• Title/Summary/Keyword: negative mutants

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Characterization of cadC and cadR Mutants in Mediating the Expression of the Salmonella typhimurium cadBA Operon (Salmonella typhimurium cadBA 오페론의 발현에 관여하는 돌연변이체의 선별 및 그 특성)

  • 방성호;박용근
    • Korean Journal of Microbiology
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    • v.37 no.4
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    • pp.259-264
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    • 2001
  • It has been well known that the expression of S. typhimurium cadBA operon requires at least two extracellular signals: low pH and high concentration of lysine. To better understand the nature of pH-dependent and lysine dependent signal transduction, mutants were isolated in JF2238(cadA-lacZ) by Tn10 insertion, spontaneous mutagenesis, and EMS treatment. Two mutants were isolated from JF2238, expressed as a cadA-lacZ operon fusion in various growth conditions, and analyzed to have mutations in cadC, a gene encoding a function necessary for transcriptional activation of cadBA. One isolate (cadC6) conferred pH-independent and lysine-independent cadBA expression and the other(cadC4) showed pH-independent and lysine-dependent cadBA expression. cadR::Tn10 and cadR4 mutants were expressed in the absence of exogenously added lysine. They were also resistant to thiosine and complemented by lysP clone from E. coli. Thus, in the absence of exogenous lysine, cadR is a negative regulator of cadBA expression. Cadaverine, the product of lysine decarboxylation, was shown to inhibit expression of cadA-lacZ fusion in cad $C^+$ cell.

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Characterization of a Novel DWD Protein that Participates in Heat Stress Response in Arabidopsis

  • Kim, Soon-Hee;Lee, Joon-Hyun;Seo, Kyoung-In;Ryu, Boyeong;Sung, Yongju;Chung, Taijoon;Deng, Xing Wang;Lee, Jae-Hoon
    • Molecules and Cells
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    • v.37 no.11
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    • pp.833-840
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    • 2014
  • Cullin4-RING ubiquitin ligase (CRL4) is a family of multi-subunit E3 ligases. To investigate the possible involvement of CRL4 in heat stress response, we screened T-DNA insertion mutants of putative CRL4 substrate receptors that exhibited altered patterns in response to heat stress. One of the mutants exhibited heat stress tolerance and was named heat stress tolerant DWD1 (htd1). Introduction of HTD1 gene into htd1-1 led to recovery of heat sensitivity to the wild type level, confirming that the decrease of HTD1 transcripts resulted in heat tolerance. Therefore, HTD1 plays a negative role in thermotolerance in Arabidopsis. Additionally, HTD1 directly interacted with DDB1a in yeast two-hybrid assays and associated with DDB1b in vivo, supporting that it could be a part of a CRL4 complex. Various heat-inducible genes such as HSP14.7, HSP21, At2g03020 and WRKY28 were hyper-induced in htd1-1, indicating that HTD1 could function as a negative regulator for the expression of such genes and that these genes might contribute to thermotolerance of htd1-1, at least in part. HTD1 was associated with HSP90-1, a crucial regulator of thermotolerance, in vivo, even though the decrease of HTD1 did not affect the accumulation pattern of HSP90-1 in Arabidopsis. These findings indicate that a negative role of HTD1 in thermotolerance might be achieved through its association with HSP90-1, possibly by disturbing the action of HSP90-1, not by the degradation of HSP90-1. This study will serve as an important step toward understanding of the functional connection between CRL4-mediated processes and plant heat stress signaling.

Use of ALLGIO Probe Assays for Detection of HBV Resistance to Adefovir in Patients with Chronic Hepatitis B, Kerman, Iran

  • Afshar, Reza Malekpour;Mollaie, Hamid Reza
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.11
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    • pp.5463-5467
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    • 2012
  • Hepatitis B virus (HBV) infection is contagious with transmissiobn vertically or horizontally by blood products and body secretions. Over 50% of Iranian carriers contracted the infection prenatally, making this the most likely route of transmission of HBV in Iran. To evaluate the resistance to adefovir (ADV) therapy in patients with chronic hepatitis B infection, a study was conducted on 70 patients (63 males and 7 females), who had received in first line lamivudine and second line adefovir. All were tested for the presence of hepatitis B surface antigen (HBsAg), hepatitis B envelope antigen (HBeAg), serum alanine amino transferase (ALT) level and HBV DNA load before and after treatment with ADV. In all samples, resistance to lamivudine and ADV was tested with real time PCR. Among seventy patients with chronic hepatitis B infection, 18 (25.7%) were resistant to LAM and 8 (11.4%) were resistant to ADV. Only one patient was negative for the presence of HBS-Ag (5.6%) and two were negative for HBe-Ag (11.1%). In this study we used a new method (ALLGIO probe assay) that has high sensitivity in detection of adefovir resistance mutants, which we recommend to other researchers. Mutant strains of the YMDD motif of HBV polymerase can be found in some patients under treatment with lamivudine and ADV. ADV has been demonstrated to be efficient in patients with lamivudine resistant HBV.

Molecular characterization of a novel rice(Oryza sativa L.) MAP kinase, OsEDRl, its role in defense signaling pathway.

  • Kim, Jung-A;Jwa, Nam-Soo
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.82-83
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    • 2003
  • Plants have evolved differently from animals having mobile activities. Thus, plants should have developed unique defense mechanisms against biotic/abiotic stresses to which plants are differently exposed, according to seasons. Most organisms have an conserved signaling network using mitogen-activated protein kinase (MAPK) cascade(s). The phenomenon implied that they are functionally very important in all organisms. In fact, they constitute one of the major components of signaling pathways involved in regulating a wide range of cellular activities from growth and development to cell death. Recently, complete MAPK cascade was first characterized in Arabidopsis from the receptor kinase (FLS2) through fellowing MEKKI -MKK4/MKK5-MPK3/MPK6-WRKY22/MRKY29 pathway. Whereas, MAPK cascade signaling pathway in monocot plant including rice (0ryza sativa L.), the most important of all food crops and an established monocot plant research model, MAPKinase kinase kinases (MAPKKK) of rice are the first upstream component of the MAPK cascade, but MAPKKK has been first identified and characterized in our lab and designated as, OsEDRl based on its homology with the Arabidopsis EDRI. The Arabidopsis EDRl was regarded as a negative regulator of defense response and the role of rice OsEDRl was analyzed. Transcriptional regulation of OsEDRl was detected under various stresses and immunoblotting analysis is going on to detect the level of OsEDRl protein in the mutants showing unique phenotype. We also introduced the constitutively active and the dominant negative forms of the OsEDRl for characterizing biological function.

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INVITED REVIEW - PHOTOTROPISM OF PHYCOMYCES SPORANGIOPHORES

  • Ootaki, Tamotsu
    • Journal of Photoscience
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    • v.1 no.1
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    • pp.69-73
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    • 1994
  • Sporangiophores (spphs) of Phycomyces blakesleeanus are positively phototropic to unilateral visible (blue) light over a range of fluence rates 10$^{-9}$ to 1 W/m$^2$. The maximal bending angle is always 70-75$\circ$ from the vertical. Many mutants with abnormal phototropism have been isolated. Complementation tests revealed that the genetic grouping is completely consistent with the phenotypic classification scheme, based on sensory responses other than those to light. The spph of the piloboloid mutant, the growth zone of which gradually ceases elongation but expands spherically, and the $\beta$-carotene-overproducing mutant show negative phototropism, in contrast to the wild type spph. We hypothesized that the phototropic orientation of spph is determined by the ratio of the maximal light fluenee rate at the proximal side to that at the distal side of the spph. Based on this hypothesis, we found that the maximal bending angle was larger in thin spphs than in thick ones, and larger in spphs containing smaller amount of $\beta$-carotene than in carotene-rich spphs. In addition to our hypothesis, gravitropic experiments revealed that the maximal bending angle of the wild type spph results from a balance among positive phototropism, negative gravitropism, and the optical properties of the spph. For further advancement of this study, we developed a mutant with a high proportion of uninucleate spores, and designed an efficient microinjection method for obtaining transformants.

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Detection of HBV Resistance to Lamivudine in Patients with Chronic Hepatitis B Using Zip Nucleic Acid Probes in Kerman, Southeast of Iran

  • Afshar, Reza Malekpour;Mollaie, Hamid Reza
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.8
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    • pp.3657-3661
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    • 2012
  • HBV infection is contagious and may be transmitted vertically or horizontally by blood products and body secretions. Over 50% of Iranian carriers have contracted the infection prenatally, making this the most likely route of transmission of HBV in Iran. This study assesses the resistance to Lamivudine in patients with chronic hepatitis B infection using a new ZNA probe Real Time PCR method. To evaluate the effectiveness of Lamivudine therapy for chronic hepatitis B infection, a study was conducted on 70 patients (63 men and 7women), who had received the drug first line. All patients were tested for the presence of HBsAg and HBeAg, the serum ALT level and the HBV DNA load before and after treatment. In all samples resistance to Lamivudine was tested with the ZNA Probe. Our results showed that ZNA Probe Real Time PCR method could detect wild type,YMDD, and its mutants, tyrosine-isoleucine-aspartate-aspartate and tyrosine-valine-aspartate-Aspartate. Among an estimated seventy patients with chronic hepatitis B infection, 18 (25.7%) were resistant to lamivudine. Only one patient was negative for presence of HBS-Ag (5.6%) and two patients were negative for HBe-Ag (11.1%). Real-time PCR with Zip nucleic acid probes is a sensitive, specific and rapid detection method for mutations in the YMDD motif, which will be essential for monitoring patients undergoing Lamivudine antiviral therapy.

Factors Affecting in Vitro Activity of LB20304, a New Flu-oroquinolone

  • Paek, Kyoung-Sook;Ahn, Mi-Jeong;Kim, Mu-Yong;Kim, In-Chull;Kwak, Jin-Hwan
    • Archives of Pharmacal Research
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    • v.19 no.2
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    • pp.143-147
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    • 1996
  • LB20304 is a novel fluoroquinolone that exhibits a potent broad spectrum antibacterial activity against both gram-positve and gram-negative bacteria. The MICs (Minimal Inhibitory Concentration) of LB20304 were determined against both gram-positve and gram-negative bacteria under various conditions including several media, pHs, and inoculum concentrations. The in vitro activity of LB20304 was not significantly affected by the changes in testing conditions such as components of media and inoculum concentrations, but it was slightly reduced by acid condition. The MICs and MBCs (Minimal Bactericidal Concentration) of LB20304 against Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa were hardly affected by the presence of 50 % human serum, mouse serum, guinea pig serum or horse serum, and the MBCs were equal to or at most four-times higher than the MiCs. The activities of LB20304 were decreased by the presence of high concentraion of $Mg^{++}$ or human urine (pH, 5.5) in the test media. The frequencies of mutants resistant to LB20304 were similar to or lower than those found in ciprofloxacin and sparfloxacin.

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Mitogen-Activated Protein Kinase Kinase 3 Is Required for Regulation during Dark-Light Transition

  • Lee, Horim
    • Molecules and Cells
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    • v.38 no.7
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    • pp.651-656
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    • 2015
  • Plant growth and development are coordinately orchestrated by environmental cues and phytohormones. Light acts as a key environmental factor for fundamental plant growth and physiology through photosensory phytochromes and underlying molecular mechanisms. Although phytochromes are known to possess serine/threonine protein kinase activities, whether they trigger a signal transduction pathway via an intracellular protein kinase network remains unknown. In analyses of mitogen-activated protein kinase kinase (MAPKK, also called MKK) mutants, the mkk3 mutant has shown both a hypersensitive response in plant hormone gibberellin (GA) and a less sensitive response in red light signaling. Surprisingly, light-induced MAPK activation in wild-type (WT) seedlings and constitutive MAPK phosphorylation in dark-grown mkk3 mutant seedlings have also been found, respectively. Therefore, this study suggests that MKK3 acts in negative regulation in darkness and in light-induced MAPK activation during dark-light transition.

Amylase Production by Continuous Cultures of Aspergillus oryzae and its Mutants (Aspergllus oryza와 그 변이주의 연적배양에 의한 amylase 생성에 관한 연구)

  • Han, Hong-Eui
    • Korean Journal of Microbiology
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    • v.15 no.2
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    • pp.63-76
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    • 1977
  • Irradiation with high doses of gamma rays induced the reduction of mycelial weight and anaylase activity, and increased relative amylase activity in surface cultures. Biphase in growth curves was shown in aeration-agitation cultures but the behavior of the first phase of growth could be eliminated by replacing the amylasehydrolysed starch substrates, so that enzyme production was shortened ca. 40 hours and relative amylase activity was increased about 3 times higher before onset of autolysis. In the effect of gibberellin on amylase production, the positive stimulation was appeared to only surface culturs of the liquid medium and the negative effect to shake-cultures in a mutant. Trials of various continuous culture were resulted not only the approalch to the value of amylase activity in surface cultures of liquid medium, but also higher productivity than in batch cultures. The culture-degeneration was observed in two-stage continuous culture, but did not appear in continuous elevation culture.

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Colistin 생산균주의 균주개량 및 productivity 증대를 위한 발효최적화

  • Ye, Byeong-Dae;Hwang, Yong-Bae;Kim, Yeong-Hui;Kim, Dong-Geon;Yang, Ho-Seok
    • 한국생물공학회:학술대회논문집
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    • 2002.04a
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    • pp.224-227
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    • 2002
  • Colistin produced from Penibacillius polymyxa was widely used as an antibiotic active against gram-negative bacteria and as feed additive. This research studied on increment of colistin productivity by mutation of P. polymyxa. As a result, several mutants were obtained from the strain by UV radiation and NTG treatment. They produced approximately 8.5${\sim}$9.0 g/L of colistin in flask and jar culture. Colistin productivity of the mutant, named Penibacillius polymyxa CBY, showed 100 times than that of wild type. When Penibacillius polymyxa CBY fermented in the optimal medium, it produced up to 18 g/L of colistin in jar fermentation.

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