Biology subjects

Yuen, A. C.

Publications and source records attributed to Yuen, A. C..

4 recordsLinked to original sources

bioRxiv2022

A kinase translocation reporter reveals real-time dynamics of ERK activity in Drosophila

Extracellular Signal-Regulated Kinase (ERK) lies downstream of a core signalling cascade that controls all aspects of development and adult homeostasis. Recent developments have led to new tools to image and manipulate the pathway. However, visualising ERK activity in vivo with high temporal resolution remains a challenge in Drosophila. We adapted a kinase translocation reporter (KTR) for use in Drosophila, which shuttles out of the nucleus when phosphorylated by ERK. We show that ERK-KTR faithfully reports endogenous ERK signalling activity in developing and adult tissues, and that it responds to genetic perturbations upstream of ERK. Using ERK-KTR in time-lapse imaging, we made two novel observations: firstly, sustained hyperactivation of ERK by expression of dominant-active Epidermal Growth Factor Receptor raised the overall level but did not alter the kinetics of ERK activity; secondly, heterogeneity in ERK activity in retinal basal glia correlated with the direction of migration of individual cells. Our results show that KTR technology can be applied in Drosophila to monitor ERK activity in real-time and suggest that this modular tool can be further adapted to study other kinases. Summary StatementWe describe a reporter to study the dynamics of ERK signalling in Drosophila, use it to measure signalling in individual cells over time, and monitor development.

developmental biology↗

bioRxiv2021

Stem cells commit to differentiation following multiple induction events in the Drosophila testis.

How and when potential becomes restricted in differentiating stem cell daughters is poorly understood. While it is thought that signals from the niche are actively required to prevent differentiation, another model proposes that stem cells can reversibly transit between multiple states, some of which are primed, but not committed, to differentiate. In the Drosophila testis, somatic cyst stem cells (CySCs) generate cyst cells, which encapsulate the germline to support its development. We find that CySCs are maintained independently of niche self-renewal signals if activity of the PI3K/Tor pathway is inhibited. Conversely, PI3K/Tor is not sufficient alone to drive differentiation, suggesting that it acts to license cells for differentiation. Indeed, we find that the germline is required for differentiation of CySCs in response to PI3K/Tor elevation, indicating that final commitment to differentiation involves several steps and intercellular communication. We propose that CySC daughter cells are plastic, that their fate depends on the availability of neighbouring germ cells, and that PI3K/Tor acts to induce a primed state for CySC daughters to enable coordinated differentiation with the germline.

developmental biology↗

bioRxiv2020

An immobilization technique for long-term time-lapse imaging of explanted Drosophila tissues

Time-lapse imaging is an essential tool to study dynamic biological processes that cannot be discerned from fixed samples alone. However, imaging cell- and tissue-level processes in intact animals poses numerous challenges if the organism is opaque and/or motile. Explant cultures of intact tissues circumvent some of these challenges, but sample drift remains a considerable obstacle. We employed a simple yet effective technique to immobilize tissues in medium-bathed agarose. We applied this technique to study multiple Drosophila tissues from first-instar larvae to adult stages in various orientations and with no evidence of anisotropic pressure or stress damage. Using this method, we were able to image fine features for up to 18 hours and make novel observations. Specifically, we report that fibers characteristic of quiescent neuroblasts are inherited by their basal daughters during reactivation; that the lamina in the developing visual system is assembled roughly 2-3 columns at a time; that lamina glia positions are dynamic during development; and that the nuclear envelopes of adult testis cyst stem cells do not break down completely during mitosis. In all, we demonstrate that our protocol is well-suited for tissue immobilization and long-term live imaging, enabling new insights into tissue and cell dynamics in Drosophila.

developmental biology↗

bioRxiv2019

Tissue mechanics regulate mitotic nuclear dynamics during epithelial development

Cell divisions are essential for tissue growth. In pseudostratified epithelia, where nuclei are staggered across the tissue, each nucleus migrates apically before undergoing mitosis. Successful apical nuclear migration is critical to preserve tissue integrity during cell division. Most previous investigations have focused on the local cellular mechanisms controlling nuclear migration. Yet, inter-species and inter-organ comparisons of different pseudostratified epithelia suggest global tissue architecture may influence nuclear dynamics, but the underlying mechanisms remain elusive. Here, we use the developing Drosophila wing disc to systematically investigate, in a single epithelial type, how changes in tissue architecture during growth influence mitotic nuclear migration. We observe distinct nuclear dynamics at discrete developmental stages, as epithelial morphology changes. We then use genetic and physical perturbations to show a direct effect of cell density on mitotic nuclear positioning. We also find Rho kinase and Diaphanous, which facilitate mitotic cell rounding in confined cell conditions, are essential for efficient apical nuclear movement. Strikingly, perturbation of Diaphanous causes increasing defects in apical nuclear migration as the tissue grows, and these defects can be reversed by acute physical reduction of cell density. Our findings reveal how the mechanical environment imposed on cells within a tissue alters the molecular and cellular mechanisms adopted by single cells for mitosis. We speculate that mechanical regulation of apical mitotic positioning could be a global mechanism for tissue growth control.

developmental biology↗