Kinase Gene (kinase + gene)

Distribution by Scientific Domains
Distribution within Life Sciences

Kinds of Kinase Gene

  • protein kinase gene
  • thymidine kinase gene

  • Selected Abstracts

    A Novel Mitogen-Activated Protein Kinase Gene in Maize (Zea mays), ZmMPK3, is Involved in Response to Diverse Environmental Cues

    Jinxiang Wang
    In search for components of mitogen-activated protein kinase (MAPK) cascades in maize (Zea mays) involved in response to abscisic acid (ABA) stimulus, a novel MAPK gene, ZmMPK3, from ABA-treated maize leaves cDNA was isolated and characterized. The full length of the ZmMPK3 gene is 1 520 bp and encodes a 376 amino acid protein with a predicted molecular mass of 43.5 kD and a pI of 5.83. ZmMPK3 contains all 11 MAPK conserved subdomains and the phosphorylation motif TEY. Amino acid sequence alignment revealed that ZmMPK3 shared high identity with group-A MAPK in plants. A time course (30,360 min) experiment using a variety of signal molecules and stresses revealed that the transcripts level of ZmMPK3 accumulated markedly and rapidly when maize seedlings were subjected to exogenous signaling molecules: ABA, H2O2, jasmonic acid and salicylic acid, various abiotic stimuli such as cold, drought, ultraviolet light, salinity, heavy metal and mechanical wounding. Its transcription was also found to be tissue-specific regulated. Here, we show that ABA and H2O2 induced a significant increase in the ZmMPK3 activity using immunoprecipitation and in-gel kinase assay. Furthermore, the results showed that the ZmMPK3 protein is localized mainly to the nucleus. These results suggest that the ZmMPK3 may play an important role in response to environmental stresses. [source]

    Methylation-Associated Silencing of Death-Associated Protein Kinase Gene in Laryngeal Squamous Cell Cancer,

    THE LARYNGOSCOPE, Issue 8 2005
    Wei-Jia Kong MD
    Abstract Objectives/Hypothesis: Death-associated protein kinase (DAPK) is a Ca2+/calmodulin-regulated Ser/Thr kinase that functions as a positive mediator of programmed cell death. It has been found that DAPK gene is frequently inactivated by its promoter hypermethylation in some cancers and tumor cell lines. However, it is not clear whether promoter hypermethylation of DAPK gene exists in laryngeal squamous cell cancer (LSCC). The aim of this study was to investigate the promoter methylation status of the DAPK gene in LSCC and the effect of 5-Aza-2'-deoxycytidine (5-Aza-CdR), a demethylating agent, on Hep-2 cells, a human laryngeal cancer cell line, and on xenografts of Hep-2. Methods: Methylation-specific polymerase chain reaction (PCR) and reverse-transcription PCR techniques were used to determine the promoter methylation status and mRNA expression of DAPK gene in LSCC. Furthermore, Hep-2 cells in vitro and in vivo were treated by 5-Aza-CdR to explore the effect of demethylating agents on DAPK mRNA expression and tumor growth. Results: Hypermethylation of DAPK gene promoter was found in 39 (67.2%) of 58 LSCC samples. There was no significant difference in the promoter hypermethylation rate among the samples of different histologic grades or samples from patients with different T stages. However, there was significant difference in methylation status of DAPK gene between the samples from patients in N0 stages and those from patients in N1 stages. No promoter hypermethylation of DAPK gene was found in any of the five normal laryngeal tissue samples. DAPK mRNA expression was not detected in tumor specimens with promoter hypermethylation. On the contrary, DAPK mRNA expression was observed in the unmethylated tumor specimens, specimens from tissues adjacent to the tumor, and normal laryngeal tissues samples. Promoter hypermethylation of DAPK gene was found, and no DAPK mRNA expression was detected in Hep-2 cells. DAPK mRNA expression in Hep-2 cells and xenografts could be restored by treating cells and xenografts with 5-Aza-CdR. The tumors' xenografts, induced by way of Hep-2 cell injection in nude mice treated with 5-Aza-CdR, were obviously smaller than those in nude mice treated with phosphate-buffered saline. Conclusions: Abnormal loss of DAPK expression could be associated with aberrant promoter region methylation in the LSCC. 5-Aza-CdR may slow the growth of Hep-2 cells in vitro and in vivo by reactivating tumor suppressor gene DAPK silenced by de novo methylation. [source]

    Cloning and Preliminary Characterization of Three Receptor-like Kinase Genes in Soybean

    Yuan-Yuan Ma
    Abstract Leaf senescence that occurs in the last stage of leaf development is a genetically programmed process. It is very significant to isolate the upstream components in the senescence signaling pathway and to elucidate the molecular mechanisms that control the initiation and progression of leaf senescence. In this study, full-length cDNAs of three receptor-like protein kinase genes, designated rlpk1, rlpk2 and rlpk3, were cloned from artificially-induced senescent soybean (Glycine max L.) primary leaves (GenBank accession AY687390, AY687391, AF338813). The deduced amino acid sequences indicated that they belonged to a receptor-like kinase family. Each of rlpk1 and rlpk2 encodes a leucine-rich repeat (LRR) receptor-like protein kinase. They both comprise a typical signal peptide, several LRR motifs, a single-pass transmem-brane domain, and a cytoplasmic protein kinase domain. No typical extracellular domain of RLPK3 was predicted. Organ-specific expression pattern analysis by reverse-transcription polymerase chain reaction (RT-PCR) revealed higher expression levels of the three genes in cotyledons, roots and flowers. Phylogenetic analysis indicated that RLPK1 and RLPK2 belonged to an independent branch, whereas RLPK3 shared common nodes with several known RLKs responding to abiotic and biotic stresses. The evident alternations of expression profiles of rlpk1 and rlpk2 induced by the artificial senescence-inducing treatment implied involvements of these two RLKs in regulating soybean leaf senescence. (Managing editor: Li-Hui Zhao) [source]

    Conditional ablation of neurones in transgenic mice

    Anthony R. Isles
    Abstract Conditional targeted ablation of specific cell populations in living transgenic animals is a very powerful strategy to determine cell functions in vivo. This approach would be of particular value to study the functions of distinct neuronal populations; however, the transgene of choice for conditional cell ablation studies in mice, the herpes simplex virus thymidine kinase gene, cannot be used to ablate neurones as its principal mode of action relies on cell proliferation. Here we report that expression of the E.coli nitroreductase gene (Ntr) and metabolism of the prodrug CB1954 (5-aziridin-1-yl-2-4-dinitrobenzamide) to its cytotoxic derivative can be used to conditionally and acutely ablate specific neuronal populations in vivo. As proof of principal, we have ablated olfactory and vomeronasal receptor neurones by expressing Ntr under the control of the olfactory marker protein (OMP) gene promoter. We demonstrate that following CB1954 administration, olfactory and vomeronasal receptor neurones expressing the transgene were selectively eliminated from the olfactory epithelium (OE), and projections to the olfactory bulb (OB) were lost. The functional efficacy of cell ablation was demonstrated using a highly sensitive behavioural test to show that ablated mice had lost the olfactory ability to discriminate distinct odors and were consequently rendered anosmic. Targeted expression of Ntr to specific neuronal populations using conventional transgenes, as described here, or by "knock-in" gene targeting using embryonic stem cells may be of significant value to address the functions of distinct neuronal populations in vivo. © 2001 John Wiley & Sons, Inc. J Neurobiol 47: 183,193, 2001 [source]

    Comparative analysis of MLL partial tandem duplication and FLT3 internal tandem duplication mutations in 956 adult patients with acute myeloid leukemia

    Christine Steudel
    Partial tandem duplication (PTD) of the MLL gene and internal tandem duplication (ITD) of the juxtamembrane region of the FLT3 receptor tyrosine kinase gene have been described in acute myeloid leukemia (AML) patients, preferentially in those with normal cytogenetics. These alterations have been associated with a poor prognosis. In our study, we analyzed the prevalence and the potential prognostic impact of these aberrations in a large unselected and well-defined cohort of 956 patients with AML. Results were correlated with cytogenetic data and clinical outcome. MLL PTD was detected by RT-PCR, subsequent nucleotide sequencing, and Southern blotting. The overall incidence was found to be 5.0% (48/956), whereas FLT3 ITD was detected in 19.2% (184/956). Sixteen cases were positive for both alterations. The rate of MLL PTD in FLT3 ITD positive patients was significantly higher than that in FLT3 ITD negative patients [16/184 (8.7%); 32/772 (4.1%); P = 0.025]. However, both aberrations were highly increased in patients with normal karyotype (MLL PTD 35/431, P = 0.004; FLT3 ITD 132/334, P < 0.001). When restricted to this subgroup, the rate of MLL PTD in patients with FLT3 mutations was not significantly increased. No statistically significant differences were detected between patients positive for MLL PTD and patients negative for MLL PTD in the rate of complete remissions or the overall survival, although we did see a significantly shorter disease-free survival in patients age 60 or younger. In conclusion, although there is an overlap in the mutational spectrum in AML with FLT3 ITD and MLL PTD mutations, our data do not support a common mechanistic basis. Although associated with inferior disease-free survival, the results of this study do not unequivocally support the notion that MLL PTD mutations represent an independent prognostic factor. © 2003 Wiley-Liss, Inc. [source]

    A variant of the myosin light chain kinase gene is associated with severe asthma in African Americans

    Carlos Flores
    Abstract Asthma is a complex phenotype influenced by environmental and genetic factors for which severe irreversible structural airway alterations are more frequently observed in African Americans. In addition to a multitude of factors contributing to its pathobiology, increased amounts of myosin light chain kinase (MLCK), the central regulator of cellular contraction, have been found in airway smooth muscle from asthmatics. The gene encoding MLCK (MYLK) is located in 3q21.1, a region noted by a number of genome-wide studies to show linkage with asthma and asthma-related phenotypes. We studied 17 MYLK genetic variants in European and African Americans with asthma and severe asthma and identified a single non-synonymous polymorphism (Pro147Ser) that was almost entirely restricted to African populations and which was associated with severe asthma in African Americans. These results remained highly significant after adjusting for proportions of ancestry estimated using 30 unlinked microsatellites (adjusted odds ratio: 1.76 [95% confidence interval, CI: 1.17,2.65], p = 0.005). Since all common HapMap polymorphisms in ,500,kb contiguous regions have low-to-moderate linkage disequilibrium with Pro147Ser, we speculate that this polymorphism is causally related to the severe asthma phenotype in African Americans. The association of this polymorphism, located in the N-terminal region of the non-muscle MLCK isoform, emphasizes the potential importance of the vascular endothelium, a tissue in which MLCK is centrally involved in multiple aspects of the inflammatory response, in the pathogenesis of severe asthma. This finding also offers a possible genetic explanation for some of the more severe asthma phenotype observed in African American asthmatics. Genet Epidemiol 2007. © 2007 Wiley-Liss, Inc. [source]

    The effect of focal adhesion kinase gene silencing on 5-fluorouracil chemosensitivity involves an Akt/NF-,B signaling pathway in colorectal carcinomas

    Yuying Chen
    Abstract Multicellular resistance (MCR) is produced because multicellular spheroids (MCSs) are formed with a broad cell,cell connection when cultured in three-dimensions, which limits the clinical treatment efficacy in solid tumors. Focal adhesion kinase (FAK) plays an important role in apoptosis, survival and cell adhesion between cells and their extracellular matrix. In this study, we investigated the expressions of FAK, Akt and NF-,B in human colorectal cancer (CRC), and the effects of FAK gene silencing on MCSs formation and 5-fluorouracil (5-FU) chemosensitivity in colon carcinoma MCSs culture cells. In CRC samples, FAK, Akt and NF-,B were overexpressed. The positive expression of FAK correlated notably with lymph node metastasis and cellular differentiation. Positive expressions of Akt and NF-,B were significantly related to cellular differentiation and lymph node metastasis, respectively. Furthermore, positive expression of FAK correlated with that of Akt and NF-,B. The expression of FAK was inhibited significantly by a small hairpin RNA targeting FAK. Knockdown of FAK reversed the formation and aggregation of MCSs, significantly decreased the 50% inhibitory concentration of 5-FU, and markedly increased MCS culture cells apoptosis. These effects were associated with reduced levels of Akt and NF-,B. These results indicate that suppressing FAK expression potentiated 5-FU-induced cytotoxicity and contributed to its chemosensitizing effect by suppressing Akt/NF-,B signaling in colon carcinoma MCS culture cells. These data also imply that FAK mediates MCR of CRC through the survival signaling pathway FAK/Akt/NF-,B. [source]

    HSPA1A is an important regulator of the stability and function of ZNF198 and its oncogenic derivative, ZNF198,FGFR1

    Chitta S. Kasyapa
    Abstract Mass spectroscopy analysis demonstrated that the HSPA1A protein is found in complex with the ZNF198 protein which is involved in a chromosome rearrangement with the FGFR1 gene in an atypical myeloproliferative disease. HSPA1A is a member of the HSP70 family of genes which has been shown to be inducible in a variety of circumstances. Exogenous expression of the ZNF198,FGFR1 fusion kinase gene as well as ZNF198 in a model cell system results in a large (>650-fold) increase in HSP70 mRNA levels. Using KNK437, a specific inhibitor of HSP70 transcription, we have demonstrated that an important function of HSPA1A is to stabilize the ZNF198 and ZNF198,FGFR1 proteins. In the absence of HSPA1A, specific functions of ZNF198,FGFR1 such as STAT3 phosphorylation is also lost. Treatment of cells with KNK437 in the presence of MG132, an inhibitor of proteasomal degradation of proteins, suggested that only the ZNF198,FGFR1 protein is subject to the proteasomal degradation pathway, while ZNF198 is not. These observations suggest an important role for HSPA1A in ZNF198 and ZNF198,FGFR1 mediated cellular function. J. Cell. Biochem. 102: 1308,1317, 2007. © 2007 Wiley-Liss, Inc. [source]

    Disruption of FRNK expression by gene targeting of the intronic promoter within the focal adhesion kinase gene

    Haruko Hayasaka
    Abstract FRNK, a non-catalytic variant of focal adhesion kinase (FAK), is expressed in major blood vessels throughout mouse development and is postulated to play a role in regulating cell adhesion and signaling in vascular smooth muscle cells (VSMCs). The FRNK transcriptional start site lies within an intron of the FAK gene, suggesting that the FRNK gene is a "gene within a gene". Here, we identified a 1 kb intronic sequence of the FAK gene that is necessary for endogenous FRNK expression. Deletion of this sequence in gene-targeted mice abolished FRNK expression, showing the direct involvement of the FAK intron in the regulation of FRNK expression. The level of FAK expression was normal in the FRNK-deficient mice, indicating that FAK and FRNK are transcriptionally regulated by distinct promoters. The FRNK-deficient mice were viable, fertile, and displayed no obvious histological abnormalities in any of the major blood vessels. Western blot analysis showed that FRNK,deficient and wild-type (WT) cells had comparable levels of steady-state and adhesion-dependent FAK autophosphorylation. Despite the fact that ectopic expression of FRNK suppresses focal adhesion formation in cultured cells, these results suggest that endogenous FRNK is not essential for development or the formation of the mouse vasculature. J. Cell. Biochem. 102: 947,954, 2007. © 2007 Wiley-Liss, Inc. [source]

    Water diffusion in a rat glioma during ganciclovir-thymidine kinase gene therapy-induced programmed cell death in vivo: Correlation with cell density

    Piia K. Valonen MSc
    Abstract Purpose To study the characteristics of diffusion magnetic resonance imaging (MRI) contrast in a rat brain BT4C glioma during progression of ganciclovir (GCV)-thymidine kinase gene therapy-induced programmed cell death (PCD) in vivo. Materials and Methods The trace of the diffusion tensor (Dav = 1/3Trace ), T2, and spin density were determined by MRI and the apparent diffusion coefficient (ADC) of water by diffusion nuclear MR (NMR) spectroscopy using largely varying b values and diffusion times (tD) at 4.7 T. Cell count and apoptotic cells were quantified by histological means. Results Decline in cell count was strongly associated with increase in both Dav and T2. Spin density ratio between tumor and contralateral parietal cortex increased with a very similar time course as Dav and T2, indicating net water gain into the eradicating tumor. Diffusion spectroscopy showed a nonmonoexponential signal decay at all tD values ranging from 14,192 msec. During PCD, the ADC of the component yielding fast diffusion coefficient (D1), as acquired with tD , 47 msec, increased with kinetics similar to those of Dav (tD = 4.8 msec). The fractional size of D1 increased by 10% to 15% throughout the entire tD range. Apparent water residence time of the slow diffusion component, D2, shortened from a value of 38.3 ± 1.7 msec on day 0 to 33.4 ± 0.5 msec by day 8. Conclusion The present results show that reduced cell density and increased water content, leading to altered water microenvironment, are associated with increased water diffusion coefficient in eradicating gliomas as a result of PCD. J. Magn. Reson. Imaging 2004;19:389,396. © 2004 Wiley-Liss, Inc. [source]

    Hyposialylation of neprilysin possibly affects its expression and enzymatic activity in hereditary inclusion-body myopathy muscle

    Aldobrando Broccolini
    Abstract Autosomal recessive hereditary inclusion-body myopathy (h-IBM) is caused by mutations of the UDP- N -acetylglucosamine 2-epimerase/N -acetylmannosamine kinase gene, a rate-limiting enzyme in the sialic acid metabolic pathway. Previous studies have demonstrated an abnormal sialylation of glycoproteins in h-IBM. h-IBM muscle shows the abnormal accumulation of proteins including amyloid-, (A,). Neprilysin (NEP), a metallopeptidase that cleaves A,, is characterized by the presence of several N-glycosylation sites, and changes in these sugar moieties affect its stability and enzymatic activity. In the present study, we found that NEP is hyposialylated and its expression and enzymatic activity reduced in all h-IBM muscles analyzed. In vitro, the experimental removal of sialic acid by Vibrio Cholerae neuraminidase in cultured myotubes resulted in reduced expression of NEP. This was most likely because of a post-translational modification consisting in an abnormal sialylation of the protein that leads to its reduced stability. Moreover, treatment with Vibrio Cholerae neuraminidase was associated with an increased immunoreactivity for A, mainly in the form of distinct cytoplasmic foci within myotubes. We hypothesize that, in h-IBM muscle, hyposialylated NEP has a role in hampering the cellular A, clearing system, thus contributing to its abnormal accumulation within vulnerable fibers and possibly promoting muscle degeneration. [source]

    Positive regulation of Bacillus subtilis ackA by CodY and CcpA: establishing a potential hierarchy in carbon flow

    Robert P. Shivers
    Summary Conversion of pyruvate to acetate via the phosphotransacetylase-acetate kinase pathway generates ATP and is a major overflow pathway under conditions of carbon and nitrogen excess. In Bacillus subtilis, this pathway is positively regulated by CcpA, a global regulator of carbon metabolism genes. Transcription of the acetate kinase gene (ackA) proved to be activated as well by a second global regulatory protein, CodY. Expression of an ackA,lacZ fusion was reduced in a codY mutant strain. CodY was found to bind in vitro to two sites in the ackA promoter region and to stimulate ackA transcription in a run-off transcription assay. This is the first known case of direct positive regulation by CodY. CodY and CcpA were found to bind to neighbouring sites and their effects were additive both in vivo and in vitro. Surprisingly, positive regulation by CodY, unlike repression, responded primarily to only one type of effector molecule. That is, branched-chain amino acids (BCAAs) served as more potent co-activators of CodY-dependent ackA transcription than did GTP. Given the roles of CcpA and CodY in regulating genes whose products determine the metabolic fate of pyruvate, these two proteins may act together to mediate a hierarchical conversion of pyruvate to its many potential products. [source]

    CPMK2, an SLT2-homologous mitogen-activated protein (MAP) kinase, is essential for pathogenesis of Claviceps purpurea on rye: evidence for a second conserved pathogenesis-related MAP kinase cascade in phytopathogenic fungi

    Géraldine Mey
    Summary Cpmk2 , encoding a mitogen-activated protein (MAP) kinase from the ascomycete Claviceps purpurea , is an orthologue of SLT2 from Saccharomyces cerevisiae , the first isolated from a biotrophic, non-appressorium-forming pathogen. Deletion mutants obtained by a gene replacement approach show impaired vegetative properties (no conidiation) and a significantly reduced virulence, although they retain a limited ability to colonize the host tissue. Increased sensitivity to protoplasting enzymes indicates that the cell wall structure of the mutants may be altered. As the phenotypes of these mutants are similar to those observed in strains of the rice pathogen, Magnaporthe grisea , that have been deprived of their MAP kinase gene mps1 , the ability of cpmk2 to complement the defects of , mps1 was investigated. Interestingly, the C. purpurea gene, under the control of its own promoter, was able to complement the M. grisea mutant phenotype: transformants were able to sporulate and form infection hyphae on onion epidermis and were fully pathogenic on barley leaves. This indicates that, despite the differences in infection strategies, which include host and organ specificity, mode of penetration and colonization of host tissue, CPMK2 / MPS1 defines a second MAP kinase cascade (after the Fus3p/PMK1 cascade) essential for fungal pathogenicity. [source]

    Role of two component signaling response regulators in acid tolerance of Streptococcus mutans

    M. Kawada-Matsuo
    Introduction:, In bacteria, two-component systems (TCS) involving the products of a histidine kinase gene (hk) and a response regulator gene (rr) play important roles in adaptation to environmental changes. Fourteen hk - rr homologs and one orphan rr homolog were identified in the Streptococcus mutans UA159 genome database. There have been no comprehensive evaluations of the roles of rr homologs in the acid tolerance of S. mutans. Methods:, The TCS genes (tcs) of S. mutans were designated smtcs01,15. Mutants of S. mutans UA159 with deletions of rr and hk-rr were constructed. Acid tolerance was evaluated by comparing the doubling times at pH 7.2 and pH 5.5 between the wild-type and mutant strains. Results:, Excluding smtcs10 and 12, for which viable mutants could not be obtained, a total of 13 rr deletion mutants were constructed. The rr deletions in smtcs03, 05, 08, and 13 resulted in diminished acid tolerance in comparison with UA159. The hk-rr double-mutants exhibited acid sensitivity levels similar to those of the corresponding rr mutants. The results of the present study reveal the involvement of the rr genes of smtcs03 and 05 in acid tolerance. Deletion of hk and/or rr in smtcs03 generated an acid-sensitive phenotype. In contrast, for smtcs05, while deletion of rr resulted in reduced acid tolerance, a single-deletion of hk had no effect on acid tolerance. Conclusions:, We implicated two rr genes in the acid tolerance of S. mutans. In particular, smtcs05 is a novel tcs, the sole rr of which is involved in the acid tolerance of S. mutans. [source]

    The cell-cycle promoter cdc2aAt from Arabidopsis thaliana is induced in the lateral roots of the actinorhizal tree Allocasuarina verticillata during the early stages of the symbiotic interaction with Frankia

    Mame Ourèye Sy
    The symbiosis between the actinorhizal tree Allocasuarina verticillata and the actinomycete Frankia leads to the formation of root nodules inside which bacteria fix atmospheric nitrogen. Actinorhizal nodule organogenesis starts with the induction of cell divisions in the root cortex and in the pericycle cells opposite protoxylem poles near Frankia -infected root hairs. To study the ability of Frankia to induce progression through the cell cycle, we monitored the expression of the ,-glucuronidase (gus) gene driven by the promoter from cdc2aAt, an Arabidopsis cyclin-dependent kinase gene that displays competence for cell division, during plant growth and nodule ontogenesis. In non-symbiotic tissues, the gus gene was mainly expressed in primary and secondary meristems of roots and shoots. Auxins and cytokinins were found to induce reporter gene activity in the root system of whole plants, showing that the promoter cdc2aAt displayed the same regulation by hormones in Allocasuarina as that reported in Arabidopsis. In transgenic nodules, gus expression was found to be restricted to the phellogen. During the early stages of the interaction between Frankia and the plant root system, cdc2aAt was strongly induced in the lateral roots surrounded by hyphae of the actinomycete. Histochemical analysis of ,-glucuronidase activity revealed that cells from the pericycle opposite protoxylem poles were very deeply stained. These data indicate that upon Frankia infection, cells from the lateral roots, and notably pericycle cells that can give rise to a nodule or a root primordium, prepare to re-enter the cell cycle. [source]

    Expression of Oryza sativa MAP kinase gene is developmentally regulated and stress-responsive

    Hao-Jen Huang
    Mitogen-activated protein kinase (MAPK) pathways are modules involved in the transduction of extracellular signals to intracellular targets in all eukaryotes. In plants, there is evidence for MAPKs playing a role in the signalling of abiotic stresses, pathogens, plant hormones, and cell cycle cues. The large number and divergence of plant MAPKs indicates that this ancient mechanism of signal transduction is extensively used in plants. However, there have been no reports of classical MAPK module in rice. In this report, we have isolated a MAPK from rice (Oryza sativa) termed OsMAPK2. The cloned cDNA is 1457 nucleotides long and the deduced amino acid sequence comprised 369 amino acid residues. Sequence analysis revealed that the predicted amino acid sequence is 72% identical to tobacco wound-induced protein kinase (WIPK). Southern analysis suggested a single OsMAPK2 gene in rice. Analysis at the mRNA level has shown that OsMAPK2 is expressed in all plant organs and high relative amounts of OsMAPK2 were detected in the mature panicles in comparison with in the immature panicles. In suspension-cultured cells, the OsMAPK2 mRNA transcript increased markedly upon temperature downshift from 26°C to 4°C and sucrose starvation. In contrast, the OsMAPK2 mRNA level rapidly declined in rice cell challenged by high temperature. A similarly rapid response of OsMAPK2 was observed in stress-treated seedlings, demonstrating that response of the MAPK pathway occurs also in intact plants. These results suggest that this OsMAPK2 may function in the stress-signalling pathway as well as panicle development in rice. [source]

    Cytotoxicity and antiangiogenesis by fibroblast growth factor 2,targeted Ad-TK cancer gene therapy,

    THE LARYNGOSCOPE, Issue 4 2009
    Koichiro Saito MD
    Abstract Objectives: Human head and neck squamous cell carcinoma (HNSCC) in addition to lung, skin, ovarian, and other cancers overexpress fibroblast growth factor (FGF) receptors on both individual tumor cells and endothelial cells within the tumor microenvironment. The purpose of this study was to investigate whether FGF2-targeted gene therapy could redirect adenoviral vectors encoding the herpes simplex virus thymidine kinase gene (Ad-TK) to FGF receptors on tumor and endothelial cells with the intent of improving both the efficiency of transgene expression and the antitumor response. Study Design and Methods: An Ad-TK vector consisting of a conjugate of FGF2 linked to a Fab, fragment against the adenoviral knob region was directly delivered to human HNSCC xenograft tumors in nude mice, which were subsequently dosed with ganciclovir. Tumor specimens were assessed for herpes simplex virus thymidine kinase (HSV- tk) transgene mRNA expression, FGF1/2 receptor expression, terminal deoxynucleotidyl transferase biotin,deoxy uridine triphosphate nick end labeling assay for apoptosis, CD31 immunohistochemistry to estimate tumor microvessel density, and tumor volume change. Results: FGF2-retargeted Ad-TK gene therapy demonstrated significant increases in both HSV- tk mRNA expression and cellular apoptosis levels, and a significant decrease in tumor volume size compared with all other groups. Furthermore, microvessel density was significantly lower in the FGF2-retargeted Ad-TK group, indicating a strong antiangiogenesis effect. Conclusions: These data suggest that FGF2-retargeted Ad-TK produces a combination of expected direct antitumor cytotoxicity and a newly reported antiangiogenesis effect that could prove promising as a novel therapeutic approach in the treatment of FGF receptor,expressing cancers. Laryngoscope, 2009 [source]

    Ethylene-induced hyponastic growth in Arabidopsis thaliana is controlled by ERECTA

    THE PLANT JOURNAL, Issue 1 2010
    Martijn Van Zanten
    Summary Plants can respond quickly and profoundly to detrimental changes in their environment. For example, Arabidopsis thaliana can induce an upward leaf movement response through differential petiole growth (hyponastic growth) to outgrow complete submergence. This response is induced by accumulation of the phytohormone ethylene in the plant. Currently, only limited information is available on how this response is molecularly controlled. In this study, we utilized quantitative trait loci (QTL) analysis of natural genetic variation among Arabidopsis accessions to isolate novel factors controlling constitutive petiole angles and ethylene-induced hyponastic growth. Analysis of mutants in various backgrounds and complementation analysis of naturally occurring mutant accessions provided evidence that the leucin-rich repeat receptor-like Ser/Thr kinase gene, ERECTA, controls ethylene-induced hyponastic growth. Moreover, ERECTA controls leaf positioning in the absence of ethylene treatment. Our data demonstrate that this is not due to altered ethylene production or sensitivity. [source]

    Acyclovir resistance in herpes simplex encephalitis

    ANNALS OF NEUROLOGY, Issue 6 2010
    Eva C. Schulte MD
    Herpes simplex virus type 1 is a common cause of severe sporadic encephalitis. Treatment with acyclovir is highly effective in this disease. We report the case of a 27-year-old, immunocompetent woman with acyclovir-resistant herpes simplex encephalitis. Although she had not been treated before, herpes simplex virus type 1 DNA from the cerebrospinal fluid showed a non-synonymous mutation in the thymidine kinase gene, which is likely to have caused resistance to acyclovir. Herpes simplex encephalitis resolved after treatment with foscarnet. To our knowledge, this is the first report of acyclovir-resistant herpes simplex virus encephalitis in an immunocompetent, previously therapy-naive adult. ANN NEUROL 2010;67:830,833 [source]

    Replication of the association between the C8orf13,BLK region and systemic lupus erythematosus in a Japanese population

    ARTHRITIS & RHEUMATISM, Issue 2 2009
    Ikue Ito
    Objective Recent genome-wide association studies identified an association between single-nucleotide polymorphisms (SNPs) in the C8orf13 region of BLK, the B lymphoid tyrosine kinase gene, with systemic lupus erythematosus (SLE) in Caucasians. The purpose of this study was to evaluate the significance of this region in the genetic background of Japanese patients with SLE. Methods Fourteen tag SNPs in the C8orf13,BLK region were genotyped in 327 Japanese patients with SLE and 322 healthy Japanese controls. The population-attributable risk percentage (PAR%) of rs13277113 in Japanese was compared with that in Caucasians as well as with that of other SLE susceptibility genes in Japanese. Results As in Caucasians, rs13277113A demonstrated the strongest association in Japanese (P = 1.73 × 10,6 for the genotype frequency, P = 4.75 × 10,7 for the allele frequency, odds ratio [OR] 2.44 [95% confidence interval (95% CI) 1.43,4.16]). The association in Japanese was consistent with a recessive model (P = 2.74 × 10,7, OR 2.27 [95% CI 1.66,3.11]). In contrast to the Caucasian population, this risk allele was the major allele in the Japanese population. Because both the risk allele frequency and the OR were higher in Japanese than in Caucasians, the PAR% of rs13277113 was estimated to be much higher in Japanese (35.4%) than in Caucasians (16.2%), and the second highest among the 6 confirmed SLE susceptibility genes in Japanese. Conclusion The association of the C8orf13,BLK region with SLE was replicated in a Japanese population. Contribution of this region to the genetic predisposition to SLE appeared to be greater in Japanese than in Caucasians. [source]

    Engineering Propionibacterium acidipropionici for enhanced propionic acid tolerance and fermentation

    An Zhang
    Abstract Propionibacterium acidipropionici, a Gram-positive, anaerobic bacterium, has been the most used species for propionic acid production from sugars. In this study, the metabolically engineered mutant ACK-Tet, which has its acetate kinase gene knocked out from the chromosome, was immobilized and adapted in a fibrous bed bioreactor (FBB) to increase its acid tolerance and ability to produce propionic acid at a high final concentration in fed-batch fermentation. After about 3 months adaptation in the FBB, the propionic acid concentration in the fermentation broth reached ,100,g/L, which was much higher than the highest concentration of ,71,g/L previously attained with the wild-type in the FBB. To understand the mechanism and factors contributing to the enhanced acid tolerance, adapted mutant cells were harvested from the FBB and characterized for their morphology, growth inhibition by propionic acid, protein expression profiles as observed in SDS,PAGE, and H+ -ATPase activity, which is related to the proton pumping and cell's ability to control its intracellular pH gradient. The adapted mutant obtained from the FBB showed significantly reduced growth sensitivity to propionic acid inhibition, increased H+ -ATPase expression and activity, and significantly elongated rod morphology. Biotechnol. Bioeng. 2009; 104: 766,773 © 2009 Wiley Periodicals, Inc. [source]

    The impact of retroviral suicide gene transduction procedures on T cells

    Waseem Qasim
    Summary., Retroviral vectors encoding the herpes simplex thymidine kinase gene have been used to render T cells sensitive to the prodrug ganciclovir. Such genetically modified T cells have been used in clinical trials for their graft-versus-leukaemia effects following allogeneic haematopoietic stem cell transplantation. In the event of graft-versus-host disease (GVHD) the cells were susceptible to elimination through exposure to ganciclovir. We have investigated the impact of T-cell activation, required for successful retrovirus-mediated gene transfer, on T-cell receptor repertoire profile, subset distribution and antiviral potential. Using a combination of antibodies against CD3 and CD28, T cells were transduced at high efficiency when exposed to retrovirus between 48 and 72 h later. Lymphocytes had undergone up to seven cycles of cell division by the end of the procedure. Although the T-cell receptor V, repertoire was not altered after retroviral transduction, there were notable shifts in subset profiles with an increased proportion of CD45RO cells in transduced populations. T cells continued to proliferate for several days after transduction and were difficult to sustain under the extended culture conditions required to generate virus-specific T cells. These observations may explain the lower than expected levels of GVHD and poor antiviral immunity reported in recent trials. [source]

    CTG repeats at the myotonic protein kinase gene in a healthy Chilean population sample

    F. Amenabar
    Objectives,,, To study the variability at the myotonic dystrophy protein kinase (DMPK) gene in a Chilean sample of healthy people. DM1 is an autosomal dominant disorder caused by an expansion of a (CTG) repeat at the 3,-UTR of the gene DMPK. Healthy individuals have alleles under 35 repeats and diseased individuals have over 50. Methods,,, Genotyping the number of (CTG) repeats at this gene in a sample of healthy Chilean people. Results,,, Allele frequencies were significantly different from those of other populations. The most frequent allele was with five repeats. The frequency of larger alleles (>18 CTG repeats) was 11%, close to the European frequency (12%) and higher than the Japanese (8%) and Aboriginal Pehuenche samples (8%). Conclusions,,, Allelic frequencies in the Chilean sample studied were intermediate between those of the two ancestral populations (European and Pehuenche). [source]

    Deletion Mutants of Human Deoxycytidine Kinase mRNA in Cells Resistant to Antitumor Cytosine Nucleosides

    CANCER SCIENCE, Issue 7 2001
    Tohru Obata
    We studied mutational events in deoxycytidine (dCyd) kinase mRNA expression, focusing on aberrant dCyd kinase mRNA, which has been frequently observed in established cell lines resistant to antitumor dCyd nucleoside analogues such as 1-,-D-arabinofuranosyl cytosine (Ara-C), gemcita-bine (dFdC) and 2,-C-cyano-2,-deoxy-l-,-D-arabinofuranosylcytosine (CNDAC). We describe here the expression of aberrant dCyd kinase mRNAs identified as splicing mutants. These mutants included deletions of the fifth exon in CNDAC-resistant cells (originating from HT-1080 cells), of the third exon in Ara-C-resistant cells (originating from SK-MEL-28 cells) and of the fourth exon in 2,-deoxy-2,-methylidenecytidine (DMDC)-resistant cells (originating from SK-MEL-28 cells). Various nucleoside-resistant cells originating from the same parental HT-1080 cells were established. The resulting cells expressed the same mRNA with deletion of the fifth exon, and the location of splicing was independent of the type of nucleosides used for the establishment of resistant cells. The deletion of the fifth exon in dCyd kinase seems to be a target for acquisition of resistance to antitumor cytosine nucleosides. However, distinct mutations in the dCyd kinase gene seem to be associated with acquisition of resistance to different antitumor cytosine nucleosides. [source]

    Cloning and Preliminary Characterization of Three Receptor-like Kinase Genes in Soybean

    Yuan-Yuan Ma
    Abstract Leaf senescence that occurs in the last stage of leaf development is a genetically programmed process. It is very significant to isolate the upstream components in the senescence signaling pathway and to elucidate the molecular mechanisms that control the initiation and progression of leaf senescence. In this study, full-length cDNAs of three receptor-like protein kinase genes, designated rlpk1, rlpk2 and rlpk3, were cloned from artificially-induced senescent soybean (Glycine max L.) primary leaves (GenBank accession AY687390, AY687391, AF338813). The deduced amino acid sequences indicated that they belonged to a receptor-like kinase family. Each of rlpk1 and rlpk2 encodes a leucine-rich repeat (LRR) receptor-like protein kinase. They both comprise a typical signal peptide, several LRR motifs, a single-pass transmem-brane domain, and a cytoplasmic protein kinase domain. No typical extracellular domain of RLPK3 was predicted. Organ-specific expression pattern analysis by reverse-transcription polymerase chain reaction (RT-PCR) revealed higher expression levels of the three genes in cotyledons, roots and flowers. Phylogenetic analysis indicated that RLPK1 and RLPK2 belonged to an independent branch, whereas RLPK3 shared common nodes with several known RLKs responding to abiotic and biotic stresses. The evident alternations of expression profiles of rlpk1 and rlpk2 induced by the artificial senescence-inducing treatment implied involvements of these two RLKs in regulating soybean leaf senescence. (Managing editor: Li-Hui Zhao) [source]

    DspA/E, a type III effector of Erwinia amylovora, is required for early rapid growth in Nicotiana benthamiana and causes NbSGT1-dependent cell death

    SUMMARY DspA/E is a pathogenicity factor of Erwinia amylovora that is translocated into the plant cell cytoplasm through an Hrp type III secretion system. Transient expression of dspA/E in Nicotiana benthamiana or yeast induced cell death, as it does in N. tabacum and apple as described previously. DspA/E-induced cell death in N. benthamiana was not inhibited by coexpression of AvrPtoB of Pseudomonas syringae pv. tomato, which inhibits programmed cell death (PCD) induced by several other elicitors in plants. Silencing of NbSGT1, the expression of which is required for PCD mediated by several resistance proteins of plants, prevented DspA/E-induced cell death in N. benthamiana. However, silencing of NbRAR1, or two MAP kinase kinase genes, which are required for PCD associated with many resistance genes in plants, did not prevent cell death induced by DspA/E. Silencing of NbSGT1 also compromised non-host resistance against E. amylovora. E. amylovora grew rapidly within the first 24 h after infiltration in N. benthamiana, and DspA/E was required for this early rapid growth. However, bacterial cell numbers decreased after 24 h in TRV-vector-transformed plants, whereas a dspA/E mutant strain grew to high populations in NbSGT1 -silenced plants. Our results indicate that DspA/E enhances virulence of E. amylovora in N. benthamiana, but the bacteria are then recognized by the plant, resulting in PCD and death of bacterial cells or restriction of bacterial cell growth. [source]

    pH and carbon supply control the expression of phosphoenolpyruvate carboxylase kinase genes in Arabidopsis thaliana

    PLANT CELL & ENVIRONMENT, Issue 12 2008
    ABSTRACT Phosphoenolpyruvate carboxylase (PEPC) is thought to play many roles in C3 plants including the provision of biosynthetic precursors and control of pH during N assimilation. Its activity is controlled via phosphorylation catalysed by PEPC kinases, which are encoded by PPCK genes. We examined PPCK expression in response to changes in the supply of N or C, and to changes in intracellular pH, using cultured Arabidopsis cells and seedlings. The results show that expression of both PPCK1 and PPCK2 is increased by C availability, but does not respond to N availability. Expression of the two PPCK genes and the phosphorylation state of PEPC are increased in response to increasing intracellular pH. Elevated pH also reduces the repression of PPCK gene expression by Pi. Expression of phosphoenolpyruvate carboxykinase (PEPCK), which catalyses the decarboxylation of oxaloacetate, is decreased in response to increasing intracellular pH. pH homeostasis may be mediated at least partly by reciprocal changes in the expression of PPCK genes and PEPCK. [source]

    Cloning of deoxynucleoside monophosphate kinase genes and biosynthesis of deoxynucleoside diphosphates

    Jie Bao
    Abstract The genes encoding four deoxynucleoside monophosphate kinase (dNMP kinase) enzymes, including ADK1 for deoxyadenylate monophosphate kinase (AK), GUK1 for deoxyguanylate monophosphate kinase (GK), URA6 for deoxycytidylate monophosphate kinase (CK), and CDC8 for deoxythymidylate monophosphate kinase (TK), were isolated from the genome of Saccharomyces cerevisiae ATCC 2610 strain and cloned into E. coli strain BL21(DE3). Four recombinant plasmids, pET17b-JB1 containing ADK1, pET17b-JB2 containing GUK1, pET17b-JB3 containing URA6, and pET17b-JB4 containing CDC8, were constructed and transformed into E. coli strain for over-expression of AK, GK, CK, and TK. The amino acid sequences of these enzymes were analyzed and a putative conserved peptide sequence for the ATP active site was proposed. The four deoxynucleoside diphosphates (dNDP) including deoxyadenosine diphosphate (dADP), deoxyguanosine diphosphate (dGDP), deoxycytidine diphosphate (dCDP), and deoxythymidine diphosphate (dTDP), were synthesized from the corresponding deoxynucleoside monophosphates (dNMP) using the purified AK, GK, CK, and TK, respectively. The effects of pH and magnesium ion concentration on the dNDP biosynthesis were found to be important. A kinetic model for the synthetic reactions of dNDP was developed based on the Bi,Bi random rapid equilibrium mechanism. The kinetic parameters including the maximum reaction velocity and Michaelis,Menten constants were experimentally determined. The study on dNDP biosynthesis reported in this article are important to the proposed bioprocess for production of deoxynucleoside triphosphates (dNTP) that are used as precursors for in vitro DNA synthesis. There is a significant advantage of using enzymatic biosyntheses of dNDP as compared to the chemical method that has been in commercial use. © 2005 Wiley Periodicals, inc. [source]