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Construction of Pathogenicity Map of hrp Genes Cluster in Xanthomonas oryzae pathovar oryzae KACC10859

Title
Construction of Pathogenicity Map of hrp Genes Cluster in Xanthomonas oryzae pathovar oryzae KACC10859
Other Titles
Xanthomonas oryzae pathovar oryzae KACC10859 균주의 hrp 유전자군의 Pathogenicity Map 작성
Authors
조희정
Issue Date
2005
Department/Major
대학원 생명과학과
Publisher
이화여자대학교 대학원
Degree
Master
Abstract
Xanthomonas oryzae pathovar oryzae (벼흰잎마름병균)는 아시아, 아프리카 등 전 세계의 벼 재배지에서 흰잎마름병 (Bacterial blight)을 유발하며 세균병 중에서 벼에 가장 큰 피해를 입히는 식물병원균이다. X. oryzae pv. oryzae는 기주식물에 병을 일으키고, 비기주식물에 과민반응 (hypersensitive response, HR)을 일으키는데 필요한 type III secretion system (TTSS)을 생성•분비하는데, 이러한 독특한 단백질 전달 체계는 32.18 kb의 hrp (hypersensitive response and pathogenicity) 유전자군에서 발현된다. hrp 유전자군에는 9개의 hrp 유전자 (hrpF, hrpE, hrpD6, hrpD5, hrpB1, hrpB2, hrpB4, hrpB5, hrpB7), 9개의 hrc (hrp conserved) 유전자 (hrcS, hrcR, hrcQ, hrcV, hrcU, hrcJ, hrcN, hrcT, hrcC), 8개의 hpa (hrp-associated) 유전자 (hpaF, hpaB, hpaP, hpa1, hpa2, hpa3, hpa4)와 7개의 PIP box (plant-inducible promoter)로 구성되어 있다. 본 연구에서는 X. oryzae pv. oryzae KACC10859 균주의 hrp 유전자군에 있는 26개의 hrp 유전자를 분리하여 그 중 24개의 hrp 유전자를 파괴한 돌연변이를 만들었다. 돌연변이는 Tn5 삽입 및 homologous recombination을 이용한 방법으로 만들었으며 그 파괴된 유전자는 hpaF, hrpF, hpaB, hrpE, hrpD6, hrpD5, hrcS, hrcR, hrcQ, hpaP, hrcV, hrcU, hrpB1, hrpB2, hrcJ, hrpB4, hrpB5, hrcN, hrpB7, hrcT, hrcC, hpa1, hpa3 및 hpa4 이다. hrp 유전자가 파괴된 X. oryzae pv. oryzae는 벼 잎에서의 병원성에 영향을 주는 것을 관찰 할 수 있었는데, 이러한 돌연변이로 hrp 유전자군 내 유전자들의 병원성을 검정하고, 이에 대한 pathogenicity map을 작성하였다. hrp 유전자군 내 24 개의 hrp 유전자 돌연변이에 대한 pathogenicity map을 살펴보면, hrpF, hrpE, hrpD6, hrpD5, hrpB1, hrpB2, hrpB4, hrpB5, hrpB7, hrcS, hrcR, hrcQ, hrcV, hrcU, hrcJ, hrcN, hrcT, hrcC 및 hpaB 유전자가 파괴된 돌연변이에서 병원성이 완전히 소실된 것을 볼 수 있었고, hpaF, hpaP, hpa1, hpa3 및 hpa4 돌연변이들는 유전자가 파괴되었음에도 병원성에 큰 영향을 미치지 않았음을 관찰 할 수 있었다. 이러한 결과로 보아, hrp 유전자군 내의 모든 hrp-hrc 유전자와 hpaB 유전자는 X. oryzae pv. oryzae의 벼흰잎마름병 발생에 중요한 요인으로 작용하는 반면, 대부분의 hpa 유전자는 병 발생에 보조적인 역할로 작용함을 알 수 있었다.;Xanthomonas oryzae pathovar oryzae is the causal agent of rice bacterial blight disease. The plant pathogenic bacterium X. oryzae pv. oryzae express a type III secretion system that is necessary for both pathogenicity in susceptible hosts and the induction of the hypersensitive response in resistant plants. This specialized protein transport system is encoded by a 32.18 kb hrp (hypersensitive response and pathogenicity) genes cluster. hrp genes cluster was composed of nine hrp (hrpF, hrpE, hrpD6, hrpD5, hrpB1, hrpB2, hrpB4, hrpB5, hrpB7), nine hrc (hrp conserved; hrcS, hrcR, hrcQ, hrcV, hrcU, hrcJ, hrcN, hrcT, hrcC), and eight hpa (hrp-associated; hpaF, hpaB, hpaP, hpa1, hpa2, hpa3, hpa4) genes with seven plant-inducible promoter (PIP) boxes. Twenty-six hrp genes in cluster of X. oryzae pv. oryzae KACC10859 (Korean Agricultural Culture Collection) had been cloned. Among 26 genes in hrp cluster, 24 genes (hpaF, hrpF, hpaB, hrpE, hrpD6, hrpD5, hrcS, hrcR, hrcQ, hpaP, hrcV, hrcU, hrpB1, hrpB2, hrcJ, hrpB4, hrpB5, hrcN, hrpB7, hrcT, hrcC, hpa1, hpa3 and hpa4) were disrupted by Tn5 insertional mutagenesis. X. oryzae pv. oryzae with mutations in the hrp region have affected on growth in rice leaves. With these mutants, pathogenicity of hrp genes cluster were characterized, and then the pathogenicity map of hrp genes cluster was constructed. The pathogenicity map of twenty-four hrp mutants was shown that each nine hrp (hrpF, hrpE, hrpD6, hrpD5, hrpB1, hrpB2, hrpB4, hrpB5 and hrpB7), nine hrc (hrcS, hrcR, hrcQ, hrcV, hrcU, hrcJ, hrcN, hrcT and hrcC) and 1 hpa (hpaB) mutants were lost their pathogenicity completely. However, each five hpa (hpaF, hpaP, hpa1, hpa3 and hpa4) mutants were retained their pathogenicity the same as those of the wild type. It was suggested that all hrp-hrc and hpaB genes encode an essential pathogenicity factor, and hpaF, hpaP, hpa1, hpa3 and hpa4 genes do not affect on the pathogenicity of X. oryzae pv. oryzae.
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