Switchgrass Contains Two Cinnamyl Alcohol Dehydrogenases Involved in Lignin Formation
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Lignin content of switchgrass (Panicum virgatum L.), a bioenergy species, is a critical determinant of biomass quality since it can negatively impact conversion of biomass into liquid fuels via biochemical platforms. Cinnamyl alcohol dehydrogenase (CAD) is a key enzyme in lignin biosynthesis. Here, we have shown that cv. Kanlow switchgrass contains at least two closely related CAD genes (PviCAD1 and PviCAD2) that code for proteins containing highly conserved domains and residues that identify them as bona fide CADs. Both recombinant proteins displayed substrate kinetics consistent with their presumed role in cell wall lignification. Proteomic and immunoblotting detected CAD containing spots in internode protein extracts, and proteomic analyses demonstrated that both CADs were expressed. In planta CAD activity, CAD protein levels were observed at all stages of tiller development. A real-time qPCR analysis of the two CADs and one CAD-like sequence indicated that transcripts coding for PviCAD1 were present in greater abundance than those coding for PviCAD2. Transcripts for a third CAD-like sequence (PviAroADH) were present at intermediate levels as compared to PviCAD1 and CAD2. The predicted protein sequence of PviAroADH indicated that it was an enzyme unrelated to lignification based on phylogenetic and protein modeling data.