Research Group Ragni
Periderm Development and Wood Formation

Secondary growth, the increase in girth of plant organs contributed to the large success of seed plants and shaped the earth’s biosphere as we know it today. Secondary growth is mainly driven by the vascular cambium, which produces daughter cells that inward will differentiate into xylem (wood) and outward into phloem (bast). In both perennial dicots and gymnosperms, wood is the principal site of biomass accumulation, and in addition to its economic value in forestry industry provides a means for the durable sequestration of CO₂. During secondary growth, the periderm replaces the epidermis as the outermost protective tissue. In stems, branches and roots of most dicots and gymnosperm, the periderm acts as the first line of defense that protects the plant against biotic and abiotic stresses limiting gas exchanges, water/solute loss and pathogen penetration.

The periderm, similar to the vascular system, comprises three tissues: the phellogen/cork cambium (a post-embryonic meristem), whose stem cells differentiate on the inside into the phelloderm and outside into the phellem/cork. The barrier property of the periderm is conferred by the cork, which is lignified and suberized and thus impermeable. Cork chemistry has been mainly studied in potato and cork oak due to its economic relevance, impacting potato conservation/production and the quality of cork in its application as wine-stoppers and insulating/building materials. For instance, a variety of potato, resistant to common scab, displays more periderm layers and increased suberin deposition in the periderm. In spite of the economical and agronomical importance, the molecular networks underlying periderm establishment and cork differentiation during plant development and in responses to stresses are largely unknown.

Our goal is to shed light on the molecular mechanisms regulating periderm growth and cork differentiation in a changing environment. We aim to understand the molecular switch that triggers cork cambium initiation and how nutrient status, biotic and abiotic stress regulate this switch. We aim to elucidate the regulatory network underlying cork-suberin and cork-lignin depositions in standard and stress conditions and how cork cambium activity and vascular cambium activity are interconnected and modulated by the environment. In the long term, our research will pave the way for obtaining plants, which are resistant to heat, drought and pathogens and helps breeding programs for biomass/wood improvement.

            The strength of our approach is that we have already developed a framework and a suite of tools (protocols for life imaging and whole root suberin pattern quantification, automated cell quantification software, reporter lines for each cell type at different developmental stages, rhizothrones to analyze roots grown on soil) to study periderm formation in the root of the amenable model plant Arabidopsis thaliana (Barbier de Reuille and Ragni 2017; Wunderling et al 2018; Campilho et al 2020; Serra et al. 2022). Thus, we can address fundamental questions of periderm growth, suberin genetics, physiology and biotic interactions using state of the art techniques, such as single cell sequencing and quantitative life imaging.

In Arabidopsis, the periderm is formed in the hypocotyl and in the root. In roots as in the majority of plants, the periderm arises from the pericycle, a single celled tissue layer surrounding the central vascular stele. To become the periderm three processes have to be coordinated: 1) reacquisition of pluripotency and specification of stem cell fate from pericycle into cork cambium stem cells 2) stem cell differentiation to form a multi-layered tissue, and 3) the shedding or death of the surrounding tissues as the pericycle, an inner tissue surrounded by the endodermis, cortex and epidermis, has to become the outer protective tissue. Thus, periderm development can be summarized in 5 characteristic stages, that take in consideration the fate of the surrounding tissues and periderm ontogenesis (Wunderling et al 2018; Serra et al 2022).

The phytohormone auxin controls many developmental processes including primary root growth, lateral root formation and cambium initiation/maintenance.We recently showed that auxin is also a key regulator of periderm development  (Xiao et al. 2020). Briefly,  auxin accumulates in the cork cambium and auxin treatment promotes periderm formation. By specifically blocking auxin signaling in the periderm, we showed that auxin is required for cork cambium initiation (from the pericycle) via AUXIN RESPONSE FACTOR 5/MONOPTEROS (ARF5/MP), ARF8 and other ARFs. Moreover, WOX4 and BP, two known vascular cambium regulators, act downstream of auxin to promote cork cambium formation revealing the first periderm regulatory hub. We are currently characterizing the targets downstream of auxin signaling, taking advantage of resources, in which auxin signaling in the periderm can be inducibly blocked, in precisely defined spatiotemporal contexts.

The periderm is not the only tissue that arises from the pericycle: lateral roots and partially, the vascular cambium also originate from the pericycle, raising the question on how auxin/output specificity is achieved and how different cell identities are established from the same tissue. The cork cambium regulatory network only partially overlaps with the auxin-induced lateral root program, which mainly act via ARF7 ARF19, LBD16 and GATA23 suggesting that different sets of TFs trigger different pericycle outputs (Xiao et al. 2020). Moreover, we demonstrated that while lateral root formation is the default auxin-induced pericycle output (after the plant body is established), auxin is not sufficient to trigger periderm formation in young seedlings as plants become competent to form a periderm with age, indicating that also developmental cues are important for pericycle output specificity (Xiao et al. 2020). As in mutants that lack lateral roots, both vascular and cork cambium are initiated earlier and only species with massive vascular cambial activity form a periderm, we hypothesized that vascular cambium formation is the developmental cue that triggers periderm initiation. Indeed, in transgenic plants with reduced vascular cambial activity, we observed delayed periderm development even upon auxin application, while ectopic induction of vascular cambium led to precocious periderm growth, showing that vascular cambium initiation is a prerequisite to trigger the periderm auxin-induced program(Xiao et al. 2020).