Elucidation of Xylem-Specific Transcription Factors and Absolute Quantification of Enzymes Regulating Cellulose Biosynthesis in Populus trichocarpa.
Cellulose, the main chemical polymer of wood, is the most abundant polysaccharide in nature.1 The ability to perturb the abundance and structure of cellulose microfibrils is of critical importance to the pulp and paper industry as well as for the textile, wood products, and liquid biofuels industries. Although much has been learned at the transcript level about the biosynthesis of cellulose, a quantitative understanding at the proteome level has yet to be established. The study described herein sought to identify the proteins directly involved in cellulose biosynthesis during wood formation in Populus trichocarpa along with known xylem-specific transcription factors involved in regulating these key proteins. Development of an effective discovery proteomic strategy through a combination of subcellular fractionation of stem differentiating xylem tissue (SDX) with recently optimized FASP digestion protocols, StageTip fractionation, as well as optimized instrument parameters for global proteomic analysis using the quadrupole-orbitrap mass spectrometer resulted in the deepest proteomic coverage of SDX protein from P. trichocarpa with 9,146 protein groups being identified (1% FDR). Of these, 20 cellulosic/hemicellulosic enzymes and 43 xylem-specific transcription factor groups were identified. Finally, selection of surrogate peptides led to an assay for absolute quantification of 14 cellulosic proteins in SDX of P. trichocarpa.
W.M. Keck FTMS Laboratory, Department of Chemistry, North Carolina State University , Raleigh, North Carolina 27695, United States.
Philip L
PL
Loziuk
W.M. Keck FTMS Laboratory, Department of Chemistry, North Carolina State University , Raleigh, North Carolina 27695, United States.
Jennifer
J
Parker
Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University , Raleigh, North Carolina 27695, United States.
Wei
W
Li
Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University , Raleigh, North Carolina 27695, United States.
Chien-Yuan
CY
Lin
Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University , Raleigh, North Carolina 27695, United States.
Jack P
JP
Wang
State Key Laboratory of Tree Genetics and Breeding, Chinese Academy of Forestry , Beijing 100091, China.
Quanzi
Q
Li
Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University , Raleigh, North Carolina 27695, United States.
Ronald R
RR
Sederoff
Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University , Raleigh, North Carolina 27695, United States.
Vincent L
VL
Chiang
W.M. Keck FTMS Laboratory, Department of Chemistry, North Carolina State University , Raleigh, North Carolina 27695, United States.
David C
DC
Muddiman
D050260:Carbohydrate Metabolism; D002482:Cellulose / Q000096:biosynthesis* / Q000235:genetics; D002853:Chromatography, Liquid; D018506:Gene Expression Regulation, Plant; D058977:Molecular Sequence Annotation; D010940:Plant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D032107:Populus / Q000235:genetics* / Q000378:metabolism; D020543:Proteome / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D040901:Proteomics; D053719:Tandem Mass Spectrometry; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D014934:Wood / Q000737:chemistry / Q000378:metabolism*; D052584:Xylem / Q000235:genetics / Q000378:metabolism
D010940:Plant Proteins; D020543:Proteome; D014157:Transcription Factors; D002482:Cellulose
PC-IDMS; SRM; absolute quantification; cellulose biosynthesis; shotgun discovery proteomics; targeted mass spectrometry; transcription factor