• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2007년도 International Meeting of the Microbiological Society of Korea
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• pp.120-122
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• 2007

All living organisms use numerous signal-transduction pathways to sense and respond to their environments and thereby survive and proliferate in a range of biological niches. Molecular dissection of these signalling networks has increased our understanding of these communication processes and provides a platform for therapeutic intervention when these pathways malfunction in disease states, including infection. Owing to the expanding availability of sequenced genomes, a wealth of genetic and molecular tools and the conservation of signalling networks, members of the fungal kingdom serve as excellent model systems for more complex, multicellular organisms. Here, we employed Cryptococcus neoformans as a model system to understand how fungal-signalling circuits operate at the molecular level to sense and respond to a plethora of environmental stresses, including osmoticshock, UV, high temperature, oxidative stress and toxic drugs/metabolites. The stress-activated p38/Hog1 MAPK pathway is structurally conserved in many organisms as diverse as yeast and mammals, but its regulation is uniquely specialized in a majority of clinical Cryptococcus neoformans serotype A and D strains to control differentiation and virulence factor regulation. C. neoformans Hog1 MAPK is controlled by Pbs2 MAPK kinase (MAPKK). The Pbs2-Hog1 MAPK cascade is controlled by the fungal "two-component" system that is composed of a response regulator, Ssk1, and multiple sensor kinases, including two-component.like (Tco) 1 and Tco2. Tco1 and Tco2 play shared and distinct roles in stress responses and drug sensitivity through the Hog1 MAPK system. Furthermore, each sensor kinase mediates unique cellular functions for virulence and morphological differentiation. We also identified and characterized the Ssk2 MAPKKK upstream of the MAPKK Pbs2 and the MAPK Hog1 in C. neoformans. The SSK2 gene was identified as a potential component responsible for differential Hog1 regulation between the serotype D sibling f1 strains B3501 and B3502 through comparative analysis of their meiotic map with the meiotic segregation of Hog1-dependent sensitivity to the fungicide fludioxonil. Ssk2 is the only polymorphic component in the Hog1 MAPK module, including two coding sequence changes between the SSK2 alleles in B3501 and B3502 strains. To further support this finding, the SSK2 allele exchange completely swapped Hog1-related phenotypes between B3501 and B3502 strains. In the serotype A strain H99, disruption of the SSK2 gene dramatically enhanced capsule biosynthesis and mating efficiency, similar to pbs2 and hog1 mutations. Furthermore, ssk2, pbs2, and hog1 mutants are all hypersensitive to a variety of stresses and completely resistant to fludioxonil. Taken together, these findings indicate that Ssk2 is the critical interface protein connecting the two-component system and the Pbs2-Hog1 pathway in C. neoformans.

• 한국균학회지
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• 제18권3호
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• pp.115-126
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• 1990

구름버섯은 항종양작용이 인정되었으며 근자에는 AIDS virus에 대한 억제효과가 보고되고 있다. 이와 같은 약효를 인정받고 있는 구름버섯 균주간의 세포융합이나 구름버섯과 타 유효한 버섯과 세포융합을 시행함으로써 더욱 효능이 우수하거나 다양한 균주의 약효를 한 균주내에서 기대할 수 있는 새로운 균주의 개발이 가능하다. 이러한 목적으로 구름버섯의 원형질체융합을 시행하기 위하여, 원형질체분리, 재생 및 두 영양요구주의 융합에 관하여 실험하였다. 균사체를 2.5일간 셀로판지위에서 배양하여 Novozym 234와 cellulase Onozuka R-10이 각각 15 mg/ml와 10 mg/ml 포함되어 있는 0.6M sucrose 용액에 넣어 $30^{\circ}C$에서 3-4.5시간 반응시킬 때 원형질체 수득률이 가장 높았다. 삼투압 안정제로 0.6M sucrose는 원형질체 형성과 재생에서 최적조건이었다. 0.6M sucrose를 포함하는 고체배지에서의 재생빈도는 3.48%이었으며 UV 조사시 생존률은 0.02-7.4%이었다. 원형질체융합은 $30^{\circ}C$에서 10분 동안 polyethylene glycol(M.W. 4,000)을 이용하여 시행하였는데 그 융합빈도는 1.86%이었다.