植物细胞壁合成与加厚调控课件.ppt

1、植物植物细胞壁合成与加厚调控细胞壁合成与加厚调控Robert Hooke观察到的植物细胞壁观察到的植物细胞壁细胞壁是最早观察到的生物微观结构细胞壁是最早观察到的生物微观结构Concrete Forest Natural Forest Common feature:Cell WallsAbundant Cell Wall Material 细胞壁的结构和组成细胞壁的结构和组成Hemicellulose(20%)Lignin(25%)Cellulose(50%)Extract(5%)植物细胞壁结构和组成植物细胞壁结构和组成 Nature Materials(2003)Cellulose Hemic

2、ellulose Pectin图12-7 植物细胞壁电镜图。A：水稻厚壁细胞电镜图。B：A图方框区域的放大。ML：中胶层；PCW：初生壁；SCW：次生壁。S1，S2，S3：次生细胞壁的三层。标尺，A：50m；B：500nm。图片修改自Cell Wall GenomicsLittle Is Known For cell wall formation:About 15%of the plant genome,possibly 3000 5000 genes.Only a small part of them characterized细胞壁纤维素的合成细胞壁纤维素的合成Cellulose Fibe

3、r StructurePear,et al.1996,PNAS,93:12637-12642From microbial cellulose to plant cellulose synthase gene (CelA-CesA)Acetobacter xylinum is Natures most prolific cellulose-producing bacterium.Arioli,T.et al.1998.Science 279:717-720 Identification of CesA genes in primary wall formationTaylor,et al.(19

4、99)Plant Cell 11:769-779 Identification of CesA genes in secondary wall formationCesA gene superfamily in Plants:Cellulose synthase(CesA);Cellulose synthase-like(Csl)Proposed CesA gene structure Kimura,et al.1999.Plant Cell 11:2075-2086.Observation of Cellulose synthase complex-“rosette”complexes.Pa

5、redez,et al.,(2006)Science 312,1491“Rosette”complex moving on the track of microtubules 纤维素由纤维素合酶复合体（纤维素由纤维素合酶复合体（CSC）合成）合成CesA-(1,4)-(1,4)-Glucan chain“rosette”complexcomplex subunitIn Arabidopsis:3 CesAs in primary cell wall 3 CesAs in secondary cell wallNature Materials(2003)LeafShoot TipPhloem X

6、ylemPtiCesA1-BPtiCesA1-BPtiCesA6-DPtiCesA4PtiCesA6-CPtiCesA6-APtiCesA7-APtiCesA8-APtiCesA6-BPtiCesA1-APtiCesA3-APtiCesA3-CPtiCesA3-BPtiCesA6-FPtiCesA7-BPtiCesA8-BPtCslA1PtCslA2PtCslA3PtCslA5PtCslC1PtCslC2PtCslC3PtCslC4PtCslC5PtCslD1PtCslD2PtCslD4PtCslD6PtCslD8PtCslD9PtCslD10PtCslE1PtCslE2PtCslE3PtCs

7、lG4PtCslG50.0200.0400.0600.0800.01000.01200.0杨树中至少杨树中至少6个个CesA基因参与次生壁纤维素合成基因参与次生壁纤维素合成(x104 copies/g total RNA)In Populus 18 CesAs30 Csls identified In Arabidopsis 10 CesA31 Csls identified6 CesA genes expressed in secondary cell wall formationcomplex subunit?CesAs 如何形成如何形成CSC？Precipitating Antibodi

8、es纤维素合酶复合体分离纤维素合酶复合体分离Immunobloting Antibodiescomplex subunitAnti-PtiCesA7-AAnti-PtiCesA1-AAnti-PtiCesA3-CAnti-PtiCesA8-BPre-immuneIgGCSC type CesA nameGene model Type IPtiCesA4eugene3.00002636PtiCesA7-AestExt_Genewise1_v1.C_LG_VI2188PtiCesA7-Bgw1.XVIII.3152.1PtiCesA8-Agw1.XI.3218.1PtiCesA8-Beugene3

9、.00040363Type IIPtiCesA1-AestExt_fgenesh4_pm.C_LG_XVIII0125PtiCesA1-Bfgenesh4_pg.C_LG_VI001789PtiCesA3-CestExt_fgenesh4_pg.C_LG_IX0979PtiCesA3-DestExt_Genewise1_v1.C_LG_I1792PtiCesA6-Efgenesh4_pm.C_LG_XIII000084PtiCesA6-Ffgenesh4_pg.C_scaffold_133000012CSCs 的蛋白组学鉴定的蛋白组学鉴定实验结果(b)(a)(c)(d)实验结果Anti-Pti

10、CesA7-AcaCesA7A in the secondary cell wall formation of differentiating xylem cells实验结果Anti-PtiCesA1-AcaCesA-1A in both the secondary and primary wall formation of differentiating xylem CesA constituents of two type CSCsPtiCesA7-APtiCesA7-BPtiCesA8-BPtiCesA8-APtiCesA4PtiCesA3-CPtiCesA3-DPtiCesA1-APt

11、iCesA6-EPtiCesA6-FType I complexType II complexRaven et al.2005 微纤丝纤维素的结构微纤丝纤维素的结构可能与可能与CSC调控相关调控相关Song,et al.New Phytologist，2010木质素的合成木质素的合成How is lignin synthesized?G“condensed lignin”difficult to break downS“relax lignin”,relatively easy to breakdownS/G,different in plants(G in softwood S/G hard

12、wood)43%efficiency improvement of lignin breakdown per S/G ratio unit increaseCH2OHOHOCH3CH2OHOHH3COOCH3CH2OHOHp-coumarylalcoholConiferylalcoholSinapyl alcoholG ligninH ligninS ligninPhenylalanine5-Hydroxyferulate 11OHOOHOHOHOHOOCH3OHOHOH3COOCH3OHOHOOCH3OHOHOOHSinapate 124-Coumarate 1Caffeate 2Ferul

13、ate 10OSHOOCH3OHCoA5-HydroxyferuloylCoA 14OSOCH3OHCoAH3COSinapoylCoA 15OSOCH3OHCoAFeruloylCoA 4OSOHOHCoACaffeoylCoA 3OSOHCoA4-CoumaroylCoA 13HHOOCH3OHO5-Hydroxyconifer-aldehyde 7OHOCH3OHH3COSinap-aldehyde 8OHOCH3OHConifer-aldehyde 5OHOHOHCaffe-aldehyde 17OHOH4-Coumar-aldehyde 16OCH3OHOHH3COSinapylal

14、cohol 9Syringyl monolignolOCH3OHOHConiferylalcohol 6Guaiacyl monolignolOCH3OHOHHO5-Hydroxyconiferylalcohol 20OHOHOHCaffeylalcohol 19OHOH4-Coumarylalcohol 18Hydroxyphenyl monolignolOHOPALC4HCinnamateC3H?OMT F5H OMT 4CL 4 CL 4CL 4CL 4CL?C3H?CoAOMT F5H?CoAOMT CCR CCR?CCR CCR?CCR CAD CAD?CAD CAD?CAD?Gua

15、iacyl-syringyl lignin GSSSSSGGGSSGGGSSSSSSSPossible Monolignol Biosynthesis PathwaysLi et al.,1997,PNAS,94:5493-5466;Li et al.,2006,Critical Review in Plant Science.25:213-233.Li et al.,1999,PMB,40:555-563;Hu et al.,1999,Nat.Biotechnol.,17:808-812Li et al.,2000,JBC,275:6537-6545;Osakabe et al.,1999,

16、PNAS,96:8955-8960;Li et al.,2005,PC&P 46:1234-1245 Li et al.,2001,the Plant Cell,13:1567-1585OHOHOOCH3OHHOOCH3OHHOOCH3OHHOOCH3OHOHOHOOCH3OHHOOCH3OHHOOCH3OHHOOCH3OH5-Hydroxyferulate 11OHOH4-Coumar-aldehyde 16OHOOH4-Coumarate 1OHOOH4-Coumarate 1OHOOHOHCaffeate 2OHOOCH3OHFerulate 10OHOOCH3OHFerulate 10

17、HOOCH3OHFerulate 10HOOCH3OHFerulate 10OHOH3COOCH3OHSinapate 12HOH3COOHSinapate 12OHOH3COOCH3OHSinapate 12HOH3COOHSinapate 12OSOHCoA4-CoumaroylCoA 13OSOHOHCoACaffeoylCoA 3OSOCH3OHCoAFeruloylCoA 4OSHOOCH3OHCoA5-HydroxyferuloylCoA 14OSOCH3OHCoAH3COSinapoylCoA 15OHOCH3OHH3COSinap-aldehyde 8OHOHOHCaffe-a

18、ldehyde 17OHOCH3OHConifer-aldehyde 5HHOOCH3OHO5-Hydroxyconifer-aldehyde 7OCH3OHOHH3COSinapylalcohol 9S-MonolignolOHOH4-Coumarylalcohol 18OHOHOHCaffeylalcohol 19OCH3OHOHConiferylalcohol 6G-monolignolOCH3OHOHHO5-Hydroxyconiferylalcohol 20PhenylalanineOHOPALCinnamateC4HCald5HAldOMTCoAOMT4CLCCRCADSADHou

19、 and Li,2011,J I P B Lu et al.,2010.GM Crops宋东亮等，宋东亮等，2010，植物生理通讯，植物生理通讯沈君辉等，沈君辉等，2010，植物生理通讯，植物生理通讯 宋东亮等，宋东亮等，2008，植物生理通讯植物生理通讯Horvath et al.,2010,Forest Products Horvath et al.,2010,Wood ResearchHorvath et al.,2010,IAWA JSuzuki et al.,2010,J.Wood Sci Lu et al.,2008,J Exp BotNakatsubo et al.,2007,C

20、ellulose Chem Technol Kasal et al.,2007,HolzforschungCell wall materialPretreatment hydrolysis fermentation Ethanol purification Lignin modification for biofuel production 0 02020404060608080100100120120140140Enzyme consumptionHydrolysisefficiencyConcentration of ethanolChange(%)ControlTransgenic-33

21、%+21%+30%Hydroxyphenyl monolignolLignin Biosynthesis Pathways in DicotsSyringyl monolignolGuaiacyl monolignol C3H?COMT F5H COMT CCR CAD CAD CAD?SADFeruloyl CoA Caffeoyl CoA 4-Coumaroyl CoA SOOCH3OHCoAOSOHOHCoAOSOHCoA5-Hydroxyconifer-aldehyde Sinap-aldehyde Conifer-aldehyde Caffe-aldehyde 4-Coumar-al

22、dehyde OHOHHHOOCH3OHOOHOCH3OHH3COOHOCH3OHOHOHOHSinapylalcohol Coniferylalcohol 5-Hydroxyconiferylalcohol 4-Coumarylalcohol OCH3OHOH H3COOCH3OHOHOCH3OHOHHOOHOHC4H5-Hydroxyferulate Sinapate 4-Coumarate Caffeate Ferulate OHOHOOCH3OHO HO H3COOCH3OHOHOOCH3OHOHO O HOOHOHCinnamate NH2 O HOPhenylalaninePALH

23、CT/C3HOMTOHOOH4CLCCRCCoAOMTF5HRice 4CLs form a different group from those of dicotsPtr:Populus trichocarpa,At:Arabidopsis thaliana Os:Oryza Japonica,Pp:Physcomitrella patens (Moss)0123456783 Weeks5 Weeks10 WeeksRelative expressionRootStemLeafSheathSpike4CL500.511.522.53Relative expression3 Weeks5 We

24、eks10 Weeks4CL1RootStemLeafSheathSpike00.10.20.30.40.50.60.70.80.93 Weeks5 Weeks10 WeeksRelative expression4CL4RootStemLeafSheathSpike00.050.10.150.20.250.30.353 Weeks5 Weeks10 WeeksRelative expression4CL2RootStemLeafSheathSpike0204060801001203 Weeks5 Weeks10 WeeksRelative expressionRootStemLeafShea

25、thSpike4CL3Expression of 4CLs in Various Tissues during Development Catalytic Properties of the Os4CLs0200040006000CIp-CACAFASAKenz(M-1S-1)4CL502004006008001000Kenz(M-1S-1)CIp-CACAFASA4CL104000800012000Kenz(M-1S-1)CIp-CACAFASA4CL40400080001200016000CIp-CACAFASAKenz(M-1S-1)4CL202000040000600008000010

26、0000CIp-CACAFASAKenz(M-1S-1)4CL3Morphological Change of 4CL Transgenic Rice(T3)Suppression of lignin biosynthesis in rice 4CL3AS transgenic plantsMonolignol(%)HGSWT1070204CL3AS87517Monolignol composition in rice Absorbance at 310nmWTTransgenicsp-CoumarateFerulate2x35S:Anti-4CL3 2x35S:4CL3oxFerulatep

27、-Coumarate10 15 20 25 30/10 15 20 25 30 Phenolic Compounds in Rice Stem4CLHCT/C3H Syringyl monolignolGuaiacyl monolignolHydroxyphenyl monolignol COMT F5H COMT 4CL CCR CCR?CCR CCR?CCR CAD CAD?CAD CAD?SAD 5-Hydroxyferuloyl CoA SinapoylCoA FeruloylCoA CaffeoylCoA 4-CoumaroylCoA OSHOOCH3OHCoAOSOCH3OH Co

28、A H3COSOOCH3OHCoAOSOHOHCoAOSOHCoA5-Hydroxyconifer-aldehyde Sinap-aldehyde Conifer-aldehyde Caffe-aldehyde 4-Coumar-aldehyde OHOHHHOOCH3OHOOHOCH3OHH3COOHOCH3OHOHOHOHSinapylalcohol Coniferylalcohol 5-Hydroxyconiferylalcohol Caffeylalcohol 4-Coumarylalcohol OCH3OHOH H3COOCH3OHOHOCH3OHOHHOOHOHOHOHOH C4H

29、5-Hydroxyferulate Sinapate 4-Coumarate Caffeate Ferulate OHOHOOCH3OHO HO H3COOCH3OHOHOOCH3OHOHO O HOOHOHOHOCinnamate NH2 O HOPhenylalaninePAL CCoAOMT CAld5H AldOMT?COMT F5H?COMTPossible 4CL-Mediated Pathways in RiceC3H?Gui et al，2011 Plant Physiology图12-2 拟南芥花序轴茎横切示不同类型细胞及其分布。不同细胞类型不同细胞类型-形成不同结结构和组成

30、的细胞壁？形成不同结结构和组成的细胞壁？不同结构与组成的细胞壁不同结构与组成的细胞壁-决定细胞的不同分化？决定细胞的不同分化？Cell elongation and wall thickening Cell elongationWall thickeningCoordination?实验结果Secondary growth-A system of wall thickening differentiationSong et al.2010,New Phytol.Song et al.2011,Plant Mol.Biol.Xu et al,2013,BMC GenomicsPlasma mem

31、brane proteomics of the wall thickening cells Cell WallPM?TransporterRLKCSC：10 CesAsPtrMAN6 localized in expanding vessels of xylemPtrMAN6 localized in plasma mebraneActivity assay of the recombinant PtrMAN6&PtrMAN4 in E.coliActivity of recombinant PtrMAN6 in plantN-glycosylation required for PtrMAN

32、6 activityCK Endo HN1 N2 N3 N4 N5Glutamine(Q)Asparagine(N)Multiple sites of N-glycosylation on PtrMAN6GGM is a main substrate of PtrMAN6 Stable oligosaccharides generated from PtrMAN6 activity Dimerization of PtrMAN6 required for the activity Protein from E.coliProtein from plantCysteineAlanineDimer

33、ization of PtrMAN6 through S-S linkages 4 5 6PtrMAN6LeMAN4aPtrMAN6LeMAN4aPtrMAN6LeMAN4aPtrMAN6LeMAN4aPtrMAN6LeMAN4aSecondary xylem differentiation is regulated by PtrMAN6PtMAN6 regulated both cell expansion and wall thickening Plants RhamnoseFucoseArabinoseXyloseMannoseGalactoseGlucoseWT6.590.131.66

34、0.05 13.350.93216.937.0117.450.8713.340.4337.471.04PtrMAN6ASLine55.600.20*1.220.04*6.910.59*280.557.32*19.870.76*9.590.41*34.691.12PtrMAN6ASLine65.190.19*1.160.03*6.560.38*288.675.23*20.190.14*8.850.37*34.440.66*PtrMAN6OELine89.720.13*3.20.07*35.960.96*165.691.83*11.830.28*31.530.61*45.940.87*PtrMAN

35、6OELine38.780.15*2.690.04*24.560.60*178.645.29*11.320.29*22.200.49*38.321.12AIR:Alcohol-insoluble residues;Value:mean(g mg-1 AIR)SE of 4 replicates.Single asterisk and double asterisks represent P 0.05 and P 0.01,respectively,in Students t test.Cell wall sugar composition regulated by PtrMAN6Wall th

36、ickening transcription program regulated by PtrMAN6Xylem differentiation treated by GGMOs MAN regulation likely mediated by GGMO signal moleculesCK GGMOsA proposed model for MAN function in regulation of SCW formationZhao,et al.2013,Plant JournalPtrWND1B occurs alternative splicingThe PtrWND1B homol

37、og also occurs AS in Eucalyptus Immunolocalization of PtrWND1B in fiber cellCa:Cambium;Ve:Vessel;Xf:Xylary fiber;Ra:Ray cell;Pf:Phloem fiberAlternative splicing of PtrWND1B is sequence-dependentPtrWND1B has a unique intron 2 sequenceAlternative splicing events of PtrWND1B were mediated by its intron

38、 2Suppression of PtrWND1B inhibited fiber secondary wall thickeningWND1B-RNAiWTXylem Phloem*NST GroupVND GroupDownsteam genesSuppressed expression of PtrWND1B inhibited fiber secondary wall thickening in PopulusWT PtrWND1B-sOE PtrWND1B-lOEXylemPhloemPtrWND1B-s and PtrWND1B-l played antagonistic roles in fiber SCW formation*PtrWND1B-s and PtrWND1B-l played antagonistic roles in fiber SCW formationA possible regulation mechanism of PtrWND1B Fiber cell wall thickening PtrWND1BAS1B-l1B-sWNDs and MYBs TFs1B-s1B-s1B-s1B-lWNDs and MYBs TFs