that zcrik mRNA was detected by RT-PCR in adult zebrafish tissues including brain, eye, ovary, testis, kidney, spleen and gill. Whole-mount in situ hybridization of different developmental stages of zebrafish embryos revealed that zcrik expressed in the entire embryo until 18 hour post fertilization and the expression of zcrik mRNA was limit to the proliferate neuroblasts and tissues that is consistent with the expression pattern in mice. Lastly, to evaluate the functions of CRIK in zebrafish development, we applied etopic expression analysis and morpholino oligonucleotides (MO) knockdown to the developing zebrafish embryos. Functional analysis results demonstrated that gain or loss of function of zCRIK resulted defects in cytokinesis, epiboly formation and embryonic death. In addition, the MO-knockdown resulted in severe defects in CNS with little or no head structure formation, which are the characteristics of ventralized embryos at 24 hpf. These results suggest that, in addition to cytokinesis and neuronal development, citron kinase is also essential for gastrulation progressing and may be involved in dorsal-ventral patterning during development.CONTENTS
CONTENTS………………………………………………………………………Ⅰ
LIST OF TABLE……………………………………………………………………Ⅲ
LIST OF FIGURES………………………………………………………………Ⅳ
中文摘要……………………………………………………………………………1
ABSTRACT…………………………………………………………………………2
INTRODUCTION……………………………………………………………4
MATERIALS AND METHODS…………………………………………………9
Cloning of citron kinase cDNA……………………………………………………9
Blasting…………………………………………………………………………9
RNA isolation…………………………………………………………………9
Single-strand cDNA synthesis ……………………………………………10
PCR amplification……………………………………………………………11
Purification of PCR products and subcloning……………………11
Mini-preparation of plasmid DNA………………………………………12
Expression vector construction and in vitro transcription………12
Subcloning of zCRIK kinase domain fragment into T7TS expression vector12
Synthesis of mRNA of zcrik-k………………………………………………12
Antisense Morpholino design and preparation…………………………………14
Microinjection and observation…………………………………………………14
Whole mount in situ hybridization……………………………………………15
Staining of actin cytoskeleton…………………………………………………15
RESULTS…………………………………………………………………………16
Isolation of the zebrafish citron kinase homolog………………………………16
Expression of zebrafish citron kinase during development……………………18
Overexpression zebrafish citron kinase domain during development…………19
Knockdown zebrafish citron kinase by morpholino antisense oligonucleotides…19
DISCUSSION………………………………………………………………………23
REFERENCES……………………………………………………………………33
TABLE………………………………………………………………………………42
FIGURES…………………………………………………………………………4
First in the Nation in Education : Final Report,1984.
No full textThis report is one step in an ongoing process of change and is a plea for commitment for high standards in education in Iowa. Contains the final reports of the six subcommittees as adopted by the Excellence in Education Task Force, and the five recommendations made by the Task Force
Variations on the Author
No full text“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Chemotaxonomic Investigations of Peel and Petitgrain Essential Oils from 17 Citron Cultivars
No full textInternational audienceChemical compositions of essential oils from 17 citron cultivars were studied using GC and GC/MS. To the best of our knowledge, chemical compositions of peel and petitgrain oils from seven of them were reported for the first time. Combined analysis of peel and petitgrain essential oils led to the identification of 37 components (amounting to 98.2-99.9% of the total oil) and 42 components (97.0-99.9%), respectively. Statistical analysis was applied to identify possible relationships between citron cultivars. The levels of seven components, i.e., limonene, -pinene, -terpinene, neral, geranial, nerol, and geraniol, indicated that the cultivars could be classified in four main chemotypes for peel and petitgrain oils. Chemotaxonomic investigations were carried out to establish relations between the morphological characteristics of citron cultivars and their corresponding oil compositions
Effects of RFI and Underground Condition on PWM Current-Mode Integrated Circuits for SMPS
No full textThe effects of a radio frequency conducted disturbing signal are evaluated on several pin-to-pin compatible control integrated circuits (IC) for PWM current-mode switching power supplies. The tested ICs were selected between some common models available on the market. The variations of the primary functional parameters (inner voltage reference, switching frequency, etc.) are investigated in conditions of different frequency and amplitude for the interfering signal. A dedicated PCB for the measurements was developed paying particular attention to HF layout. Also, the impact of underground condition (voltage at one pin below ground reference) on ICs’ behaviour is analysed. The paper gives a comparative view of the results, focusing on the key macroscopic effects
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