Regulation of Gene Expression in Fungi for Breaching the Plant Host Barriers

调控真菌基因表达以突破植物宿主屏障

• 批准号: 0313968
• 负责人: P. Kolattukudy
• 金额: $ 30万
• 依托单位: The University of Central Florida Board of Trustees
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-12-01 至 2005-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0313968&HistoricalAwards=false
• 关键词: Regulation; Gene; Expression; Fungi; Breaching

The plant cuticle, being the outermost barrier, is the first point of contact in the interaction between plants and fungal pathogens. The cuticle is composed of an insoluble biopolyester, cutin, and a complex mixture of soluble, non-polar lipids collectively called waxes. Cutin is the first structural barrier that the fungal infection peg must penetrate in order to infect the plant. Prior support from NSF was primarily responsible for the basic information obtained about the chemistry and biosynthesis of cutin, as well as the role of the cuticle in plant-fungus interaction. Cutinase secreted by the invading fungus helps it to attach to the host and penetrate the outer barrier. According to previously obtained evidence, when fungal conidia contact a plant surface, the small amount of constitutively expressed cutinase present on the conidia releases hydroxy fatty acid monomers and oligomers from cutin, and these soluble products cause induction of high levels of cutinase that helps the infection process. With cloned cutinase genes and transcription factors, it has been shown that in Nectria haematococca one transcription factor, CTF1b, regulates a constitutively expressed cutinase gene (cut2), and another transcription factor, CTF1a, is involved in cutin monomer induction of another cutinase gene, cut1. The mechanism by which hydroxy fatty acids or their oligomers induce cut1 remains to be elucidated. Cut1 induction involves a palindromic promoter element that contains 2 overlapping palindromes. Palindrome 2 is required for cut1 induction by cutin monomers. Palindrome binding protein (PBP) binds palindrome1 of cut1 and acts as a repressor that prevents CTF1a binding to palindrome 2. Since palindrome 1 of cut2 has 2 nucleotide substitutions, PBP cannot bind to it and suppress cut2 expression, and thus this gene is constitutively expressed. It is postulated that ctf1a induction by cutin monomers allows CTF1a to outcompete with PBP to bind cut1 promoter and cause cut1 induction. Cutin monomer or oligomer-stimulated phosphorylation of this cutin response element binding protein (CREBP) allows it to bind CRE and transcriptionally activate ctf1a gene. It is also possible that phosphorylation of PBP might weaken its binding to palindrome1 of cut1 and thus help relieve the suppression. This project will test this hypothesis and define these regulatory processes at a molecular level. Disruption of ctf1a gene drastically reduces virulence of N. haematococca, but the drastic decrease in cutinase level resulting from this disruption cannot explain the loss of virulence because supplementation of the inoculum with cutinase or mechanical breaching of the cuticle does not recover virulence of the disruptants. Obviously, this Cys6Zn2 binuclear protein (CTF1a) also regulates other gene(s) required for virulence. This project will seek the identity of these virulence genes. The results expected from this project will elucidate novel molecular mechanisms involved in some of the fundamental processes in plant-fungus interaction, possibly suggesting novel approaches to control fungal diseases. Fungal infection causes by far the most damage to food and fiber production. This project could make a significant contributions to an understanding of this interaction that may help manage damaging fungal diseases of plants.

植物角质层作为最外层的屏障，是植物与真菌病原体相互作用的第一个接触点。角质层由不溶性生物聚酯、角质层和一种称为蜡质的可溶性非极性脂质的复杂混合物组成。角质层是真菌感染钉必须穿透的第一个结构屏障，以便感染植物。NSF的前期支持主要负责角质层的化学和生物合成的基本信息，以及角质层在植物-真菌相互作用中的作用。入侵真菌分泌的角质酶帮助它附着在宿主上并穿透外部屏障。根据先前获得的证据，当真菌分生孢子接触植物表面时，分生孢子上存在的少量组成性表达的角质酶会从角质素中释放出羟基脂肪酸单体和低聚物，这些可溶性产物会诱导高水平的角质酶，从而帮助感染过程。利用克隆的角质酶基因和转录因子，研究表明，在赤血菌中，一个转录因子CTF1b调节一个组成性表达的角质酶基因（cut2），另一个转录因子CTF1a参与另一个角质酶基因cut1的角质蛋白单体诱导。羟基脂肪酸或其低聚物诱导cut1的机制仍有待阐明。Cut1诱导包括一个包含2个重叠回文的回文启动子元件。切割素单体诱导切割1需要回文2。Palindrome binding protein （PBP）结合cut1的Palindrome 1，并作为抑制因子阻止CTF1a结合Palindrome 2。由于cut2的回文1有2个核苷酸取代，PBP不能与它结合并抑制cut2的表达，因此该基因是组成性表达的。假设切割素单体诱导ctf1a使ctf1a能够与PBP竞争结合cut1启动子并引起cut1诱导。角质素单体或寡聚物刺激的这种角质素反应元件结合蛋白（CREBP）磷酸化使其能够结合CRE并转录激活ctf1a基因。PBP的磷酸化也可能削弱其与cut1的回文蛋白1的结合，从而帮助缓解抑制。该项目将测试这一假设，并在分子水平上定义这些调节过程。ctf1a基因的破坏大大降低了嗜血嗜血杆菌的毒力，但由于这种破坏导致的角质酶水平的急剧下降并不能解释毒力的丧失，因为补充接种物的角质酶或机械破坏角质层并不能恢复破坏物的毒力。显然，这个Cys6Zn2双核蛋白（CTF1a）也调节其他毒力所需的基因。该项目将寻求这些毒力基因的身份。该项目的结果将阐明植物与真菌相互作用的一些基本过程的新分子机制，可能为控制真菌疾病提供新方法。迄今为止，真菌感染对食品和纤维生产造成的损害最大。该项目可以为理解这种相互作用做出重大贡献，这可能有助于管理植物的破坏性真菌疾病。