Imaging with the fluorogenic dye Basic Fuchsin reveals subcellular patterning and ecotype variation of lignification in Brachypodium distachyon

Nikki Kapp; William J Barnes; Tom L Richard; Charles T Anderson

doi:10.1093/jxb/erv158

Imaging with the fluorogenic dye Basic Fuchsin reveals subcellular patterning and ecotype variation of lignification in Brachypodium distachyon

J Exp Bot. 2015 Jul;66(14):4295-304. doi: 10.1093/jxb/erv158. Epub 2015 Apr 28.

Authors

Nikki Kapp¹, William J Barnes², Tom L Richard³, Charles T Anderson⁴

Affiliations

¹ Center for Lignocellulose Structure and Formation, The Pennsylvania State University, University Park, PA 16802, USA Department of Biology, The Pennsylvania State University, University Park, PA 16802, USA Department of Agricultural and Biological Engineering, The Pennsylvania State University, University Park, PA 16802, USA.
² Department of Biology, The Pennsylvania State University, University Park, PA 16802, USA.
³ Center for Lignocellulose Structure and Formation, The Pennsylvania State University, University Park, PA 16802, USA Department of Agricultural and Biological Engineering, The Pennsylvania State University, University Park, PA 16802, USA.
⁴ Center for Lignocellulose Structure and Formation, The Pennsylvania State University, University Park, PA 16802, USA Department of Biology, The Pennsylvania State University, University Park, PA 16802, USA cta3@psu.edu.

Abstract

Lignin is a complex polyphenolic heteropolymer that is abundant in the secondary cell walls of plants and functions in growth and defence. It is also a major barrier to the deconstruction of plant biomass for bioenergy production, but the spatiotemporal details of how lignin is deposited in actively lignifying tissues and the precise relationships between wall lignification in different cell types and developmental events, such as flowering, are incompletely understood. Here, the lignin-detecting fluorogenic dye, Basic Fuchsin, was adapted to enable comparative fluorescence-based imaging of lignin in the basal internodes of three Brachypodium distachyon ecotypes that display divergent flowering times. It was found that the extent and intensity of Basic Fuchsin fluorescence increase over time in the Bd21-3 ecotype, that Basic Fuchsin staining is more widespread and intense in 4-week-old Bd21-3 and Adi-10 basal internodes than in Bd1-1 internodes, and that Basic Fuchsin staining reveals subcellular patterns of lignin in vascular and interfascicular fibre cell walls. Basic Fuchsin fluorescence did not correlate with lignin quantification by acetyl bromide analysis, indicating that whole-plant and subcellular lignin analyses provide distinct information about the extent and patterns of lignification in B. distachyon. Finally, it was found that flowering time correlated with a transient increase in total lignin, but did not correlate strongly with the patterning of stem lignification, suggesting that additional developmental pathways might regulate secondary wall formation in grasses. This study provides a new comparative tool for imaging lignin in plants and helps inform our views of how lignification proceeds in grasses.

Keywords: Acetyl bromide soluble lignin; Basic Fuchsin; Brachypodium distachyon; Wiesner staining.; ecotype; flowering time; fluorescence imaging; lignin.

Publication types

Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Brachypodium / chemistry*
Fluorescent Dyes / chemistry*
Lignin / chemistry*
Subcellular Fractions / chemistry*

Substances

Fluorescent Dyes
Lignin