The formation of seeds represented a remarkable innovation that affects plant evolutionary fitness. For humans, cereal grains have become essential for our dietary needs, for animal feeding and industrial processing. Current climate change and the associated increase in extreme weather events represent an additional constraint to agricultural practice and productivity.

Our general goal is to understand molecular control mechanisms of seed formation at the cell, tissue and whole-organ level and to establish novel approaches for improving seed yield and quality in cereal species, particular for barley and wheat, but also for other crop plants, like rice, rapeseed, sugar beet and sorghum.

We especially focus on uncovering gene regulatory networks by studying transcriptional regulators, plant hormone and reactive oxygen species (ROS) signalling cascades, and nutrient transport during grain filling. Aim of our work is to improve or stabilise the yield potential and quality of crop seeds under current and future climate conditions.

Alternation of generations and embryogenesis

The conquest of the land by plants is one of the most remarkable transitions of life on earth that altered the atmosphere and paved the way for the evolution of complex animals. Land plants (embryophytes) most likely evolved from charophycean freshwater green algae around 450 million years ago. Terrestrialization required major adaptations, including the development of mechanical support for the plant body, adaptation to a gaseous and relative dry environment, the formation of novel organs for the acquisition of water and nutrients, the ability to cope with novel abiotic and biotic stresses but also a reprogramming of the reproduction through the evolution of embryogenesis. In the frame of MAdLand DFG priority programme 2237 we study the role of ROS scavenging and producing enzymes in the mosses Marchantia polymorpha and Physcomitrium patens, which can both reproduce asexual and sexual, and thereby represent unique organisms for studying genes essential for embryogenesis.

In addition, we study selected transcription factors (TFs) that show embryo-developmental stage specific expression patterns in Arabidopsis. Based upon the gene regulatory networks (GRNs) that these thus far uncharacterised TFs control we can assign a biological function for them and specifically analyse their role during embryogenesis. Furthermore, our group is part of the BMBF funded AVATARS project and focusses on rapeseed embryogenesis during beneficial and detrimental conditions. Within this project, gene expression analysis of dissected rapeseed embryos will be performed and used for the modelling of gene regulatory networks

TCS signal transduction and regulatory mechanisms of tissue differentiation in cereal grains

Developing cereal grains are complex structures consisting of maternal and filial tissues. Sequences of cell proliferation, differentiation, maturation and disintegration occur in the different grain tissues in a coordinated manner. Since a long time, we are studying tissue-specific processes in cereal and legume seeds, and established laser capture microdissection (LCM)-based RNA-seq analysis for cereals (Brandt et al., 2018) and other species. Tissue-specific RNA-seq of developing barley endosperm transfer cells (ETCs) identified the Two-Component Signaling (TCS) system as essential signal transduction pathway for early endosperm specification and differentiation. Current functional genomics studies focus on the role of selected TCS elements on barley endosperm and grain development. Histidine kinase 1 (HvHK1) was identified as a receptor component with a unique expression in the syncytial endosperm domain at the maternal-filial boundary. Knock-down of HvHK1 by RNAi impairs transfer cell specification in the central ETC region, abolishes cell wall ingrowths and produces smaller grains with reduced starch accumulation (Hertig et al., 2020). RNA-seq of the altered cell type confirmed loss of transfer cell identity and gene regulatory network modelling predicted main regulatory links governing cellularization of ETCs. New models for modification of further TCS genes with potential functions in grain (and also spike) development have been established using the CRISPR/Cas9-technology in frame of a DFG project. Phenotypes of mutated plants will be characterised by genetic and histological analysis of the progenies (ongoing work). In addition, downstream transcription factors of the TCS pathways are functionally characterised.

Elucidation of gene regulatory networks in cereals using DAP-seq

Both the response of a plant to its environment and how plant organs grow to characteristic sizes and shapes is genetically controlled. Intriguingly, the cause behind improved plant traits is often due to altered gene regulatory networks. Transcription factors are key determinants in regulating growth and adaptation responses by modulating the activity of tens to hundreds of genes. In contrast to Arabidopsis thaliana, we virtually have no idea on the gene networks controlled by transcription factors in crops. Recently, we managed to setup DNA-affinity purification and sequencing (DAP-seq) in barley through an IPK-funded Flagship project. This technique allows us to yield insight into the biology and binding site architecture of numerous barley TFs. In addition, we use tissue-specific libraries to study the binding profiles of the selected TFs in their natural context. Currently we are optimizing the method for wheat and in the near future plant to adapt it for Sorghum bicolor.

Based upon the identified networks we select promising candidate TFs with a putative role in grain filling for functional analysis. Currently, several NAC and AP2/ERF transcription factors have been selected for which transgenic models will be established to validate our approach.

Regulation of cell proliferation, differentiation and signalling by reactive oxygen species

Reactive oxygen species (ROS) are inevitable byproducts of several metabolic processes or cascade of oxidation-reduction reactions that take place in aerobic organisms. Initially mainly recognised for their potential to induce damage to cells, it has become clear that plants actively produce and scavenge ROS to modulate cellular processes and trigger a ROS burst for stress signalling. Interestingly, in the apical regions of plants a ROS gradient between superoxide and hydrogen peroxide is observed, in which the first is linked to stemness and the second to differentiation.

Recently we uncovered a family of myb-like transcription factors that modulate cell expansion in plants. These KUODA (Chinese for enlarge) TFs are plant specific and conserved in all plant species, suggesting that they have a universal role in plants. Currently we investigate their role in sugar beet in collaboration with KWS. In addition, we are highly interested in other TFs that specifically regulate ROS-related genes during development and stress. In addition, we study ROS signalling during the initial phase of salt stress in rice in collaboration with the Plant Biotechnology group of the University of Bielefeld. Salt stress in rice provokes a ROS burst that triggers the activity of several transcriptional cascades. Currently we mainly focus on the SERF1 cascade and the branching of the ROS signal.

Role of protein stability during development

Wear and tear on proteins due to the environment in which they act and the biochemical processes they control, can result in misfolding, aggregation, or mistargeting. Therefore, cells monitor the state of each protein through either chaperones or components of diverse proteolytic pathways. Still, the largest fraction of the proteins that are removed through proteostasis systems are actually undamaged. Based on the demand for specific proteins during cellular processes, unwanted proteins are removed. For instance, during cell proliferation and cell differentiation, which are sequential processes that involve specific protein sets during the course of development. In addition, many proteins have a short half-life, which potentially minimises the risk of protein damage to prevent the accumulation of aberrant and damaging proteins. Protein turnover is performed mostly through the ubiquitin–proteasome system (UPS) and autophagy. Here we study the transcriptional regulation of the proteasome during development. In addition, we characterise labile transcription factors with a role in plant development and a need for ROS gradients.

Legend:

Barley HISTIDINE KINASE1 (HvHK1) coordinates endosperm transfer cell (ETC) specification in the young endosperm required for efficient grain filling. Gene regulatory network modeling predicted distinct phosphorelay and hormone signalling modules as main regulatory links that are directed by HvHK1.

Legende:

Gerste HISTIDINE KINASE1 (HvHK1) koordiniert die Spezifikation der Endosperm-Transferzellen (ETC) im jungen Endosperm, die für eine effiziente Kornfüllung erforderlich sind. Mit Hilfe der Modellierung genregulatorischer Netzwerke konnten verschiedene Phosphorelisierungs- und Hormon-Signalwege als wichtigste regulatorische Verbindungen von HvHK1 vorhergesagt werden.