Institute of Plant and Microbial Biology

實驗室研究主題

植物抗病性之研究發展與應用
本實驗室長期從事茄科植物與植物病原微生物交互作用的研究，發現在甜椒中有兩種會影響植物過敏反應發生的植物蛋白，分別命名為PFLP與HRAP。這兩種蛋白質在與細菌的致敏蛋白harpinPss混合接種之後，均能在接種植物的細胞間隙中促進harpinPss所引發的過敏反應。在經過基因選殖與許多的定性分析之後，我們發現這兩種蛋白及其基因的重要性質如下：

1.	PFLP (Plant Ferredoxin-Like Protein) PFLP是屬於Ferredoxin(硫鐵蛋白)一類的蛋白質，位於植物細胞內的葉綠體中。植物病原細菌的接種會壓抑植物光合作用硫鐵蛋白的產生(圖一)，並減弱植物的抗病力。在轉殖菸草的實驗中顯示，當致敏原刺激時(如harpinPss接種)，增加PFLP的表現量能增加致敏原所引發的過氧化物，進而降低植物細胞表現過敏反應之細胞死亡的門檻。藉由這種機制，許多原本會在菸草上致病的病原菌(包括Pseudomonas屬與Erwinia屬之細菌性病原菌，以及Botrytis屬之真菌性病原菌)，在pflp的轉殖菸草上卻引起抗病的過敏反應，而使其無法寄生，造成轉基因植物對植物病害更具抗性。這項發現提供農作物在選育抗病品種時一項有效策略之應用。根據此項發現我們已獲得抗白葉枯病的水稻、抗軟腐病的青花菜、抗青枯病與軟腐病的蕃茄(圖二)以及抗軟腐病的彩色海芋(圖三)等。該基因並於2006年8月11日，本著人道協助非洲農民解決香蕉萎凋病之精神，由中央研究院與非洲農業技術基金會簽署授權合約，期能協助非洲農民解決香蕉生產困境，改善糧食問題。

	圖一、注射接種非親合性Pseudomonas syringae pv. syringae(Pss)的細菌懸浮液後，在接種區域內Fd-I的含量明顯增加(A)；而接種親合性Erwinia carotovora subsp. carotovora(Ecc)細菌懸浮液後，接種區域內Fd-I的含量則會減少(B)。Mock為僅注射接種磷酸緩衝液之對照組。


	圖二、硫鐵蛋白轉殖番茄品系(24-18-7)相較於未轉殖植物(Cln1558a)，對細菌性軟腐病(A)及茄科青枯病(B)皆顯現出較強的抗病性。


	圖三、八週大(a)及四週大(b)之硫鐵蛋白轉殖彩色海芋品系(Fa7及Fa24)相較於未轉殖植物，對細菌性軟腐病皆可顯現出較強的抗病性。

2.	HRAP (Hypersensitive Response Assisting Protein) HRAP的基因hrap是一個新發現的基因，與已知功能的蛋白或基因序列均不相似。依序列的分析顯示，HRAP極可能位於植物細胞表面，而有機會與病原菌所分泌的分子發生交互作用。在生化實驗中，HRAP蛋白能解離harpinPss的多聚體，使得harpin蛋白在植物細胞上更容易引起過敏反應。另一方面，HRAP蛋白也可在植物細胞間隙中降低細菌性病原菌的致病力。在天然甜椒植株中，hrap是要在非親合性病原菌的感染之下才會被誘發表現。但在hrap轉殖菸草中，HRAP的持續表現，造成轉殖植物對Pseudomonas屬，Xanthomonas屬與Erwinia屬等細菌性病原菌都具有效的抗性。

pflp與hrap兩個基因在多種植物抗病性上的應用以及在逆境下的反應，正在多方面的進行中。而pflp在抗病上的應用，已獲得歐美與本地的專利。

│研究員簡介│研究領域│重要著作│實驗室成員│

近期著作

相關資料

Hsiang-En Huang, Chien-An Liu, Mei-Jiuan Lee, C-George Kuo, Huei-Mei Chen, Mang-Jye Ger, Yu-Chih Tsai, Yen-Ru Chen, Ming-Kun Lin, and T-Y Feng 2007. Resistance enhancement of transgenic tomato to bacterial pathogens by the heterologous expression of sweet pepper ferredoxin-I protein (PFLP). Phytopathology 97:900-906.
Yip MK,Huang HE,Ger MJ, Chen CY, Chiu SH, Tsai YC, Lin CI and TY. Feng. 2007. Production of soft rot resistant cally lily by expressing a ferredoxin-like protein gene (pflp) in transgenic plants. Plant Cell Report 26:449-457.
Huang HE,Ger MJ, Chen CY, Pandey AK, Yip MK, Chou HW and TY. Feng. 2007. Dosease resisitance to bacterial pathogen effected by the qunatity of ferredoxin-I protein in plants. Molecular Plant pathology 8:129-137.
Tang, K.X., Sun, X.F., Hu, Q.N., Wu, A.Z., Lin, C.H., Lin, H.-J., Twyman, R.M., Christou, P., and Feng, T.-Y.(2001).
Trangenic rice plants expressing the ferredoxin-like proein (AP1) from sweet pepper show enhanced resistance to Xanthomonas oryzae pv. oryzae. Plant Sci. 160, 1035-1042.
Podile, A.R., Lin, H.J., Stainforth, V., Sripriya, P., Chen, L.-F.O., and Feng, T.-Y.(2001).
Conditional expression of harpinpss causes yeast cell death that shares features of cell death pathway with harpinpss-mediated hypersensitive response (HR). Physiol. Mol. Plant Pathol. 58, 267-276.

Dayakar, B.V., Lin, H.-J., Chen, C.-H., Ger, M.-J., Lee, B.-H., Pai, C.-H., Chow, D., Huang, H.-E., Hwang, S.-Y., Chung, M.-C., and Feng, T.-Y.(2003).
Ferredoxin from sweet pepper (Capsicum annuum L.) intensifying harpinpss-mediated hypersensitive response shows an enhanced production of active oxygen species (AOS). Plant Mol. Biol. 51, 913-924.

Ger, M.-J., Chen, C.-H., Hwang, S.-Y., Huang, H.-E., Podile, A.R., Dayakar, B.V., and Feng, T.-Y.(2003).
Constitutive expression of hrap gene in transgenic tobacco plant enhances resistance against virulent bacterial pathogens by induction of a hypersensitive response. Mol. Plant-Microbe Interact. 15, 764-773.

Liau, C.-H., Lu, J.-C., Prasad, V., Hsiao, H.-H., You, S.-J., Lee, J.-T., Yang, N.-S., Huang, H.-E., Feng, T.-Y., Chen, W.-H., and Chan, M.-T.(2003).
The sweet pepper ferredoxin-like protein (pflp) conferred resistance against soft rot disease in Oncidium orchid. Transgenic Res. 12, 329-336

Lu, J.-C., Liau, C.-H., Prasad, V., Lee, J.-T., Hsiao, H.-H., You, S.-J., Yang, N.-S., Huang, S.-E., Feng, T.-Y., Chen, W.-H., and Chan, M.-T.(2003).
Oncidium orchid ectopically expressing the sweet pepper ferredoxin-like protein (AP1) conferred resistance against soft rot disease. Transgenic Res. 12, 329-336.

You, S.-J., Liau, C.-H., Huang, H.-E., Feng, T.-Y., Prasad, V., Hsiao, H.-H., Lu, J.-C., and Chan, M.-T.(2003).
Sweet pepper ferredoxin-like protein (pflp) gene as a novel selection marker for orchid transformation. Planta 217, 60-65.

Huang, H.-E., Ger, M.-J., Yip, M.-K., Chen, C.-Y., Pandeya, A.-K., and Feng, T.-Y.(2004).
A hypersensitive response was induced by virulent bacteria in transgenic tobacco plants overexpressing a plant ferredoxin-like protein (PFLP). Physiol. Mol. Plant Pathol. 64, 103-110.

Pandey, A.-K., Ger, M.-J., Huang, H.-E., Yip, M.-K., Zeng, J., and Feng, T.-Y.(2005).
Expression of the hypersensitive response-assisting protein in Arabidopsis results in harpin-dependent hypersensitive cell death in response to Erwinia carotovora. Plant Mol. Biol. 59, 771-780.

Huang, H.-E., Ger, M.-J., Chen, C.-Y., Yip, M.-K., Chung, M.-C., and Feng, T.-Y.(2006).
Plant ferredoxin-like protein (PFLP) exhibits an anti-microbial ability against soft-rot pathogen Erwinia carotovora subsp. carotovora in vitro and in vivo. Plant Sci. 171, 17-23.

Huang, H.-E., Ger, M.-J., Chen, C.-Y., Pandey, A.-K., Yip, M.-K., Chou, H.-W., and Feng, T.-Y.(2007).
Disease resistance to bacterial pathogens affected by the amount of ferredoxin-I protein in plants. Mol. Plant Pathol. 8, 129-137.

Huang, H.-E., Liu, C.-A., Lee, M.-J., Kuo, C.-G., Chen, H.-M., Ger, M.-J., Tsai, Y.-C., Chen, Y.-R., Lin, M.-K., and Feng, T.-Y.(2007).
Resistance enhancement of transgenic tomato to bacterial pathogens by the heterologous expression of sweet pepper ferredoxin-I protein. Phytopathol. 97, 900-906.

Yip, M.-K., Huang, H.-E., Ger, M.-J., Chiu, S.-H., Tsai, Y.-C., Lin, C.-I., and Feng, T.-Y. (2007).
Production of soft rot resistant calla lily by expressing a ferredoxin-like protein gene (pflp) in transgenic plants. . Plant Cell Rep. 26, 449-457.

專利

Lin, H.-J., and FENG, T.-Y. (1999. Oct.).
The use of a ferredoxin-like gene to protect plants against bacterial diseases. (USA patent) patent no. 5968804
Lin, H.-J., and FENG, T.-Y.(2001. Oct 2.).
The use of a ferredoxin-like gene to protect plants against bacterial diseases. (USA patent) patent no. 6297360.
Lin, H.-J., and Feng, T.-Y. (2002. Sep. 13.).
Amphipathic protein-1. (Netherlands patent) patent no. 1011688.