bluuebunny/crossref_metadata_2025_split huggingface dataset:
Importance of spatial activation of Cdc42 and Rac small G proteins by frabin for microspike formation in MDCK cells
Authors: Yasuda Takeo , Ohtsuka Toshihisa , Inoue Eiji , Yokoyama Shigekazu , Sakisaka Toshiaki , Kodama Atsuko , Takaishi Kenji , Takai Yoshimi 
Published: March 2003

BackgroundFrabin is an actin filament (F‐actin)‐binding protein that shows GDP/GTP exchange activity for Cdc42 small G protein (Cdc42). Frabin furthermore induces indirect activation of Rac small G protein (Rac) in intact cells. We have recently shown that in nonepithelial cells, frabin induces the formation of both filopodia‐ and lamellipodia‐like processes through the activation of Cdc42 and Rac, respectively. In epithelial cells such as MDCK cells, Cdc42 and Rac regulate cell‐cell adherens junctions (AJs) via the accumulation of F‐actin and E‐cadherin, although neither Cdc42 nor Rac induces the formation of filopodia or lamellipodia. In this study, we have examined the effects of frabin on the reorganization of the actin cytoskeleton in MDCK cells.ResultsFrabin induces the formation of microspikes at the basal area of the lateral membranes through the activation of Cdc42 and Rac in MDCK cells, although a dominant active mutant of Cdc42 or Rac alone, or both, did not induce the formation of microspikes. Furthermore, frabin weakly increased the accumulation of F‐actin and E‐cadherin at cell‐cell AJs and the formation of stress fibres through the activation of Cdc42 and Rac, under conditions where the dominant active mutant of Cdc42 or Rac markedly showed these effects. The Cdc42‐ and Rac‐induced formation of stress fibres was dependent on the activation of Rho small G protein.ConclusionThese results indicate that the frabin‐dependent spatial activation of Cdc42 and Rac is important for the formation of microspikes.
The role of tightly bound ATP in Escherichia coli tRNA nucleotidyltransferase
Authors: Tomari Yukihide , Suzuki Tsutomu , Watanabe Kimitsuna , Ueda Takuya 
Published: March 2003

BackgroundThe CCA‐adding enzyme [ATP(CTP): tRNA nucleotidyltransferase (EC. 2.7.7.25)] catalyses the addition of the conserved CCA sequence to the 3′‐terminus of tRNAs. All CCA‐adding enzymes are classified into the nucleotidyltransferase superfamily. In the absence of ATP, the Escherichia coli CCA‐adding enzyme displays anomalous poly(C) polymerase activity.ResultsWe show that CCA‐adding enzyme over‐expressed in E. coli exists in an ATP‐bound form. The affinities of ATP and CTP towards the enzyme were estimated by several methods, and the dissociation constants for ATP and CTP were determined to be 6.3 and 188 μm, respectively. AMP‐incorporation terminated the nucleotidyltransferase reaction, while in the absence of ATP, the enzyme continued poly(C) polymerization. In the case of a tRNA substrate with a mutation in the T‐loop region, normal CC was added at a much slower rate compared with the wild‐type, but anomalous poly(C) polymerization occurred at the same rate as in the wild‐type.ConclusionBased on the findings outlined above, we concluded that the E. coli CCA‐adding enzyme possesses at least two distinct nucleotide binding sites, one responsible for ATP binding and the other(s) for CTP binding. The addition of ATP from the tight ATP binding site terminates nucleotide incorporation, thus limiting poly(C) polymerization to CCA. It is also suggested that during anomalous poly(C) polymerization, tRNA translocates from the tRNA binding site upon the third C addition.
Expression of herpes simplex virus type 2 US3 affects the Cdc42/Rac pathway and attenuates c‐Jun N‐terminal kinase activation
Authors: Murata Takayuki , Goshima Fumi , Daikoku Tohru , Takakuwa Hiroki , Nishiyama Yukihiro 
Published: March 2003

Although the US3 gene product of herpes simplex virus (HSV) has been identified as a serine/threonine protein kinase (PK), the functions are poorly understood.We found that US3 PK of HSV‐2 induced disruption of actin filaments, cell rounding and accumulation of binucleate cells in HEp2 cells. Cell rounding was abrogated by expression of either kinase‐dead forms of US3 PK or a mutant protein lacking the acidic cluster in the kinase regulatory domain. Co‐expression of dominant active forms of Cdc42/Rac inhibited cell rounding, suggesting that a signal transduction pathway involving Cdc42/Rac may play a role in the morphological changes induced by the kinase. Cdc42 and Rac, members of the Rho family of small GTPases, function as molecular switches changing actin cytoskeletal organization, influencing transcription and controlling apoptotic cell death. By computed homology search, we noticed significant similarities between US3 PK and p21‐activated kinase (PAK), which is activated by the Cdc42 or Rac. We also found that the expression of US3 suppressed the activation of c‐Jun N‐terminal kinase (JNK), a kinase that is downstream of PAK.These observations suggest that the US3 PK affects the Cdc42/Rac pathway and can act as an upstream suppressor of JNK in the stress‐activated signalling pathway.
Regulation of gene expression, cellular localization, and in vivo function of Caenorhabditis elegans DNA topoisomerase I
Authors: Lee Myon Hee , Park Hyungki , Shim Gaegal , Lee Junho , Koo Hyeon‐Sook 
Published: March 2003

BackgroundDNA topoisomerase I is dispensable in yeast, but is essential during the embryogenesis of Drosophila and mouse. In order to determine functions of the enzyme in the development of Caenorhabditis elegans, phenotypes resulting from the deficiency were observed and correlated with the expression of the gene.ResultsThe transcriptional regulation of the C. elegans DNA topoisomerase I gene was investigated by mRNA localization and reporter gene expression in C. elegans. The mRNA was expressed in the gonad and in the early embryos, followed by a rapid decrease in its level during the late embryonic stage. A reporter gene expression induced by the 5′‐upstream DNA sequence appeared at the comma stage of embryos, continued through the L1 larval stage, and began to decrease gradually afterwards. The DNA topoisomerase I protein was immuno‐localized in the nuclei of meiotic gonad cells and interphase embryonic cells, and unexpectedly in centrosomes of mitotic embryonic cells. Double‐stranded RNA interference of DNA topoisomerase I gene expression resulted in pleiotropic phenotypes showing abnormal gonadogenesis, oocyte development and embryogenesis.ConclusionThese phenotypes, along with expressional regulations, demonstrate that DNA topoisomerase I plays important roles in rapidly growing germ cells and embryonic cells.
Arabidopsis transcriptional regulation by light stress via hydrogen peroxide‐dependent and ‐independent pathways
Authors: Kimura Mitsuhiro , Yoshizumi Takeshi , Manabe Katsushi , Yamamoto Yoshiharu Y. , Matsui Minami 
Published: March 2003

Background High (intense) light stress causes the formation of oxygen radicals in chloroplasts and has the potential to damage them. However, plants are able to respond to this stress and protect the chloroplasts by various means, including transcriptional regulation in the nucleus. Although the corresponding signalling pathway is largely unknown, the high light response in the expression of the Arabidopsis APX2 gene is reported to be mediated by hydrogen peroxide.Results We characterized light stress signalling by analysing expression profiles of another high light‐inducible gene of Arabidopsis, ELIP2, with the aid of an ELIP2 promoter‐luciferase gene fusion. The established ELIP2::LUC transgenic Arabidopsis showed activation by high light, but not by hydrogen peroxide. On the other hand, the native ELIP2 gene as well as the APX2 gene was activated by the hydrogen peroxide. The activation of ELIP2::LUC by intense light was not inhibited by K252a but by okadaic acid.Conclusion The light stress signalling from the chloroplast to the nucleus is revealed to be mediated through at least two pathways: both hydrogen peroxide‐dependent and –independent. The latter pathway is thought to be mediated by the protein phosphatase 2A/1 activity that is suppressed by okadaic acid.
Bir1/Cut17 moving from chromosome to spindle upon the loss of cohesion is required for condensation, spindle elongation and repair
Authors: Morishita Jun , Matsusaka Takahiro , Goshima Gohta , Nakamura Takahiro , Tatebe Hisashi , Yanagida Mitsuhiro 
Published: March 2003

Background In mammals, proteins containing BIR domains (IAPs and survivin) are implicated in inhibiting apoptosis and sister chromatid separation. In the nematode, Bir1 is required for a proper localization of aurora kinase, which moves from the mitotic chromosome in metaphase to the spindle midzone in anaphase as a passenger. Fission yeast Bir1/Pbh1 is essential for normal mitosis.Results A temperature sensitive mutant cut17‐275 exhibits the defect in condensation and spindle elongation at 36 °C, while securin is degraded. Gene cloning shows that the cut17+ gene is identical to bir1+/pbh1+. At 26 °C, cut17‐275 is UV sensitive as the repair of DNA damage is severely compromised. Bir1/Cut17 is a nuclear protein in interphase, which is then required for recruiting condensin to the mitotic nucleus, and concentrates to form a discrete number of dots from prometaphase to metaphase. Once the chromatids are separated, Bir1/Cut17 no longer binds to kinetochores and instead moves to the middle of spindle. Chromatin immunoprecipitation suggested that Bir1/Cut17 associates with the outer repetitious centromere region in metaphase. Following the initiation of anaphase the protein switches from being a chromosomal protein to a spindle protein. This transit is stringently regulated by the state of sister chromatid cohesion proteins Mis4 and Rad21. Ark1, is an aurora kinase homologue whose mitotic distribution is identical to, and under the control of Bir1/Cut17.Conclusions Bir1/Cut17 and Ark1 act as ‘passengers’ but they may play a main role as a recruitment factor, essential for condensation, spindle elongation and DNA repair. Bir1/Cut17 should have roles both in mitotic and in interphase chromosome. The proper location of Ark1 requires Bir1/Cut17, and the mitotic localization of Bir1/Cut17 requires sister cohesion.
Regulation of the activity of the transcription factor Runx2 by two homeobox proteins, Msx2 and Dlx5
Authors: Shirakabe Kyoko , Terasawa Kazuya , Miyama Katsuyoshi , Shibuya Hiroshi , Nishida Eisuke 
Published: March 2003

Background Runx2, formerly called PEBP2αA or Cbfa1, is a transcription factor whose deletion causes a complete lack of ossification. It directly regulates the expression of osteoblast‐specific genes through the osteoblast‐specific cis‐acting element found in the promoter region of these genes.Results In this study, we have found conditions in which induction of the expression of Runx2 is not accompanied by expression of an osteoblast‐specific gene, osteocalcin in C2C12 cells. This finding suggests the existence of a repressor of the activity of Runx2. We have then found that the homeobox protein Msx2 is able to repress the transcription activity of Runx2 by interacting with it. Furthermore, our results have shown that the other homeobox protein Dlx5 has an activity which interferes with both abilities of Msx2 to interact with Runx2 and repress its transcription activity. It has previously been shown that a missense mutation of Msx2 (P148H) causes Boston‐type craniosynostosis in humans. Interestingly, while this mutant form of Msx2 was able to bind to Runx2 and repress its activity, these abilities of Msx2 (P148H) were not subject to regulation by Dlx5.Conclusion These results suggest that regulation of the activity of Runx2 by Msx2 and Dlx5 plays an important role in the mammalian skull development.
ICRF‐193, a catalytic inhibitor of DNA topoisomerase II, inhibits re‐entry into the cell division cycle from quiescent state in mammalian cells
Authors: Hossain Muktadir S. , Akimitsu Nobuyoshi , Takaki Tohru , Hirai Hiroshi , Sekimizu Kazuhisa 
Published: March 2003

AbstractBackground: To describe the requirement of DNA topoisomerase II (topo II) during transition from the quiescent state (G0 phase) to the cell division cycle in mammalian cells, we examined the influence of ICRF‐193, a catalytic inhibitor of topo II, on re‐entry into the cell division cycle of quiescent cells in response to appropriate growth stimuli.Results: The re‐entry into the S phase of cultured cell lines arrested at the quiescent (G0) phase by serum‐starvation was sensitive to 10 µm ICRF‐193. DNA syntheses induced by lipopolysaccharide in murine spleen cells or by release from contact‐inhibition were also inhibited by ICRF‐193. The cell lines with a high‐level of resistance toward ICRF‐193 due to a point mutation in the topo IIα gene entered into the S phase from quiescence in the presence of ICRF‐193. The drug did not inhibit entry into the S phase in cultured cells released from arrest at the metaphase or G1 phase.Conclusion: There is an ICRF‐193‐sensitive step during re‐entry of quiescent mammalian cells into the cell division cycle upon growth stimulation and the drug targets topo IIα during the process.
Genome‐wide analysis of changes in early gene expression induced by oestrogen
Authors: Watanabe Hajime , Suzuki Atsuko , Mizutani Takeshi , Khono Satomi , Lubahn Dennis B. , Handa Hiroshi , Iguchi Taisen 
Published: March 2003

AbstractBackground: The sex hormone 17β‐oestradiol (E2) has profound effects on many aspects of reproduction, development, as well as behaviour. Although the oestrogen receptor is well characterized on a molecular level, relatively few genes affected by E2 have been identified, and the mechanisms underlying the physiological changes caused by E2 are largely unknown. In order to identify oestrogen‐regulated genes in vivo, early uterine gene expression profiles were developed using DNA microarrays.Results: Ovariectomized mice were exposed to 17β‐oestradiol for 6 h, and mRNA expression analysis for 9977 genes was performed. Although a large number of genes was affected by oestrogen administration, the genes that showed higher reproducibility in repetitive experiments were selected and further examined. For most of the selected genes, expression was induced in a dose‐dependent manner, and gene expression was not altered following oestrogen treatment in oestrogen receptor‐α (ERα)‐deficient mice. In combination with the estimation of gene expression levels using quantitative PCR, it was revealed that multiple genes related to sterol biosynthesis, tRNA synthesis, RNA processing, and growth signalling were activated. Based on the microarray data, we selected additional genes related to sterol biosynthesis and tRNA synthesis and confirmed that these genes are also activated by oestrogen.Conclusion: Genes suggesting a basis for the drastic uterotrophic effect observed several days following oestrogen administration were identified. These findings not only reveal the diverse effect of oestrogen signalling on transcript levels in vivo but also demonstrate the ability of DNA microarrays to identify cellular pathways affected by oestrogen.
MCM3‐binding GANP DNA‐primase is associated with a novel phosphatase component G5PR
Authors: Kono Yoshihiko , Maeda Kazuhiko , Kuwahara Kazuhiko , Yamamoto Hideyuki , Miyamoto Eishichi , Yonezawa Kazuyoshi , Takagi Katsumasa , Sakaguchi Nobuo 
Published: March 2003

AbstractBackground: GANP, carrying DNA‐primase and MCM3‐binding domains, is up‐regulated in germinal centre B cells. To understand the regulatory function of GANP upon MCM complex, we searched for GANP‐associated molecules by yeast two‐hybrid screening.Results: Using the 1 kb fragment (G5) of the ganp cDNA, we identified a clone named G5PR that is structurally homologous to known regulatory subunits of protein phosphatases (PPases) and determined the association of G5PR with GANP in vivo in the DNA transfectant. G5PR is associated with protein phosphatase 5 (PP5) through its tetratricopeptide‐repeat (TPR) domain. Pull‐down assays demonstrated that G5PR is also associated with protein phosphatase 2A (PP2A), the complex of A subunit (PR65) and the catalytic (C) subunit (PP2Ac), similar to the B′′ subunit. The G5PR‐associated complex had phosphatase activity on casein, histone H1 and MCM3 in vitro, but the addition of G5PR did not stimulate or inhibit the phosphatase activities of PP5 and PP2A. The cellular localization of G5PR in transfected cells varies during cell cycling, appearing in the nucleus during prophase, in the peri‐chromatin during mitotic phase, and in the cytoplasm after cell division.Conclusion: G5PR is capable of recruiting two kinds of PPases, PP5 and PP2A, into the GANP/MCM3 complex, which might regulate its phosphorylation state during cell cycle progression.
An algebraic formulation of geophysical inverse problems
Authors: Vasco D. W. 
Published: March 2003

Summary
Many geophysical inverse problems derive from governing partial differential equations with unknown coefficients. Alternatively, inverse problems often arise from integral equations associated with a Green's function solution to a governing differential equation. In their discrete form such equations reduce to systems of polynomial equations, known as algebraic equations. Using techniques from computational algebra one can address questions of the existence of solutions to such equations as well as the uniqueness of the solutions. The techniques are enumerative and exhaustive, requiring a finite number of computer operations. For example, calculating a bound to the total number of solutions reduces to computing the dimension of a linear vector space. The solution set itself may be constructed through the solution of an eigenvalue problem. The techniques are applied to a set of synthetic magnetotelluric values generated by conductivity variations within a layer. We find that the estimation of the conductivity and the electric field in the subsurface, based upon single-frequency magnetotelluric field values, is equivalent to a linear inverse problem. The techniques are also illustrated by an application to a magnetotelluric data set gathered at Battle Mountain, Nevada. Surface observations of the electric (E y) and magnetic (H x) fields are used to construct a model of subsurface electrical structure. Using techniques for algebraic equations it is shown that solutions exist, and that the set of solutions is finite. The total number of solutions is bounded above at 134 217 728. A numerical solution of the algebraic equations generates a conductivity structure in accordance with the current geological model for the area.
Comparison of six different methods for calculating traveltimes
Authors: Leidenfrost  , Ettrich  , Gajewski  , Kosloff  
Published: March 2003

We study six different methods for the calculation of seismic traveltimes. All methods yield traveltimes at all points of a regular grid.The methods examined comprise three different variants of finite‐difference (FD) eikonal solvers, the graph method, wavefront construction and a combined FD and Runge–Kutta method.The main points of investigation are computational time, accuracy and memory requirements. We took measures to obtain a high level of both general validity and clear understanding of the results. We used a profiling program to be able to measure the time that the actual core algorithm needs, thus avoiding any overhead of highly system‐dependent in‐/output operations.The comparison shows that no single method is the most appropriate but that the choice depends on the task to be fulfilled. The FD eikonal solver that uses expanding squares proves to be best suited for models which are not too complicated because it offers the best compromise between speed and accuracy, whereas wavefront construction should be applied to complex media because of its superior reliability which then justifies the much higher computational times.
Depth of detection of highly conducting and volume polarizable targets using induced polarization
Authors: Apparao A. , Srinivas G.S. , Subrahmanya Sarma V. , Thomas P.J. , Joshi M.S. , Rajendra Prasad P. 
Published: March 2003

AbstractWe define the apparent frequency effect in induced polarization (IP) as the relative difference between apparent resistivities measured using DC excitation on the one hand and high‐frequency excitation (when the IP effect vanishes) on the other. Assuming a given threshold for the minimum detectable anomaly in the apparent frequency effect, the depth of detection of a target by IP can be defined as that depth below which the target response is lower than the threshold for a given electrode array. Physical modelling shows that for the various arrays, the depth of detection of a highly conducting and volume polarizable target agrees closely with the depth of detection of an infinitely conducting and non‐polarized body of the same shape and size. The greatest depth of detection is obtained with a two‐electrode array, followed by a three‐electrode array, while the smallest depth of detection is obtained with a Wenner array when the array spread is in‐line (i.e. perpendicular to the strike direction). The depth of detection with a Wenner array improves considerably and is almost equal to that of a two‐electrode array when the array spread is broadside (i.e. along the strike direction).
Biophysical stratification of the Amazon basin
Authors: Prince Stephen D. , Steininger MarC. K. 
Published: March 2003

AbstractIn field measurement programmes, stratified sampling can optimize sampling efficiency, but stratification is often undertaken subjectively, and is frequently based on a priori classification schemes such as those used for vegetation maps. In order to avoid the problems associated with a priori subjective schemes, we explore here an objective procedure, Regression Tree Analysis (RTA). RTA has previously been used in local‐scale studies, but here we apply it to a very large study domain, namely the entire humid tropical zone of South America. The aim of the study was to develop an optimal sampling design in preparation for the Large Scale Biosphere‐Atmosphere Experiment in Amazonia (LBA). Co‐registered spatially continuous fields of rainfall, temperature, photosynthetically active radiation (PAR), the normalized difference index (NDVI), an index of surface moisture, and other independent variables were used to predict three dependent variables, annual net radiation (Rn), latent heat (LE) and net primary production (NPP). Rather than simply dividing the study area based on differing levels of the three dependent variables, empirical models were developed using RTA to indicate how the relationships between these and possible forcing variables vary across the study area. For each variable long‐term seasonal indices such as annual average, monthly minimum and amplitude were used to exclude effects of temporal phase differences between the hemispheres. The resulting hierarchical models revealed variations in the interdependence of the forcing variables throughout the study area and therefore provided a basis for a stratified sampling and identifying the most important variables to be collected in LBA for the Amazon basin as a whole as well as optimizing the sampling scheme for scaling up findings from the field scale to larger areas.