Gene regulatory networks for lignin biosynthesis in switchgrass <i>(Panicum virgatum</i>)

<jats:title>Summary</jats:title><jats:p>Cell wall recalcitrance is the major challenge to improving saccharification efficiency in converting lignocellulose into biofuels. However, information regarding the transcriptional regulation of secondary cell wall biogenesis remains poor in switchgrass (<jats:italic>Panicum virgatum</jats:italic>), which has been selected as a biofuel crop in the United States. In this study, we present a combination of computational and experimental approaches to develop gene regulatory networks for lignin formation in switchgrass. To screen transcription factors (<jats:styled-content style="fixed-case">TF</jats:styled-content>s) involved in lignin biosynthesis, we developed a modified method to perform co‐expression network analysis using 14 lignin biosynthesis genes as bait (target) genes. The switchgrass lignin co‐expression network was further extended by adding 14 <jats:styled-content style="fixed-case">TF</jats:styled-content>s identified in this study, and seven <jats:styled-content style="fixed-case">TF</jats:styled-content>s identified in previous studies, as bait genes. Six <jats:styled-content style="fixed-case">TF</jats:styled-content>s (Pv<jats:styled-content style="fixed-case">MYB</jats:styled-content>58/63, Pv<jats:styled-content style="fixed-case">MYB</jats:styled-content>42/85, Pv<jats:styled-content style="fixed-case">MYB</jats:styled-content>4, Pv<jats:styled-content style="fixed-case">WRKY</jats:styled-content>12, Pv<jats:styled-content style="fixed-case">SND</jats:styled-content>2 and Pv<jats:styled-content style="fixed-case">SWN</jats:styled-content>2) were targeted to generate overexpressing and/or down‐regulated transgenic switchgrass lines. The alteration of lignin content, cell wall composition and/or plant growth in the transgenic plants supported the role of the <jats:styled-content style="fixed-case">TF</jats:styled-content>s in controlling secondary wall formation. <jats:styled-content style="fixed-case">RNA</jats:styled-content>‐seq analysis of four of the transgenic switchgrass lines revealed downstream target genes of the secondary wall‐related <jats:styled-content style="fixed-case">TF</jats:styled-content>s and crosstalk with other biological pathways. <jats:italic>In vitro</jats:italic> transactivation assays further confirmed the regulation of specific lignin pathway genes by four of the <jats:styled-content style="fixed-case">TF</jats:styled-content>s. Our meta‐analysis provides a hierarchical network of <jats:styled-content style="fixed-case">TF</jats:styled-content>s and their potential target genes for future manipulation of secondary cell wall formation for lignin modification in switchgrass.</jats:p>