Abstract:
Leaves are the key organs for plants to grow and flourish, as they allow efficient harvesting of the solar energy required for plant survival. Leaf organogenesis begins at the periphery of the shoot apical meristem (SAM), where local auxin maxima define the positions of incipient primordia. Yet, how auxin signaling translates into precisely orchestrated and robust developmental programs remained unclear.
To address this, we leveraged single-cell RNA-sequencing (scRNA-seq) to generate a high-resolution transcriptomic atlas of the Arabidopsis vegetative shoot apex. Contrary to the previously published atlas, we successfully captured rare SAM cells, which allowed targeted analysis of the early events in leaf organogenesis. A high-resolution leaf developmental trajectory describing the transcriptomic dynamics of leaf outgrowth revealed distinct phases of gene expression corresponding to meristematic, organ initiation/cell fate acquisition, and differentiation phases. A gene regulatory network (GRN) underlying the lateral organ founder cells within the initiation phase revealed a core auxin-responsive module of eight key transcriptional factors (TFs). Within this module, we showed that PLETHORA 3 (PLT3) and AUXIN RESPONSE TRANSCRIPTION FACTOR 3 (ARF3) operate in a positive feedback loop that plausibly facilitates rapid amplification of the initial auxin signal, leading to establishment of robust transcriptional output and consequent organ outgrowth coupled with patterning.
Contrary to the SAM, where stem cells give rise to organ founder cells that progressively acquire distinct cell fates, cell identities in the root are specified already within the stem cell niche. The tissue initials undergo several rounds of cell division before initiating the bespoke tissue maturation programs. The timing of the switch from proliferation to differentiation needs to be precisely controlled and coordinated across the cell files to allow correct root functioning. To investigate the mechanism that coordinates this transition, we combined single-cell transcriptomic and chromatin accessibility data and reconstructed developmental trajectories of the root ground tissue. Using this model, we described phased transcriptional programs governing the transition from stem cell to early differentiation, in which chromatin accessibility marks the activation of central developmental regulators. A major transcriptional switch in the late meristematic zone, underpinned by extensive chromatin remodeling, drives the transition from cell division to differentiation. Central to this transition are GATA TFs, which regulate both lineage-specific differentiation and general maturation. We propose a feedback-driven gene regulatory network in which GATA TFs, in interplay with PLT and ARR-B TFs, integrate inputs from auxin and cytokinin to balance proliferation and differentiation, ensuring temporal precision in root development at both the tissue and organ scales.
The single-cell data presented in this thesis hold a substantial potential for re-analysis from perspectives beyond those highlighted here. To facilitate this, we developed an intuitive online single-cell data browser for seamless data exploration. This platform allows a flexible analytical workspace accessible without a requirement for expertise in single-cell data analysis, thus providing access to the data to the broader research community.
RNA-seq