Depletion of UDP-D-apiose/UDP-D-xylose synthases results in rhamnogalacturonan-II deficiency, cell wall thickening, and cell death in higher plants.
D-apiose serves as the binding site for borate cross-linking of rhamnogalacturonan II (RG-II) in the plant cell wall, and biosynthesis of D-apiose involves UDP-D-apiose/UDP-D-xylose synthase catalyzing the conversion of UDP-D-glucuronate to a mixture of UDP-D-apiose and UDP-D-xylose. In this study we have analyzed the cellular effects of depletion of UDP-D-apiose/UDP-D-xylose synthases in plants by using virus-induced gene silencing (VIGS) of NbAXS1 in Nicotiana benthamiana. The recombinant NbAXS1 protein exhibited UDP-D-apiose/UDP-D-xylose synthase activity in vitro. The NbAXS1 gene was expressed in all major plant organs, and an NbAXS1-green fluorescent protein fusion protein was mostly localized in the cytosol. VIGS of NbAXS1 resulted in growth arrest and leaf yellowing. Microscopic studies of the leaf cells of the NbAXS1 VIGS lines revealed cell death symptoms including cell lysis and disintegration of cellular organelles and compartments. The cell death was accompanied by excessive formation of reactive oxygen species and by induction of various protease genes. Furthermore, abnormal wall structure of the affected cells was evident including excessive cell wall thickening and wall gaps. The mutant cell walls contained significantly reduced levels of D-apiose as well as 2-O-methyl-L-fucose and 2-O-methyl-D-xylose, which serve as markers for the RG-II side chains B and A, respectively. These results suggest that VIGS of NbAXS1 caused a severe deficiency in the major side chains of RG-II and that the growth defect and cell death was likely caused by structural alterations in RG-II due to a D-apiose deficiency.
Laboratory of Plant Genomics, Korea Research Institute of Bioscience and Biotechnology, Taejon 305-333, Korea.
Joon-Woo
JW
Ahn
Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut 06269-3125.
Rajeev
R
Verma
BioNanotechnology Center, Korea Research Institute of Bioscience and Biotechnology, Taejon 305-333, Korea.
Moonil
M
Kim
Department of Biology, Yonsei University, Seoul 120-749, Korea.
Jae-Yong
JY
Lee
Laboratory of Plant Genomics, Korea Research Institute of Bioscience and Biotechnology, Taejon 305-333, Korea.
Yu-Kyung
YK
Kim
Department of Biology, Chungnam National University, Taejon 305-764, Korea.
Jae-Wook
JW
Bang
Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut 06269-3125. Electronic address: wdreiter@uconn.edu.
Wolf-Dieter
WD
Reiter
Department of Biology, Yonsei University, Seoul 120-749, Korea,. Electronic address: hspai@yonsei.ac.kr.
Hyun-Sook
HS
Pai
D019731:Carbon-Nitrogen Ligases / Q000172:deficiency* / Q000378:metabolism; D016923:Cell Death; D002473:Cell Wall / Q000378:metabolism*; D020868:Gene Silencing; D008969:Molecular Sequence Data; D010368:Pectins / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology* / Q000201:enzymology* / Q000378:metabolism; D014539:Uridine Diphosphate Sugars / Q000378:metabolism*; D014994:Xylose / Q000378:metabolism*
D010940:Plant Proteins; C511616:UDP-apiose; D014539:Uridine Diphosphate Sugars; C042492:rhamnogalacturonan II; D010368:Pectins; D014994:Xylose; D019731:Carbon-Nitrogen Ligases