A genetic screen for plant mutants with altered nodulation phenotypes in response to rhizobial glycan mutants

Abstract

Nodule primordia induced by rhizobial glycan mutants often remain uninfected. To identify processes involved in infection and organogenesis we used forward genetics to identify plant genes involved in perception and responses to bacterial glycans. To dissect the mechanisms underlying the negative plant responses to the Mesorhizobium loti R7AexoU and ML001cep mutants, a screen for genetic suppressors of the nodulation phenotypes was performed on a chemically mutagenized Lotus population. Two mutant lines formed infected nitrogen-fixing pink nodules, while five mutant lines developed uninfected large white nodules, presumably altered in processes controlling organogenesis. Genetic mapping identified a mutation in the cytokinin receptor Lhk1 resulting in an alanine to valine substitution adjacent to a coiled-coil motif in the juxta-membrane region of LHK1. This results in a spontaneous nodulation phenotype and increased ethylene production. The allele was renamed snf5, and segregation studies of snf5 together with complementation studies suggest that snf5 is a gain-of-function allele. This forward genetic approach to investigate the role of glycans in the pathway synchronizing infection and organogenesis shows that a combination of plant and bacterial genetics opens new possibilities to study glycan responses in plants as well as identification of mutant alleles affecting nodule organogenesis.

Nitrate restricts nodule organogenesis through inhibition of cytokinin biosynthesis in Lotus japonicus

“…Synthesis of trans-Zeatin by LjCyp735a is induced during nodule development, but knock-out mutants do not display a nodulation phenotype in L. japonicus 8 . Stimulation of cytokinin signalling through biosynthesis and receptor activation is also sufficient to trigger nodule development, even in the absence of rhizobia 7 , 8 , 15 , 16 . This cytokinin signalling regulates expression of central components of nodulation signalling, including the transcription factor NIN (Nodule Inception) 17 via distal cis -elements in the Nin promoter 18 .…”

A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules

“…The major plant hormones auxin and cytokinin have important roles in both nodule organogenesis and infection (Tirichine et al, 2006, 2007; Murray et al, 2007; Suzaki et al, 2012; Breakspear et al, 2014; Reid et al, 2017; Liu et al, 2018; Nadzieja et al, 2018) and we therefore focused our tool-box development on these two hormones. Transcriptional (promoter based) markers have traditionally been used, however present problems associated with gene silencing (Zürcher et al, 2013).…”

Natural variation identifies a Pxy gene controlling vascular organisation and formation of nodules and lateral roots in Lotus japonicus

“…PXY suppresses ethylene signalling, and several ERF transcription factors showed increased expression in Arabidopsis pxy mutants (Etchells et al ., 2012). Ethylene, auxin and cytokinin are all involved in nodule organogenesis (Lin et al ., 2020), and hyperactivation of cytokinin can induce spontaneous nodule formation without rhizobia, which in turn facilitate infection (Tirichine et al ., 2007; Madsen et al ., 2010; Liu et al ., 2018a). Allelic differences in Pxy , and in other genes associated with the exoU QTL reported here, could therefore promote exoU nodulation through modulation of hormone signalling.…”