Functional genetics of barley stamen maturation

Unravelling the molecular mechanisms of stamen maturation in cereals is necessary to advance crop breeding

 

In flowering plants, proper stamen and pistil development are essential for successful plant reproduction and critical for crop yield. Although there is a large body of work on molecular factors of stamen development in the model plants Arabidopsis, rice and maize, such knowledge remains limited for the important temperate crops barley and wheat. We focus on the molecular mechanisms of stamen maturation in barley, which encompasses post-meiotic pollen development, anther opening and filament elongation. Because plant hormones are involved in some of these processes, our studies also aim at uncovering general principles of hormone biology that remain unresolved, such as how specific cell responses are achieved and which cellular functions activated by hormone signalling ultimately drive plant development. Importantly, a deep understanding of stamen maturation will provide ideal targets to effectively control the timing of male fertility in crops, potentially revolutionizing crop breeding through hybrid seed production. This is one of the promising solutions to the challenges of food security but remains costly and inefficient in temperate cereal crops. We are also interested in understanding the effects of and tolerance mechanisms to abiotic stress during barley stamen maturation.

Our research program combines approaches on genetics, genomics, development, physiology, histology, biological imaging, and quantitative analyses. We use the historic collection of barley male sterile genetic (msg) mutants to identify and characterize factors necessary for stamen maturation, following our established three-tier approach. First, we describe stamen and pollen development in msg mutants with morphological, histological and electron microscopy analyses. Second, we identify causal mutations with RNA or genome sequencing and confirm these mutations with new mutant alleles generated by gene editing with the CRISPR-Cas9 system. Third, we apply a battery of functional genetics analysis to understand the roles of MSG factors in stamen maturation. These include transcriptomics; metabolomics; spatiotemporal analysis of gene expression with qRT-PCR and in situ hybridization, or of protein expression with fluorescence reporters in stably transformed plants. A game-changer for these analyses has been our mastering of barley transformation in “Golden Promise Fast”, a Ppd-H1 introgression line that removes all the bottlenecks traditionally associated with this technology in barley. This line grows more robustly (no special conditions required), produces transformable embryos when plants are 8 or 9 weeks old, provides material to study stamen maturation in 5-7 weeks and completes its life cycle in 3 months.

 

 

Using this approach, we have discovered that the MSG38 gene encodes an auxin biosynthesis enzyme specifically expressed in pollen grains, which autonomously produce auxin to drive their own maturation. Our transcriptomic and metabolomic evidence indicates that auxin is required for a boost in energy production pathways, which enables timely starch accumulation, a hallmark of pollen maturity in cereals. These findings, recently published in Current Biology (https://doi.org/10.1016/j.cub.2022.02.073), reveal a concrete cellular output of auxin signaling, unveil a previously unrecognized function of plant hormones in controlling metabolic reprogramming during plant development, and establish cereal pollen as a simplified model to study the local effects of auxin synthesis and signaling.

Male reproductive development in cereals is extremely sensitive to plant abiotic stress. We are part of two different research consortia where we use some aspects of our three-tier approach to study how drought or heat stresses affect stamen maturation. These initiatives are the DFG-funded PhD School GRK2064 on drought based at the University of Bonn (https://www.grk2064.uni-bonn.de/de), and a BMBF-funded project on heat stress that is part of GENDIBAR, a multidisciplinary consortium within the Partnership on Research and Innovation in the Mediterranean Area (PRIMA), an initiative of the European Union (https://www.gendibar.com/).

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