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In wounded Arabidopsis thaliana leaves, four 13S-lipoxygenases (AtLOX2, AtLOX3, AtLOX4, AtLOX6) act in a hierarchical manner to contribute to the jasmonate burst. This leads to defense responses with LOX2 playing an important role in plant resistance against caterpillar herb-ivory. In this study, we sought to characterize the impact of AtLOX2 on wound-induced phytohormonal and transcriptional responses to foliar mechanical damage using wildtype (WT) and lox2 mutant plants. Compared with WT, the lox2 mutant had higher constitutive levels of the phytohormone salicylic acid (SA) and enhanced expression of SA-responsive genes. This suggests that AtLOX2 may be involved in the biosynthesis of jasmonates that are involved in the antagonism of SA biosynthesis. As expected, the jasmonate burst in response to wounding was dampened in lox2 plants. Generally, 1 h after wounding, genes linked to jasmonate biosynthesis, jasmonate signaling attenuation and abscisic acid-responsive genes, which are primarily involved in wound sealing and healing, were differentially regulated between WT and lox2 mutants. Twelve h after wounding, WT plants showed stronger expression of genes associated with plant protection against insect herbivory. This study highlights the dynamic nature of jasmonate-responsive gene expression and the contribution of AtLOX2 to this pathway and plant resistance against insects.

Summary: Adrenal cortex and gonads represent the two major steroidogenic organs in mammals. Both tissues are considered to share a common developmental origin characterized by the expression of Nr5a1/Sf1. The precise origin of adrenogonadal progenitors and the processes driving differentiation toward the adrenal or gonadal fate remain, however, elusive. Here, we provide a comprehensive single-cell transcriptomic atlas of early mouse adrenogonadal development including 52 cell types belonging to twelve major cell lineages. Trajectory reconstruction reveals that adrenogonadal cells emerge from the lateral plate rather than the intermediate mesoderm. Surprisingly, we find that gonadal and adrenal fates have already diverged prior to Nr5a1 expression. Finally, lineage separation into gonadal and adrenal fates involves canonical versus non-canonical Wnt signaling and differential expression of Hox patterning genes. Thus, our study provides important insights into the molecular programs of adrenal and gonadal fate choice and will be a valuable resource for further research into adrenogonadal ontogenesis.

In general, caterpillar herbivores with a narrow host preference (specialists) have evolved mechanisms to circumvent specific plant defenses. In contrast, caterpillars with a broader host range (generalists) may manipulate phytohormone pathways common to many plant species to attenuate induced defenses. Many studies have compared plant responses to specialist versus generalist caterpillars. In contrast, this study evaluates the induced response of Arabidopsis thaliana to two generalist caterpillar species, the cabbage looper, Trichoplusia ni, and the beet armyworm, Spodoptera exigua. Although both caterpillars are considered generalists, S. exigua has a broader plant host range, whereas T. ni prefers Brassicaceous plants. Our study shows that most responses to caterpillar herbivory, such as the jasmonate burst, are similar in plants attacked by either insect species; however, we do observe dynamic and temporal differences in specific responses. Expression of AtZAT10, a 12-oxo-phytodienoic acid-responsive gene, is only induced in response to T. ni damage. In comparison, only S. exigua herbivory activates the salicylic acid/NPR1-dependent pathway, as observed by the expression of the marker gene AtPR1. Even though both species induce AtPDF1.2 expression, we found caterpillar-specific temporal differences: T. ni herbivory results in sustained expression over time, whereas gene expression is sharply downregulated at 36 h in S. exigua-attacked plants. Although damage by these two caterpillar species induced AtMYB28 and AtMYB34 expression, specific short- and long-chain aliphatic and indolic glucosinolates accumulate only in response to S. exigua herbivory. These species-specific, plant-induced responses likely reflect differences in effectors found in caterpillar oral secretions.[Figure: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Retinoic acid (RA) is an essential signaling molecule for cardiac development and plays a protective role in the heart after myocardial infarction (MI). In both cases, the effect of RA signaling on cardiomyocytes, the principle cell type of the heart, has been reported to be indirect. Here we have developed an inducible murine transgenic RA-reporter line using CreERT2 technology that permits lineage tracing of RA-responsive cells and faithfully recapitulates endogenous RA activity in multiple organs during embryonic development. Strikingly, we have observed a direct RA response in cardiomyocytes during mid-late gestation and after MI. Ablation of RA signaling through deletion of the Aldh1a1/a2/a3 genes encoding RA-synthesizing enzymes leads to increased cardiomyocyte apoptosis in adults subjected to MI. RNA sequencing analysis reveals Tgm2 and Ace1, two genes with well-established links to cardiac repair, as potential targets of RA signaling in primary cardiomyocytes, thereby providing novel links between the RA pathway and heart disease.

Programmed cell death is an important part of tissue development, and traditionally it is considered that neuronal death is a stochastic process in response to neurotrophic factor deprivation. Here the authors show that for TrkC+ proprioreceptors, which neurons die is predetermined molecularly by how much TrkC is present, as well as by a gene expression signature.

In response to physiological demand, the pituitary gland generates new hormone-secreting cells from committed progenitor cells throughout life. It remains unclear to what extent pituitary stem cells (PSCs), which uniquely express SOX2, contribute to pituitary growth and renewal. Moreover, neither the signals that drive proliferation nor their sources have been elucidated. We have used genetic approaches in the mouse, showing that the WNT pathway is essential for proliferation of all lineages in the gland. We reveal that SOX2+ stem cells are a key source of WNT ligands. By blocking secretion of WNTs from SOX2+ PSCs in vivo, we demonstrate that proliferation of neighbouring committed progenitor cells declines, demonstrating that progenitor multiplication depends on the paracrine WNT secretion from SOX2+ PSCs. Our results indicate that stem cells can hold additional roles in tissue expansion and homeostasis, acting as paracrine signalling centres to coordinate the proliferation of neighbouring cells.

Coordinated development of muscles, tendons, and their attachment sites ensures emergence of functional musculoskeletal units that are adapted to diverse anatomical demands among different species. How these different tissues are patterned and functionally assembled during embryogenesis is poorly understood. Here, we investigated the morphogenesis of extraocular muscles (EOMs), an evolutionary conserved cranial muscle group that is crucial for the coordinated movement of the eyeballs and for visual acuity. By means of lineage analysis, we redefined the cellular origins of periocular connective tissues interacting with the EOMs, which do not arise exclusively from neural crest mesenchyme as previously thought. Using 3D imaging approaches, we established an integrative blueprint for the EOM functional unit. By doing so, we identified a developmental time window in which individual EOMs emerge from a unique muscle anlage and establish insertions in the sclera, which sets these muscles apart from classical muscle-to-bone type of insertions. Further, we demonstrate that the eyeballs are a source of diffusible all-trans retinoic acid (ATRA) that allow their targeting by the EOMs in a temporal and dose-dependent manner. Using genetically modified mice and inhibitor treatments, we find that endogenous local variations in the concentration of retinoids contribute to the establishment of tendon condensations and attachment sites that precede the initiation of muscle patterning. Collectively, our results highlight how global and site-specific programs are deployed for the assembly of muscle functional units with precise definition of muscle shapes and topographical wiring of their tendon attachments.

During kidney development, WNT/β-catenin signalling has to be tightly controlled to ensure proliferation and differentiation of nephron progenitor cells. Here, we show in mice that the signalling molecules RSPO1 and RSPO3 act in a functionally redundant manner to permit WNT/β-catenin signalling and their genetic deletion leads to a rapid decline of nephron progenitors. By contrast, tissue specific deletion in cap mesenchymal cells abolishes mesenchyme to epithelial transition (MET) that is linked to a loss of Bmp7 expression, absence of SMAD1/5 phosphorylation and a concomitant failure to activate Lef1, Fgf8 and Wnt4, thus explaining the observed phenotype on a molecular level. Surprisingly, the full knockout of LGR4/5/6, the cognate receptors of R-spondins, only mildly affects progenitor numbers, but does not interfere with MET. Taken together our data demonstrate key roles for R-spondins in permitting stem cell maintenance and differentiation and reveal Lgr-dependent and independent functions for these ligands during kidney formation.

Pheochromocytoma (PCC) and paraganglioma (PGL) are rare neuroendocrine tumors associated with high cardiovascular morbidity and variable risk of malignancy. The current therapy of choice is surgical resection. Nevertheless, PCCs/PGLs are associated with a lifelong risk of tumor persistence or recurrence. A high rate of germline or somatic mutations in numerous genes has been found in these tumors. For some, the tumorigenic processes are initiated during embryogenesis. Such tumors carry gene mutations leading to pseudohypoxic phenotypes and show more immature characteristics than other chromaffin cell tumors; they are also often multifocal or metastatic and occur at an early age, often during childhood. Cancer stem cells (CSCs) are cells with an inherent ability of self-renewal, de-differentiation, and capacity to initiate and maintain malignant tumor growth. Targeting CSCs to inhibit cancer progression has become an attractive anti-cancer therapeutic strategy. Despite progress for this strategy for solid tumors such as neuroblastoma, brain, breast, and colon cancers, no substantial advance has been made employing similar strategies in PCCs/PGLs. In the current review, we discuss findings related to the identification of normal chromaffin stem cells and CSCs, pathways involved in regulating the development of CSCs, and the importance of the stem cell niche in development and maintenance of CSCs in PCCs/PGLs. Additionally, we examine the development and feasibility of novel CSC-targeted therapeutic strategies aimed at eradicating especially recurrent and metastatic tumors.

Congenital abnormalities of the kidney and urinary tract (CAKUT) are a highly diverse group of diseases that together belong to the most common abnormalities detected in the new-born child. Consistent with this diversity, CAKUT are caused by mutations in a large number of genes and present a wide spectrum of phenotypes. In this review, we will focus on duplex kidneys, a relatively frequent form of CAKUT that is often asymptomatic but predisposes to vesicoureteral reflux and hydronephrosis. We will summarise the molecular programs responsible for ureter induction, review the genes that have been identified as risk factors in duplex kidney formation and discuss molecular and cellular mechanisms that may lead to this malformation.

The predicted future increase in tropospheric carbon dioxide (CO2) levels will have major effects on C3 plants and their interactions with other organisms in the biosphere. In response to attack by chewing arthropod herbivores or nectrotrophic pathogens, many plants mount a rapid and intense increase in jasmonate-related phytohormones that results in a robust defense response; however, previous studies have shown that C3 plants grown at elevated CO2 may have lower induced jasmonate levels, particularly in well nitrate-fertilized plants. Given the relationship between atmospheric CO2, photorespiration, cellular reductant and redox status, nitrogen assimilation and phytohormones, we compared wound-induced responses of the C3 plant Arabidopsis thaliana. These plants were fertilized at two different rates (1 or 10 mM) with nitrate or ammonium and grown at ambient or elevated CO2. In response to artificial wounding, an increase in cellular oxidative status leads to a strong increase in jasmonate phytohormones. At ambient CO2, increased oxidative state of nitrate-fertilized plants leads to a robust 7-iso-jasmonyl-L-isoleucine increase; however, the strong fertilizer rate-associated increase is alleviated in plants grown at elevated CO2. As well, the changes in ascorbate in response to wounding and wound-induced salicylic acid levels may also contribute to the suppression of the jasmonate burst. Understanding the mechanism underlying the attenuation of the jasmonate burst at elevated CO2 has important implications for fertilization strategies under future predicted climatic conditions.

Coronary arteries are essential to support the heart with oxygen, and coronary heart disease is one of the leading causes of death worldwide. The coronary arteries form at highly stereotyped locations and are derived from the primitive vascular plexus of the heart. How coronary arteries are remodeled and the signaling molecules that govern this process are poorly understood. Here, we have identified the Wnt-signaling modulator Rspo3 as a crucial regulator of coronary artery formation in the developing heart. Rspo3 is specifically expressed around the coronary stems at critical time points in their development. Temporal ablation of Rspo3 at E11.5 leads to decreased β-catenin signaling and a reduction in arterial-specific proliferation. As a result, the coronary stems are defective and the arterial tree does not form properly. These results identify a mechanism through which localized expression of RSPO3 induces proliferation of the coronary arteries at their stems and permits their formation.

Glucosinolates (GSLs) evolved in Brassicaceae as chemical defenses against herbivores. The GSL content in plants is affected by both abiotic and biotic factors, but also depends on the genetic background of the plant. Since the bitter taste of GSLs can be unfavorable for both livestock and human consumption, several plant varieties with low GSL seed or leaf content have been bred. Due to their lower GSL levels, such varieties can be more susceptible to herbivore pests. However, low GSL varieties may quickly increase GSL levels upon herbivore feeding by activating GSL biosynthesis, hydrolysis, or transporter genes. To analyze differences in herbivore-induced GSL responses in relation to constitutive GSL levels, we selected four Brassica rapa varieties, containing either low or high root GSL levels. Plants were infested either with Delia radicum or Delia floralis larvae. The larvae of both root flies are specialists on Brassica plants. Root samples were collected after 3, 5, and 7 days. We compared the effect of root herbivore damage on the expression of GSL biosynthesis (CYP79A1, CYP83B2), transporter (GTR1A2, GTR2A2), and GSL hydrolysis genes (PEN2, TGG2) in roots of low and high GSL varieties in conjugation with their GSL levels. We found that roots of high GSL varieties contained higher levels of aliphatic, indole, and benzyl GSLs than low GSL varieties. Infestation with D. radicum larvae led to upregulation of indole GSL synthesis genes in low and high GSL varieties. High GSL varieties showed no or later responses than low varieties to D. floralis herbivory. Low GSL varieties additionally upregulated the GSL transporter gene expression. Low GSL varieties did not show a stronger herbivore-induced response than high GSL varieties, which indicates that there is no trade-off between constitutive and induced GSLs.

Many adrenocortical diseases are more prevalent in women than in men, but the reasons underlying this sex bias are still unknown. Recent studies involving gonadectomy and sex hormone replacement experiments in mice have shed some light onto the molecular basis of sexual dimorphism in the adrenal cortex. Indeed, it has been shown that gonadal hormones influence many aspects of adrenal physiology, ranging from stem cell-dependent tissue turnover to steroidogenesis and X-zone dynamics. This article reviews current knowledge on adrenal cortex sexual dimorphism and the potential mechanisms underlying sex hormone influence of adrenal homeostasis. Both topics are expected to contribute to personalized and novel therapeutic approaches in the future.

The adrenal cortex undergoes functional zonation to generate an outer zona glomerulosa (ZG) and inner zona fasciculata (ZF), but how this is regulated at a molecular level is unclear. Here, the authors show that ZG differentiation is stimulated by WNT signalling and that PKA blocks WNT signalling to allow ZF differentiation and also prevents WNT-induced cancer development.

Liver zonation, the spatial separation of different metabolic pathways along the liver sinusoids, is fundamental for proper functioning of this organ, and its disruption can lead to the development of metabolic disorders such as hyperammonemia. Metabolic zonation involves the induction of β-catenin signaling around the central veins, but how this patterned activity is established and maintained is unclear. Here, we show that the signaling molecule Rspondin3 is specifically expressed within the endothelial compartment of the central vein. Conditional deletion of Rspo3 in mice disrupts activation of central fate, demonstrating its crucial role in determining and maintaining β-catenin-dependent zonation. Moreover, ectopic expression of Rspo1, a close family member of Rspo3, induces the expression of pericentral markers, demonstrating Rspondins to be sufficient to imprint a more central fate. Thus, Rspo3 is a key angiocrine factor that controls metabolic zonation of liver hepatocytes.

After acute kidney injury (AKI), surviving cells within the nephron proliferate and repair. We identify Sox9 as an acute epithelial stress response in renal regeneration. Translational profiling after AKI revealed a rapid upregulation of Sox9 within proximal tubule (PT) cells, the nephron cell type most vulnerable to AKI. Descendants of Sox9+ cells generate the bulk of the nephron during development and regenerate functional PT epithelium after AKI-induced reactivation of Sox9 after renal injury. After restoration of renal function post-AKI, persistent Sox9 expression highlights regions of unresolved damage within injured nephrons. Inactivation of Sox9 in PT cells pre-injury indicates that Sox9 is required for the normal course of post-AKI recovery. These findings link Sox9 to cell intrinsic mechanisms regulating development and repair of the mammalian nephron.

In mammals, male sex determination is governed by SRY-dependent activation of Sox9, whereas female development involves R-spondin1 (RSPO1), an activator of the WNT/beta-catenin signaling pathway. Genetic analyses in mice have demonstrated Sry and Sox9 to be both required and sufficient to induce testicular development. These genes are therefore considered as master regulators of the male pathway. Indeed, female-to-male sex reversal in XX Rspo1 mutant mice correlates with Sox9 expression, suggesting that this transcription factor induces testicular differentiation in pathological conditions. Unexpectedly, here we show that testicular differentiation can occur in XX mutants lacking both Rspo1 and Sox9 (referred to as XX Rspo1(KO)Sox9(cKO) ()), indicating that Sry and Sox9 are dispensable to induce female-to-male sex reversal. Molecular analyses show expression of both Sox8 and Sox10, suggesting that activation of Sox genes other than Sox9 can induce male differentiation in Rspo1(KO)Sox9(cKO) mice. Moreover, since testis development occurs in XY Rspo1(KO)Sox9(cKO) mice, our data show that Rspo1 is the main effector for male-to-female sex reversal in XY Sox9(cKO) mice. Thus, Rspo1 is an essential activator of ovarian development not only in normal situations, but also in sex reversal situations. Taken together these data demonstrate that both male and female sex differentiation is induced by distinct, active, genetic pathways. The dogma that considers female differentiation as a default pathway therefore needs to be definitively revised.

Vertebrate genes often play functionally distinct roles in different subsets of cells; however, tools to study the cell-specific function of gene products are poorly developed. Therefore, we have established a novel mouse model that enables the visualization and manipulation of defined subpopulations of neurons. To demonstrate the power of our system, we dissected genetic cascades in which Pax6 is central to control tangentially migrating neurons of the mouse brainstem. Several Pax6 downstream genes were identified and their function was analyzed by over-expression and knock-down experiments. One of these, Pou4f2, induces a prolonged midline arrest of growth cones to influence the proportion of ipsilaterally versus contralaterally settling neurons. These results demonstrate that our approach serves as a versatile tool to study the function of genes involved in cell migration, axonal pathfinding, and patterning processes. Our model will also serve as a general tool to specifically over-express any gene in a defined subpopulation of neurons and should easily be adapted to a wide range of applications.