A network of CLAVATA receptors buffers auxin-dependent meristem maintenance.

Plant body plans are elaborated in response to both environmental and endogenous cues. How these inputs intersect to promote growth and development remains poorly understood. During reproductive development, central zone stem cell proliferation in inflorescence meristems is negatively regulated by the CLAVATA3 (CLV3) peptide signalling pathway. In contrast, floral primordia formation on meristem flanks requires the hormone auxin. Here we show that CLV3 signalling is also necessary for auxin-dependent floral primordia generation and that this function is partially masked by both inflorescence fasciation and heat-induced auxin biosynthesis. Stem cell regulation by CLAVATA signalling is separable from primordia formation but is also sensitized to temperature and auxin levels. In addition, we uncover a novel role for the CLV3 receptor CLAVATA1 in auxin-dependent meristem maintenance in cooler environments. As such, CLV3 signalling buffers multiple auxin-dependent shoot processes across divergent thermal environments, with opposing effects on cell proliferation in different meristem regions.

BAM1/2 receptor kinase signaling drives CLE peptide-mediated formative cell divisions in Arabidopsis roots.

Cell division is often regulated by extracellular signaling networks to ensure correct patterning during development. In Arabidopsis, the SHORT-ROOT (SHR)/SCARECROW (SCR) transcription factor dimer activates CYCLIND6;1 (CYCD6;1) to drive formative divisions during root ground tissue development. Here, we show plasma-membrane-localized BARELY ANY MERISTEM1/2 (BAM1/2) family receptor kinases are required for SHR-dependent formative divisions and CYCD6;1 expression, but not SHR-dependent ground tissue specification. Root-enriched CLE ligands bind the BAM1 extracellular domain and are necessary and sufficient to activate SHR-mediated divisions and CYCD6;1 expression. Correspondingly, BAM-CLE signaling contributes to the restriction of formative divisions to the distal root region. Additionally, genetic analysis reveals that BAM-CLE and SHR converge to regulate additional cell divisions outside of the ground tissues. Our work identifies an extracellular signaling pathway regulating formative root divisions and provides a framework to explore this pathway in patterning and evolution.

A PXY-Mediated Transcriptional Network Integrates Signaling Mechanisms to Control Vascular Development in Arabidopsis.

The cambium and procambium generate the majority of biomass in vascular plants. These meristems constitute a bifacial stem cell population from which xylem and phloem are specified on opposing sides by positional signals. The PHLOEM INTERCALATED WITH XYLEM (PXY) receptor kinase promotes vascular cell division and organization. However, how these functions are specified and integrated is unknown. Here, we mapped a putative PXY-mediated transcriptional regulatory network comprising 690 transcription factor-promoter interactions in Arabidopsis (Arabidopsis thaliana). Among these interactions was a feedforward loop containing transcription factors WUSCHEL HOMEOBOX RELATED14 (WOX14) and TARGET OF MONOPTEROS6 (TMO6), each of which regulates the expression of the gene encoding a third transcription factor, LATERAL ORGAN BOUNDARIES DOMAIN4 (LBD4). PXY signaling in turn regulates the WOX14, TMO6, and LBD4 feedforward loop to control vascular proliferation. Genetic interaction between LBD4 and PXY suggests that LBD4 marks the phloem-procambium boundary, thus defining the shape of the vascular bundle. These data collectively support a mechanism that influences the recruitment of cells into the phloem lineage, and they define the role of PXY signaling in this context in determining the arrangement of vascular tissue.

Evolution of buffering in a genetic circuit controlling plant stem cell proliferation.

Precise control of plant stem cell proliferation is necessary for the continuous and reproducible development of plant organs1,2. The peptide ligand CLAVATA3 (CLV3) and its receptor protein kinase CLAVATA1 (CLV1) maintain stem cell homeostasis within a deeply conserved negative feedback circuit1,2. In Arabidopsis, CLV1 paralogs also contribute to homeostasis, by compensating for the loss of CLV1 through transcriptional upregulation3. Here, we show that compensation4,5 operates in diverse lineages for both ligands and receptors, but while the core CLV signaling module is conserved, compensation mechanisms have diversified. Transcriptional compensation between ligand paralogs operates in tomato, facilitated by an ancient gene duplication that impacted the domestication of fruit size. In contrast, we found little evidence for transcriptional compensation between ligands in Arabidopsis and maize, and receptor compensation differs between tomato and Arabidopsis. Our findings show that compensation among ligand and receptor paralogs is critical for stem cell homeostasis, but that diverse genetic mechanisms buffer conserved developmental programs.

Modulation of Asymmetric Division Diversity through Cytokinin and SPEECHLESS Regulatory Interactions in the Arabidopsis Stomatal Lineage.

Coordinated growth of organs requires communication among cells within and between tissues. In plants, leaf growth is largely dictated by the epidermis; here, asymmetric and self-renewing divisions of the stomatal lineage create two essential cell types-pavement cells and guard cells-in proportions reflecting inputs from local, systemic, and environmental cues. The transcription factor SPEECHLESS (SPCH) is the prime regulator of divisions, but whether and how it is influenced by external cues to provide flexible development is enigmatic. Here, we show that the phytohormone cytokinin (CK) can act as an endogenous signal to affect the extent and types of stomatal lineage divisions and forms a regulatory circuit with SPCH. Local domains of low CK signaling are created by SPCH-dependent cell-type-specific activity of two repressive type-A ARABIDOPSIS RESPONSE REGULATORs (ARRs), ARR16 and ARR17, and two secreted peptides, CLE9 and CLE10, which, together with SPCH, can customize epidermal cell-type composition.

Cutting Edge Genetics: CRISPR/Cas9 Editing of Plant Genomes.

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 (Cas9) system is a genome editing technology transforming the field of plant biology by virtue of the system's efficiency and specificity. The system has quickly evolved for many diverse applications including multiplex gene mutation, gene replacement and transcriptional control. As CRISPR/Cas9 is increasingly applied to plants, it is becoming clear that each component of the system can be modified to improve editing results. This review aims to highlight common considerations and options when conducting CRISPR/Cas9 experiments.

CRISPR/Cas9-mediated resistance to cauliflower mosaic virus.

Viral diseases are a leading cause of worldwide yield losses in crop production. Breeding of resistance genes (R gene) into elite crop cultivars has been the standard and most cost-effective practice. However, R gene-mediated resistance is limited by the available R genes within genetic resources and in many cases, by strain specificity. Therefore, it is important to generate new and broad-spectrum antiviral strategies. The CRISPR-Cas9 (clustered regularly interspaced palindromic repeat, CRISPR-associated) editing system has been employed to confer resistance to human viruses and several plant single-stranded DNA geminiviruses, pointing out the possible application of the CRISPR-Cas9 system for virus control. Here, we demonstrate that strong viral resistance to cauliflower mosaic virus (CaMV), a pararetrovirus with a double-stranded DNA genome, can be achieved through Cas9-mediated multiplex targeting of the viral coat protein sequence. We further show that small interfering RNAs (siRNA) are produced and mostly map to the 3' end of single-guide RNAs (sgRNA), although very low levels of siRNAs map to the spacer region as well. However, these siRNAs are not responsible for the inhibited CaMV infection because there is no resistance if Cas9 is not present. We have also observed edited viruses in systematically infected leaves in some transgenic plants, with short deletions or insertions consistent with Cas9-induced DNA breaks at the sgRNA target sites in coat protein coding sequence. These edited coat proteins, in most cases, led to earlier translation stop and thus, nonfunctional coat proteins. We also recovered wild-type CP sequence in these infected transgenic plants, suggesting these edited viral genomes were packaged by wild-type coat proteins. Our data demonstrate that the CRISPR-Cas9 system can be used for virus control against plant pararetroviruses with further modifications.

Modification of the Mycotoxin Deoxynivalenol Using Microorganisms Isolated from Environmental Samples.

The trichothecene mycotoxin deoxynivalenol (DON) is a common contaminant of wheat, barley, and maize. New strategies are needed to reduce or eliminate DON in feed and food products. Microorganisms from plant and soil samples collected in Blacksburg, VA, USA, were screened by incubation in a mineral salt media containing 100 µg/mL DON and analysis by gas chromatography mass spectrometry (GC/MS). Two mixed cultures derived from soil samples consistently decreased DON levels in assays using DON as the sole carbon source. Nuclear magnetic resonance (NMR) analysis indicated that 3-keto-4-deoxynivalenol was the major by-product of DON. Via 16S rRNA sequencing, these mixed cultures, including mostly members of the genera Acinetobacter, Leadbetterella, and Gemmata, were revealed. Incubation of one of these mixed cultures with wheat samples naturally contaminated with 7.1 µg/mL DON indicated nearly complete conversion of DON to the less toxic 3-epimer-DON (3-epi-DON). Our work extends previous studies that have demonstrated the potential for bioprospecting for microorganisms from the environment to remediate or modify mycotoxins for commercial applications, such as the reduction of mycotoxins in fuel ethanol co-products.

Ready, aim, shoot: stem cell regulation of the shoot apical meristem.

Plant shoot meristems contain stem cells that are continuously renewed to replenish cells that exit and differentiate during lateral organ formation. Complex cell-to-cell signaling systems balance division and differentiation. These center on ligand-receptor networks, hormone pathways, and transcriptional regulators that function in an integrated manner. In this review, we aim to highlight new findings in shoot stem cell regulation across species.