Award details

de novo specification of a post-embryonic stem cell population in plants

Reference	BB/V008129/1
Principal Investigator / Supervisor	Dr Peter Etchells
Co-Investigators / Co-Supervisors
Institution	Durham University
Department	Biosciences
Funding type	Research
Value (£)	460,814
Status	Current
Type	Research Grant
Start date	09/08/2021
End date	08/08/2024
Duration	36 months

Abstract

Xylem and phloem are essential plant tissues responsible for the movement of water and nutrients. They arise from opposing sides of a bifacial stem cell population referred to as the cambium. The xylem constitutes the woody tissue, thus cambial divisions are responsible for generating the majority of plant biomass. In Arabidopsis, the cambium is formed post-embryonically within the vascular cylinder, in cells characterised by high levels of the phytohormone auxin, and elevated expression of homeodomain transcription factors. Events downstream of these regulators are poorly understood. However, in our preliminary data, we have shown that PXY and ER receptor kinases and their paralogues are essential cambium formation. Thus, experiments in this proposal will enable us to understand how these receptors function in this process. We aim to test whether PXY, ER, and their paralogues interact at the plasma membrane, determine in which cell types and at what time-points they are able to promote cambium formation, and we will define their transcriptional targets. We have already identified members of the PLETHORA family of transcription factors as transcriptional targets of PXY signalling, and will expand on this analysis, determining whether PXY and ER signalling can regulate each other at the transcriptional level. Plant development is entirely the consequence of differential growth which is characterised by only two factors, cell division, and changes to cell size. Despite this apparent limitation, higher plant body plans are remarkably complex. The experimental programme described here will contribute to our understanding of one mechanism by which plants generate this complexity, i.e. by generating de novo, a stem cell population post embryonically.

Summary

Plants draw carbon dioxide from the air and convert it to sugars. Much of the sugar is utilised in the most abundant biopolymer on Earth, cellulose. Cellulose is the main constituent of wood, and wood represents a renewable biomaterial and a source of bioenergy. This proposal aims to shed light on how woody tissue is formed. Wood is part of the plant vascular system and is made up of xylem cells. The xylem arises from a group of stem cells referred to as the cambium. Understanding of how the cambium functions could be used to increase wood formation in a forestry setting, but in these experiments we will use the model plant, Arabidopsis, because it is easy to manipulate in the lab. In both plants and animals, stem cells are typically formed in the embryo. The cambium is unusual because its stem cell population is formed after embryogenesis, making it an interesting system for understanding stem cell initiation. Previously we have identified six signalling proteins, so-called receptor kinases, which act together to promote the cell divisions necessary for cambium formation. However, while we know that they are important in this process, we don't know exactly how they work. In this proposal we aim to answer this question. The receptor kinases are localised to the plasma membrane in the cambium cells. One hypothesis is that protein-protein interactions occur between some of the six receptor kinases. We will test this hypothesis in two ways. In the first, we will tag the receptor kinases in question with different fluorescent proteins. We can determine if energy transfer between two different fluorescent proteins has occurred (referred to as FRET) using microscopy. Because energy transfer is only be possible if the two proteins are in very close proximity, we would only detect FRET if protein-protein interactions were occurring between the tagged receptor kinases. The second method that we will use is referred to as co-immunoprecipitation. Here we will use an antibody that targets a tag which we have placed on a receptor kinase. If the receptor kinase forms part of a protein complex we will be able to identify other parts of the complex. Receptor kinases are molecules that send signals to the cells on the membranes of which they are present. The six receptor kinases that we have identified are present on several cell types, but which cell type(s) are required for normal cambium initiation? Can these proteins initiate cambium in several cell types? We aim to address this question in our proposal. We have generated a mutant in which all six receptor kinases are missing. We propose to replace these proteins in specific cell types including the xylem and cambium, but other vascular cell types too. These experiments will tell us if the consequences of the receptor kinase signalling are restricted to the cell in which they are present (referred to as cell-autonomous), or if they influence the behaviour of neighbouring cells. Finally, we propose to determine what the outcome of the signalling is. Receptor kinases signal to the nucleus to alter gene activity, but we don't which genes respond to the receptor kinases that are the focus of this study. We propose to increase signalling through the receptor kinases and detect which genes respond by sequencing the mRNA before and after the signalling increase. Changes in mRNA levels will tell us which genes have been regulated. It will also tell us whether some of the receptor kinases can influence the levels at which the other five are expressed. Together, these experiments will tell us how the cambium functions. They will also provide information on how stem cell populations can be set up following embryogenesis. They may also suggest ways in which we can manipulate wood formation to increase forest productivity and improve carbon capture.

Committee	Research Committee B (Plants, microbes, food & sustainability)
Research Topics	Plant Science, Stem Cells
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search