Biology subjects

Muroyama, A.

Publications and source records attributed to Muroyama, A..

6 recordsLinked to original sources

bioRxiv2025

A user-friendly machine-learning program to quantify stomatal features from fluorescence images

In nearly all plants, pores on the leaf surface called stomata are essential for photosynthesis and gas exchange. The shape and distribution of stomata on the leaf varies widely between plants and is directly connected to photosynthetic efficiency. However, our understanding of the factors, both genetic and environmental, that exert subtle but significant effects on stomatal morphology is limited by the time required to manually annotate stomata in large imaging datasets. Here, we present a lightweight and efficient tool, QuickSpotter, for semi-automated stomatal annotation from fluorescence images. First, we establish QuickSpotters ability to automatically and accurately annotate mature stomata across developmental time. We also introduce an optional, speedy proofreading utility, StomEdit, that allows the researcher to quickly validate and correct machine-generated annotations. We use QuickSpotter and StomEdit to quantify how stomatal morphology evolves at the population level during cotyledon development and demonstrate how the programs can be used to extract subtle differences in stomatal development following pharmacological treatments. Finally, we describe PairCaller, a pair-calling classifier that accompanies QuickSpotter and can be used to identify stomatal clusters, a physiologically relevant and widely studied developmental phenotype. Taken together, our suite of programs facilitates quantitative analyses of stomatal development at scale, enabling high-throughput analyses of leaf phenotypes under varied conditions.

plant biology↗

bioRxiv2025

Force-responsive symmetric cell divisions orient stomata along global tissue axes

Stomata, microscopic pores that regulate gas exchange in plants, are patterned according to conserved pathways that regulate their physiology. Here, we identify a new mode of stomatal patterning that depends on previously unrecognized regulation of the final symmetric cell division that creates paired guard cells. Symmetric cell divisions are aligned by tensile stress at both the subcellular and supracellular scales, creating a globally polarized stomatal field that tracks the major axes of tissue growth. By identifying KATANIN as a critical regulator of symmetric division orientation, we show that stress-based division orientation is required to prepattern stomatal morphology and pore creation. We find that expansion of neighboring cells non-autonomously controls symmetric division orientation, linking stomatal alignment to overall leaf shape. Finally, we show that polarized stomatal fields are widespread across plant genera and their species-specific alignment patterns are consistent with the force-based mechanism we identify in Arabidopsis. This force-responsive pathway provides a unifying model that explains long-standing observations of stomatal organization across species.

developmental biology↗

bioRxiv2022

Cortical polarity ensures its own asymmetric inheritance in the stomatal lineage to pattern the leaf surface

Asymmetric cell divisions (ACDs) specify differential cell fates across kingdoms. In metazoans, preferential inheritance of fate determinants into one daughter cell frequently depends on polarity-cytoskeleton interactions (1, 2). Despite the prevalence of ACDs during plant development, evidence for analogous mechanisms that segregate fate determinants during ACD remain elusive. Here, we describe a mechanism in the Arabidopsis thaliana leaf epidermis that ensures unequal inheritance of a fate-enforcing polarity domain during the creation of stomata, essential two-celled valves that mediate gas exchange between the plant and environment. Formation of a plasma membrane-associated polarity domain, defined by BREAKING OF ASYMMETRY IN THE STOMATAL LINEAGE (BASL), overrides default division patterns in stomatal precursors. The polarity domain exerts this control by constraining formation of the preprophase band of microtubules that mark the cortical division site and are a hallmark of plant mitosis. Experimentally uncoupling preprophase band establishment from the polarity domain results in aberrant polarity inheritance and subsequent fate errors. Mechanistically, our analyses of the interactions between microtubules and BASL in native and heterologous contexts revealed that the stomatal lineage polarity domain locally depletes cortical microtubules by altering microtubule stability. As the inherited cortical BASL crescent scaffolds a MAPK cascade to suppress progenitor identity in one daughter post-division, we propose that BASL-microtubule interactions represent a novel strategy to link cell identity to division orientation. Together, our data highlight how a common biological module, coupling the cytoskeleton to fate segregation via cell polarity, has been configured to accommodate the unique features of plant development.

cell biology↗

bioRxiv2021

Arabidopsis stomatal polarity protein BASL mediates distinct processes before and after cell division to coordinate cell size and fate asymmetries

In many land plants, asymmetric cell divisions (ACDs) create and pattern differentiated cell types on the leaf surface. In the Arabidopsis stomatal lineage, BREAKING OF ASYMMETRY IN THE STOMATAL LINEAGE (BASL) regulates multiple aspects of ACD including division plane placement and cell fate enforcement. Polarized subcellular localization of BASL is initiated before the ACD and persists for many hours after the division in one of the two daughters. Untangling the respective contributions of polarized BASL before and after division is essential to gain a better understanding of its roles in regulating stomatal lineage ACDs and to uncover the rules that guide leaf pattern. Here we combine quantitative imaging and lineage tracking with genetic tools that provide temporally-restricted BASL expression. We find that pre-division BASL is required for division orientation, whereas BASL polarity post-division ensures proper cell fate commitment. These genetic manipulations allowed us to uncouple daughter-cell size asymmetry from polarity crescent inheritance, revealing independent effects of these two asymmetries on subsequent cell behavior. Finally, we show that there is coordination between the division frequencies of sister cells produced by ACDs, and this coupling requires BASL as an effector of peptide signaling.

plant biology↗

bioRxiv2020

Mechanical Regulation of Stem Cell Proliferation and Fate Decisions by their Differentiated Daughters

Basal stem cells fuel development, homeostasis, and regeneration of the epidermis. The proliferation and fate decisions of these cells are highly regulated by their microenvironment, including the basement membrane and underlying mesenchymal cells. Basal progenitors give rise to differentiated progeny that serve an essential role in generating the epidermal barrier. Here, we present data that differentiated progeny also regulate the proliferation, differentiation, and migration of basal progenitor cells. Using two distinct mouse lines, we found that increasing contractility of differentiated cells resulted in non-cell autonomous hyperproliferation of stem cells and prevented their commitment to a hair follicle lineage. These phenotypes were rescued by pharmacological inhibitors of contractility. Live-imaging revealed that increasing the contractility of differentiated cells resulted in stabilization of adherens junctions and impaired movement of basal progenitors during hair placode morphogenesis, as well as a defect in migration of melanoblasts. These data suggest that intra-tissue tension regulates stem cell proliferation, fate decisions and migration, similar to the known roles of extracellular matrix rigidity. Additionally, this work demonstrates that differentiated epidermal keratinocytes are a component of the stem cell niche that regulates development and homeostasis of the skin.

cell biology↗

bioRxiv2017

A transgenic toolkit for visualizing and perturbing microtubules reveals unexpected functions in the epidermis

Differentiation induces reorganization of microtubules (MTs) into non-centrosomal arrays in a variety of tissues. The physiological functions of these microtubule arrays are just beginning to be understood as few tools currently exist to genetically perturb microtubule organization in vivo, particularly in mammals. We developed a genetic toolkit that can be broadly applied to the study of microtubule dynamics and function in many cell types. Using a TRE-EB1-GFP mouse we demonstrate that distinct differentiation transitions in the epidermis cause a decrease in microtubule growth rates and microtubule growth lifetimes, resulting in strong suppression of dynamics. To understand the physiological functions of these stable, non-centrosomal microtubules, we generated a TRE-spastin mouse, which can be used to perturb microtubule organization in a wide-variety of tissues in vivo. Unexpectedly, microtubule perturbation exclusively in post-mitotic keratinocytes had profound consequences on epidermal morphogenesis. We uncoupled novel cell-autonomous roles for MTs in differentiation-driven cell flattening from non-cell autonomous functions in regulating proliferation, differentiation, and tissue architecture. Taken together, we have created tools that will be broadly useful for the study of microtubule dynamics and function in mammalian tissue physiology and have used them to uncover previously unknown functions for non-centrosomal microtubules during mammalian epidermal development.

cell biology↗