Root genomics: towards digital in situ hybridization

Separation of cell types and developmental stages in the Arabidopsis root and subsequent expression profiling have yielded a valuable dataset that can be used to select candidate genes for detailed study and to start probing the complexities of gene regulation in plant development

Cell-by-cell dissection of phloem development links a maturation gradient to cell specialization

Publisher Copyright: Copyright © 2021 The Authors, some rights reserved;In the plant meristem, tissue-wide maturation gradients are coordinated with specialized cell networks to establish various developmental phases required for indeterminate growth. Here, we used single-cell transcriptomics to reconstruct the protophloem developmental trajectory from the birth of cell progenitors to terminal differentiation in the Arabidopsis thaliana root. PHLOEM EARLY DNA-BINDING-WITH-ONE-FINGER (PEAR) transcription factors mediate lineage bifurcation by activating guanosine triphosphatase signaling and prime a transcriptional differentiation program. This program is initially repressed by a meristem-wide gradient of PLETHORA transcription factors. Only the dissipation of PLETHORA gradient permits activation of the differentiation program that involves mutual inhibition of early versus late meristem regulators. Thus, for phloem development, broad maturation gradients interface with cell-type-specific transcriptional regulators to stage cellular differentiation.Peer reviewe

Identification of factors required for meristem function in Arabidopsis using a novel next generation sequencing fast forward genetics approach

<p>Abstract</p> <p>Background</p> <p>Phenotype-driven forward genetic experiments are powerful approaches for linking phenotypes to genomic elements but they still involve a laborious positional cloning process. Although sequencing of complete genomes now becomes available, discriminating causal mutations from the enormous amounts of background variation remains a major challenge.</p> <p>Method</p> <p>To improve this, we developed a universal two-step approach, named 'fast forward genetics', which combines traditional bulk segregant techniques with targeted genomic enrichment and next-generation sequencing technology</p> <p>Results</p> <p>As a proof of principle we successfully applied this approach to two Arabidopsis mutants and identified a novel factor required for stem cell activity.</p> <p>Conclusion</p> <p>We demonstrated that the 'fast forward genetics' procedure efficiently identifies a small number of testable candidate mutations. As the approach is independent of genome size, it can be applied to any model system of interest. Furthermore, we show that experiments can be multiplexed and easily scaled for the identification of multiple individual mutants in a single sequencing run.</p

Probing the roles of LRR RLK genes in Arabidopsis thaliana roots using a custom T-DNA insertion set

Leucine-rich repeat receptor-like protein kinases (LRR RLKs) represent the largest group of Arabidopsis RLKs with approximately 235 members. A minority of these LRR RLKs have been assigned to diverse roles in development, pathogen resistance and hormone perception. Using a reverse genetics approach, a collection of homozygous T-DNA insertion lines for 69 root expressed LRR RLK genes was screened for root developmental defects and altered response after exposure to environmental, hormonal/chemical and abiotic stress. The obtained data demonstrate that LRR RLKs play a role in a wide variety of signal transduction pathways related to hormone and abiotic stress responses. The described collection of T-DNA insertion mutants provides a valuable tool for future research into the function of LRR RLK genes

Arabidopsis Root Development

In plant development, the basic body plan is laid down during embryogenesis. Development carries on postembryogenically above and below ground with the continuous formation and outgrowth of lateral organs shaping the adult plant. In the past two decades, molecular genetics has been the preferred approach to study Arabidopsis thaliana root development. These efforts have resulted in the identification of numerous genes, involved in as many regulatory processes of root growth and development. Incidentally, conserved mechanisms and genetic factors that act in root and shoot growth have been uncovered, revealing general principles of plant development. Transport-mediated graded distribution of the phytohormone auxin, for example, acts as a global organizer that is locally translated into distinct cellular responses by specific auxin/indole-3-acetic acid-AUXIN RESPONSE FACTOR pairs. In the root, these responses promote expression of the PLETHORA regulators that act dose-dependently in controlling root morphology. The stem cell niche is uniquely defined by the combinatorial activity of the PLETHORA and SHORT-ROOT/SCARECROW transcription factors yet deploy signaling mechanisms that are conserved in root and shoot stem cell maintenance. Perpetual divisions of the stem cells are tightly regulated, interconnecting epigenetic factors, hormonal control and core cell cycle components. In this chapter, we will focus on recent advances in our understanding of Arabidopsis root development. Taking embryogenesis as a starting point, we will describe the genes and mechanisms involved in root meristem and stem cell patterning and maintenance

Induction of 35S-PLT2-GR in whole seedlings

Organ formation in animals and plants relies on precise control of cell state transitions to turn stem cell daughters into fully differentiated cells. In plants, cells cannot rearrange due to shared cell walls. Thus, differentiation progression and the accompanying cell expansion must be tightly coordinated. PLETHORA (PLT) transcription factor gradients were shown to guide the progression of cell differentiation at different positions in the Arabidopsis root. While well-described transcription factor gradients in animals specify distinct cell fates within an essentially static context, the PLT gradient is unique in its ability to control cell differentiation in a growing organ during continuous production and expansion of cells. To understand the output of these gradients we studied the gene set transcriptionally controlled by PLTs. Our work reveals how the PLT gradient regulates cell state by region-specific induction of cell proliferation genes and repression of differentiation. Moreover, PLT targets include major patterning genes and autoregulatory feedback components, reinforcing their role as master regulators of organ development

A dialogue between generations

Arabidopsis embryonic root development involves the formation of distinct cell types and tissues in a tightly regulated and thereby highly predictable spatio-temporal manner. A crosstalk between maternal and embryonic genes orchestrates division orientation and fate specification to control root development

Induction of pPLT2-PLT2-GR in Quiescent Center (QC) cells

Organ formation in animals and plants relies on precise control of cell state transitions to turn stem cell daughters into fully differentiated cells. In plants, cells cannot rearrange due to shared cell walls. Thus, differentiation progression and the accompanying cell expansion must be tightly coordinated. PLETHORA (PLT) transcription factor gradients were shown to guide the progression of cell differentiation at different positions in the Arabidopsis root. While well-described transcription factor gradients in animals specify distinct cell fates within an essentially static context, the PLT gradient is unique in its ability to control cell differentiation in a growing organ during continuous production and expansion of cells. To understand the output of these gradients we studied the gene set transcriptionally controlled by PLTs. Our work reveals how the PLT gradient regulates cell state by region-specific induction of cell proliferation genes and repression of differentiation. Moreover, PLT targets include major patterning genes and autoregulatory feedback components, reinforcing their role as master regulators of organ development

Induction of pPLT2-PLT2-GR in Quiescent Center (QC) cells

Organ formation in animals and plants relies on precise control of cell state transitions to turn stem cell daughters into fully differentiated cells. In plants, cells cannot rearrange due to shared cell walls. Thus, differentiation progression and the accompanying cell expansion must be tightly coordinated. PLETHORA (PLT) transcription factor gradients were shown to guide the progression of cell differentiation at different positions in the Arabidopsis root. While well-described transcription factor gradients in animals specify distinct cell fates within an essentially static context, the PLT gradient is unique in its ability to control cell differentiation in a growing organ during continuous production and expansion of cells. To understand the output of these gradients we studied the gene set transcriptionally controlled by PLTs. Our work reveals how the PLT gradient regulates cell state by region-specific induction of cell proliferation genes and repression of differentiation. Moreover, PLT targets include major patterning genes and autoregulatory feedback components, reinforcing their role as master regulators of organ development

Induction of PLT1, PLT3, PLT4, PLT5 and PLT7 in whole seedlings

Organ formation in animals and plants relies on precise control of cell state transitions to turn stem cell daughters into fully differentiated cells. In plants, cells cannot rearrange due to shared cell walls. Thus, differentiation progression and the accompanying cell expansion must be tightly coordinated. PLETHORA (PLT) transcription factor gradients were shown to guide the progression of cell differentiation at different positions in the Arabidopsis root. While well-described transcription factor gradients in animals specify distinct cell fates within an essentially static context, the PLT gradient is unique in its ability to control cell differentiation in a growing organ during continuous production and expansion of cells. To understand the output of these gradients we studied the gene set transcriptionally controlled by PLTs. Our work reveals how the PLT gradient regulates cell state by region-specific induction of cell proliferation genes and repression of differentiation. Moreover, PLT targets include major patterning genes and autoregulatory feedback components, reinforcing their role as master regulators of organ development