植物激素——油菜素内酯

1、BrassinosteroidsPlant Steroid HormonesBrassinosteroidsBrassinolide (BL)Brassinosteroids (BRs) are a family of 50 structurally related compounds that contribute to:GrowthCell division, elongation, and differentiationDefense against pathogensStress toleranceReproductive developmentCastasterone (CS)Pol

2、len extract was found to promote cell and stem elongation (1941)Mitchell, J.W., and Whitehead, M.R. (1941) Responses of vegetative parts of plants following application of extract of pollen from Zea mays. Bot. Gaz. 102: 770-791 POLLEN EXTRACTCONTROLBeans treated with pollen extract showed dramatic s

3、tem and cell elongation as compared to control beans. The novel compound was partially purified and named “brassin” (1970)Reprinted by permission from Macmillan Publishers Limited, Nature: Mitchell, J.W., Mandava, N., Worley, J.F., Plimmer, J.R., and Smith, M.V. (1970). Brassins - a new family of pl

4、ant hormones from rape pollen. Nature 225: 1065-1066, copyright 1970. ControlGibberellin BrassinosteroidIn 1970 Mitchell and colleagues showed that their newly purified compound promotes stem elongation. The structure of brassinolide (BL) was identifiedBrassinolide (BL)Starting with pollen collected

5、 by bees, brassinolide was purified and its structure determined (1979).Reprinted by permission from Macmillan Publishers Limited, Nature: Grove, M.D., Spencer, G.F., Rohwedder, W.K., Mandava, N., Worley, J.F., Warthen, J.D., Steffens, G.L., Flippen-Anderson, J.L., and Cook, J.C. (1979). Brassinolid

6、e, a plant growth-promoting steroid isolated from Brassica napus pollen. Nature 281: 216-217; copyright 1979.Like other hormones, BL is not very abundant in plant tissuesReprinted with permission from from Mandava, N., Kozempel, M., Worley, J.F., Matthees, D., Warthen, J.D., Jacobson, M., Steffens,

7、G.L., Kenney, H., and Grove, M.D. (1978). Isolation of brassins by extraction of rape (Brassica napus L.) pollen. Ind. Eng. Chem. Prod. Res. Dev. 17: 351-354, copyright 1978 American Chemical Society. 230 kgs pollen10 mg BLBrassinolide and castasterone are steroids like some animal hormones InsectPl

8、antMammalBishop, G.J., and Koncz, C. (2002). Brassinosteroids and plant steroid hormone signaling. Plant Cell 14: S97-110.Characterization of det2 and cpd showed that BRs are essentialFrom Li, J. et al. (1996) A role for brassinosteroids in light-dependent development of Arabidopsis. Science 272: 39

9、8401; reprinted with permission from AAAS. Reprinted from Szekeres, M. et al. (1996) Brassinosteroids rescue the deficiency of CYP90, a cytochrome P450, controlling cell elongation and de-etiolation in Arabidopsis. Cell 85: 171182 with permission from Elsevier. WTdet2det2 + BLWTdet2det2 + BLWTcpdIn

10、1996, two Arabidopsis mutants, de-etiolated2 (det2) and constitutive photomorphogenesis and dwarfism (cpd) were shown to be BR-deficient. Lecture outline BR biosynthesis, homeostasis and transport Brassinosteroid receptors BR signaling from the receptor to nucleus BRs in whole-plant physiology Ongoi

11、ng researchCatharanthus roseus cultured cells helped define the synthesis pathwayPhoto credit titanium22The products of a radioactively-labeled precursor can be identifiedNoguchi, T., Fujioka, S., Takatsuto, S., Sakurai, A., Yoshida, S., Li, J., and Chory, J. (1999). Arabidopsis det2 Is defective in

12、 the conversion of (24R)-24-methylcholest-4-en-3-one to (24R)-24-methyl-5alpha -cholestan-3-one in brassinosteroid biosynthesis. Plant Physiol. 120: 833-840.The white circles indicate introduced compounds and the black circles the products formed. W = wild-type ArabidopsisV = Catharanthus roseus.Blo

13、cked steps in biosynthetic mutants can be identifiedNoguchi, T., Fujioka, S., Takatsuto, S., Sakurai, A., Yoshida, S., Li, J., and Chory, J. (1999). Arabidopsis det2 Is defective in the conversion of (24R)-24-methylcholest-4-en-3-one to (24R)-24-methyl-5alpha -cholestan-3-one in brassinosteroid bios

14、ynthesis. Plant Physiol. 120: 833-840.d = det2 mutant of Arabidopsis det2 is blocked at this stepThese studies suggested biosynthesis involves a “metabolic grid”ABThere are many ways to get from Point A (campesterol) to Point B (brassinolide).CampesterolBrassinolideA likely pathway for BR synthesisR

15、eprinted from Ohnishi, T., Yokota, T., and Mizutani, M. (2009). Insights into the function and evolution of P450s in plant steroid metabolism. Phytochemistry 70: 1918-1929 with permission from Elsevier.CampesterolCastasteroneBrassinolideDET2DWF4DWF1 / LKB / BRD2DWARF(tomato)CampesterolPlants mutated

16、 in the production of campesterol are dwarfedChoe, S., et al. (1999). The Arabidopsis dwarf1 mutant is defective in the conversion of 24-methylenecholesterol to campesterol in brassinosteroid biosynthesis. Plant Physiol. 119: 897-908; Hong, Z., et al. (2005). The rice brassinosteroid-deficient dwarf

17、2 mutant, defective in the rice homolog of Arabidopsis DIMINUTO/DWARF1, is rescued by the endogenously accumulated alternative bioactive brassinosteroid, dolichosterone. Plant Cell 17: 2243-2254; Nomura, T., et al., (1997). Blockage of brassinosteroid biosynthesis and sensitivity causes dwarfism in

18、garden pea. Plant Physiol. 113: 31-37.Mutants blocked in this step are called dwarf1 (Arabidopsis), lkb (pea) and brassinosteroid-deficient dwarf2 (rice). lkbWild typeLKBBRD2Wild typebrd2Wild typedwf1Many BR-synthesis enzymes are cytochrome P450s (CYPs)Reprinted from Bishop, G.J. (2007). Refining th

19、e plant steroid hormone biosynthesis pathway. Trends Plant Sci. 12: 377-380, with permission from Elsevier. cpddwf4C6-oxidaseC22-oxidaseC23-oxidaseC6-oxidase*dx (tomato)C22-oxidaseC3-oxidaseFor simplicity, only Arabidopsis genes are shown hereDWF4 encodes a CYP C22-oxidase dwf4 wild-typeDWF4 encodes

20、 cytochrome P450 CYP90B1, a C22-oxidase. Loss-of-function mutants are dwarfed.DWF4Four dwarf4 mutantsAzpiroz, R., Wu, Y., LoCascio, J.C., and Feldmann, K.A. (1998). An Arabidopsis brassinosteroid-dependent mutant Is blocked in cell elongation. Plant Cell 10: 219-230; Choe, S., Dilkes, B.P., Fujioka,

21、 S., Takatsuto, S., Sakurai, A., and Feldmann, K.A. (1998). The DWF4 gene of Arabidopsis encodes a cytochrome P450 that mediates multiple 22-hydroxylation steps in brassinosteroid biosynthesis. Plant Cell 10: 231-244.DWF4 encodes a CYP C22-oxidase dwf4 wild-typeDWF4 encodes cytochrome P450 CYP90B1,

22、a C22-oxidase. Loss-of-function mutants are dwarfed.DWF4Four dwarf4 mutantsAzpiroz, R., Wu, Y., LoCascio, J.C., and Feldmann, K.A. (1998). An Arabidopsis brassinosteroid-dependent mutant Is blocked in cell elongation. Plant Cell 10: 219-230; Choe, S., Dilkes, B.P., Fujioka, S., Takatsuto, S., Sakura

23、i, A., and Feldmann, K.A. (1998). The DWF4 gene of Arabidopsis encodes a cytochrome P450 that mediates multiple 22-hydroxylation steps in brassinosteroid biosynthesis. Plant Cell 10: 231-244.DWF4 is expressed in young, elongating tissuesKim, H.B., Kwon, M., Ryu, H., Fujioka, S., Takatsuto, S., Yoshi

24、da, S., An, C.S., Lee, I., Hwang, I., and Choe, S. (2006). The regulation of DWARF4 expression is likely a critical mechanism in maintaining the homeostasis of bioactive brassinosteroids in Arabidopsis. Plant Physiol. 140: 548-557.The accumulation pattern of BRs is closely correlated with DWF4 expre

25、ssionDE-ETIOLATED2 encodes a 5 -reductase Chory, J., Nagpal, P., and Peto, C.A. (1991). Phenotypic and genetic analysis of det2, a new mutant that affects light-regulated seedling development in Arabidopsis. Plant Cell 3: 445-459; Nomura, T., et al. (2004). Brassinosteroid deficiency due to truncate

26、d steroid 5-reductase Causes dwarfism in the lk mutant of pea. Plant Physiol. 135: 2220-2229. WTdet2DET2WTlkLK is the pea orthologue of DET2DE-ETIOLATED2 encodes a 5 -reductase WTdet2WTlkLK is the pea orthologue of DET2Chory, J., Nagpal, P., and Peto, C.A. (1991). Phenotypic and genetic analysis of

27、det2, a new mutant that affects light-regulated seedling development in Arabidopsis. Plant Cell 3: 445-459; Nomura, T., et al. (2004). Brassinosteroid deficiency due to truncated steroid 5-reductase Causes dwarfism in the lk mutant of pea. Plant Physiol. 135: 2220-2229. DET2In dicots, two genes enco

28、ding CYP85A are functionally differentNomura, T., Kushiro, T., Yokota, T., Kamiya, Y., Bishop, G.J., and Yamaguchi, S. (2005). The last reaction producing brassinolide is catalyzed by cytochrome P-450s, CYP85A3 in tomato and CYP85A2 in Arabidopsis. J. Biol. Chem. 280: 17873-17879.MonofunctionalBifun

29、ctionalOne synthesizes castasterone, the other brassinolideNomura, T., Kushiro, T., Yokota, T., Kamiya, Y., Bishop, G.J., and Yamaguchi, S. (2005). The last reaction producing brassinolide is catalyzed by cytochrome P-450s, CYP85A3 in tomato and CYP85A2 in Arabidopsis. J. Biol. Chem. 280: 17873-1787

30、9.CYP85A1CYP85A2 or 3CYP85A2 or 3In tomato and Arabidopsis, CYP85A1 makes castasterone (CS). CYP85A2 or A3 has a second catalytic property that converts CS to BL. In tomato, CS and BL accumulate in different tissuesNomura, T., Kushiro, T., Yokota, T., Kamiya, Y., Bishop, G.J., and Yamaguchi, S. (200

31、5). The last reaction producing brassinolide is catalyzed by cytochrome P-450s, CYP85A3 in tomato and CYP85A2 in Arabidopsis. J. Biol. Chem. 280: 17873-17879.CYP85A1 is expressed in vegetative tissues and converts 6-deoxocastasterone to castasteroneCastasterone is abundant in vegetative tissuesCYP85

32、A1BL accumulates in fruit where CYP85A3 is expressedNomura, T., Kushiro, T., Yokota, T., Kamiya, Y., Bishop, G.J., and Yamaguchi, S. (2005). The last reaction producing brassinolide is catalyzed by cytochrome P-450s, CYP85A3 in tomato and CYP85A2 in Arabidopsis. J. Biol. Chem. 280: 17873-17879.CYP85

33、A3CYP85A3 is expressed in developing fruit and converts castasterone to brassinolideBrassinolide is abundant in tomato fruitCYP85A1CYP85A3The dx mutant is dwarfed but with normal fruitNomura, T., Kushiro, T., Yokota, T., Kamiya, Y., Bishop, G.J., and Yamaguchi, S. (2005). The last reaction producing

34、 brassinolide is catalyzed by cytochrome P-450s, CYP85A3 in tomato and CYP85A2 in Arabidopsis. J. Biol. Chem. 280: 17873-17879.The plants do not produce BR in vegetative tissues because of the dx mutation in the CYP85A1 gene. The CYP85A3 gene produces brassinolide only in fruit.Monocots have only a

35、single CYP85 gene and may not produce BLReprinted from Kim, B.K., Fujioka, S., Takatsuto, S., Tsujimoto, M., and Choe, S. (2008). Castasterone is a likely end product of brassinosteroid biosynthetic pathway in rice. Biochemical and Biophysical Research Communications 374: 614-619 with permission fro

36、m Elsevier. Dicots have pairs of genesThere are several pathways for BL deactivationReprinted from Ohnishi, T., Yokota, T., and Mizutani, M. (2009). Insights into the function and evolution of P450s in plant steroid metabolism. Phytochemistry 70: 1918-1929 with permission from Elsevier.BRs are deact

37、ivated by CYP72B1Turk, E.M., Fujioka, S., Seto, H., Shimada, Y., Takatsuto, S., Yoshida, S., Denzel, M.A., Torres, Q.I., and Neff, M.M. (2003). CYP72B1 inactivates brassinosteroid hormones: An intersection between photomorphogenesis and plant Steroid Signal Transduction. Plant Physiol. 133: 1643-165

38、3.Loss-of-functionIncreased hypocotyl lengthGain-of-functionDecreased hypocotyl lengthCYP72B1ACTIVEINACTIVEBRs may also be deactivated by glucosylation and other reactionsPoppenberger, B., Fujioka, S., Soeno, K., George, G.L., Vaistij, F.E., Hiranuma, S., Seto, H., Takatsuto, S., Adam, G., Yoshida,

39、S., and Bowles, D. (2005). The UGT73C5 of Arabidopsis thaliana glucosylates brassinosteroids. Proc. Natl. Acad. Sci. USA 102: 15253-15258, copyright 2005 National Academy of Sciences, USA. When overexpressed, the UDP-glycosyltransferase UGT73C5 deactivates brassinosteroids and limits plant growth. S

40、ymons GM, Reid, JB. (2004) Brassinosteroids do not undergo long-distance transport in pea. Implications for the regulation of endogenous brassinosteroid levels, Plant Physiol. 135: 21962206.Wild-type pea roots cannot rescue BR-deficient shoots, indicating that localized BR synthesis is required for

41、stem elongation. Similar results have been obtained from tomato. Grafting studies suggest that endogenous BR is not transported long-distancesBL negatively regulates expression of BL biosynthesis genesBancos, S., et al. (2002). Regulation of Transcript Levels of the Arabidopsis Cytochrome P450 Genes

42、 Involved in Brassinosteroid Biosynthesis. Plant Physiol. 130: 504-513; Kim, H.B., et al. (2006). The regulation of DWARF4 expression is likely a critical mechanism in maintaining the homeostasis of bioactive brassinosteroids in Arabidopsis. Plant Physiol. 140: 548-557.; Shimada, Y., et al. (2003).

43、Organ-Specific Expression of Brassinosteroid-Biosynthetic Genes and Distribution of Endogenous Brassinosteroids in Arabidopsis. Plant Physiol. 131: 287-297.BrassinolideCampesterolExpression levels of most of the BL biosynthesis genes are negatively regulated by BL.CYP85A2 mRNA; hours after BL additi

44、on+ BLCPDDWF4rRNACPDDWF4UBQ10CYP85A1CYP85A2CYP90C1CYP90D1BR synthesis is also controlled by light and circadian cyclesBancos, S., Szatmari, A.-M., Castle, J., Kozma-Bognar, L., Shibata, K., Yokota, T., Bishop, G.J., Nagy, F., and Szekeres, M. (2006). Diurnal regulation of the brassinosteroid-biosynt

45、hetic CPD gene in Arabidopsis. Plant Physiol. 141: 299-309.CPDpro:LUCCYP85A2pro:LUCRegulation of biosynthetic gene transcription is being investigatedBrassinolideCampesterolRAVL1TCP1Guo, Z., Fujioka, S., Blancaflor, E.B., Miao, S., Gou, X., and Li, J. (2010) TCP1 modulates brassinosteroid biosynthes

46、is by regulating the expression of the key biosynthetic gene DWARF4 in Arabidopsis thaliana. Plant Cell 22: 1161-1173.Overexpression of the TCP1 transcription factor leads to elevated expression of DWF4 and enhanced growth.Several transcription factors that directly regulate BR synthesis genes have

47、recently been identifiedAccumulation of active BRs - summary BRs biosynthesis mutants are dwarfed; a family of cytochrome P450 genes is central to their synthesis DWARF4 expression is closely correlated with BR levels BRs are deactivated by reversible conjugation and irreversible modifications BRs a

48、ppear to be synthesized at their site of action; there is no evidence for long-distance transport of endogenous BRs BR synthesis is controlled by negative feedback, light and circadian cycles, and interactions with other hormonesBrassinosteroid signalingBRI1 encodes the main BR receptor; bri1 mutant

49、s are BR-insensitiveClouse, S.D., Langford, M., and McMorris, T.C. (1996). A brassinosteroid-insensitive mutant in Arabidopsis thaliana exhibits multiple defects in growth and development. Plant Physiol. 111: 671-678.bri1Wild-typeWild-typebri1bri1In wild-type plants, root growth is inhibited by elev

50、ated BR levelsThe Arabidopsis bri1 mutant is very small (below) and insensitive to exogenous BRbri receptor mutants have been identified in rice and tomatoYamamuro, C., et al. (2000). Loss of runction of a rice brassinosteroid insensitive1 homolog prevents internode elongation and bending of the lam

51、ina joint. Plant Cell 12: 1591-1606. Montoya, T., et al. (2002). Cloning the tomato Curl3 gene highlights the putative dual role of the leucine-rich repeat receptor kinase tBRI1/SR160 in plant steroid hormone and peptide hormone signaling. Plant Cell 14: 3163-3176.Receptor mutantBiosynthesis mutantW

52、ild typeosbri1The BRI1 receptor is plasma-membrane localizedBRI1Leucine-rich repeat (LRR)Trans-membrane domainKinase DomainReprinted from Li, J., and Chory, J. (1997). A putative leucine-rich repeat receptor kinase involved in brassinosteroid signal transduction. Cell 90: 929-938, with permission fr

53、om Elsevier; Bishop, G.J., and Koncz, C. (2002). Brassinosteroids and plant steroid hormone signaling. Plant Cell 14: S97-110.BRI1 is a Leucine-rich repeat receptor like kinase (LRR-RK)bri1The BRI1 gene was identified through map-based cloning By contrast, animal steroid receptors are intracellularT

54、ranscriptionSteroids freely cross plasma membraneSteroid receptorWhen the receptor binds the hormone, the complex moves into nucleus, binds DNA and initiates transcriptionLeucine-rich repeat receptor-like kinases (LRR-RLKs) are prevalentLRR-RLKs are very abundant in plant and animal cells and bind a

55、 wide variety of signaling molecules.In plants LRR-RLKs are involved in developmental signaling and pathogen responses.PLANTANIMALBishop, G.J., and Koncz, C. (2002). Brassinosteroids and plant steroid hormone signaling. Plant Cell 14: S97-110.Two additional BR receptors have restricted expression pa

56、tternsReproduced with permission from Cao-Delgado, A., Yin, Y., Yu, C., Vafeados, D., Mora-Garca, S., Cheng, J.-C., Nam, K.H., Li, J., and Chory, J. (2004). BRL1 and BRL3 are novel brassinosteroid receptors that function in vascular differentiation in Arabidopsis. Development 131: 5341-5351.The BRI1

57、 gene is expressed throughout the plant.BRL1 and BRL3 are primarily expressed in vascular tissues.The protein encoded by BRL1 and BRL3 can function as receptorsWhen BRL1 or BRL3 proteins are expressed from the BRI1 promoter, they rescue the bri loss-of-function mutant. bri1 loss-of-function mutantRe

58、scued by BRL1 or BRL3 expressed from BRI1 promoterThe BRL genes may encode receptors with cell-specific functions. Reproduced with permission from Cao-Delgado, A., Yin, Y., Yu, C., Vafeados, D., Mora-Garca, S., Cheng, J.-C., Nam, K.H., Li, J., and Chory, J. (2004). BRL1 and BRL3 are novel brassinost

59、eroid receptors that function in vascular differentiation in Arabidopsis. Development 131: 5341-5351.The rice BRL genes have a different expression pattern than BRI1Nakamura, A., Fujioka, S., Sunohara, H., Kamiya, N., Hong, Z., Inukai, Y., Miura, K., Takatsuto, S., Yoshida, S., Ueguchi-Tanaka, M., H

60、asegawa, Y., Kitano, H., and Matsuoka, M. (2006). The role of OsBRI1 and its homologous gGenes, OsBRL1 and OsBRL3, in rice. Plant Physiol. 140: 580-590.The rice OsBRI1 gene is preferentially expressed in shoots. A loss-of-function mutant shows severe dwarfing in the shoot but little change in root g