Biology subjects

Leong, J. X.

Publications and source records attributed to Leong, J. X..

5 recordsLinked to original sources

bioRxiv2026

The lymphangiogenic Ccbe1-Adamts-Vegfc axis shapes cerebrovascular regionalization via selective patterning of fenestrated vascular beds

Organ development and function require regional specialization of vascular networks, yet the underlying molecular mechanisms remain poorly understood. Here, we identify a non-canonical role for the lymphangiogenic Ccbe1-Adamts-Vegfc axis in shaping cerebrovascular regionalization in zebrafish. A ccbe1 knock-in reporter revealed highly restricted expression in the developing brain, with its genetic loss causing early vascularization defects at the pineal gland-choroid plexus interface. Simultaneous Vegfa signaling deficiency markedly exacerbated these regional vascular defects, accompanied by disrupted blood flow, local hemorrhage, and choroid plexus epithelial disorganization, while largely preserving neighboring barrier-forming vessels. Loss of vegfc closely phenocopied ccbe1 mutant defects, with fenestrated vascular patterning requiring both Vegfr2 and Vegfr3 signaling, whereas disruption of adamts3 and adamts14 indicated region-specific requirements for Vegfc-activating proteases. Conversely, pineal gland-specific overexpression of activated Vegfc partially rescued the vascularization defects. Collectively, these findings suggest that spatially restricted Ccbe1-Adamts-Vegfc activity provides angiogenic signaling specificity enabling fenestrated cerebrovascular patterning and regionalization.

developmental biology↗

bioRxiv2024

A molecular mechanism for blood-brain barrier and fenestrated vessel segregation from a multipotent cerebrovascular niche

Cerebrovascular endothelial cell (EC) subtypes characterized by blood-brain barrier (BBB) properties or fenestrated pores are essential components of brain-blood interfaces, supporting brain function and homeostasis. To date, the origins and developmental mechanisms underlying this heterogeneous EC network remain largely unclear. Using single-cell-resolution lineage tracing in zebrafish, we discover a multipotent vascular niche at embryonic capillary borders that generates ECs with BBB or fenestrated molecular identity. RNAscope analysis demonstrates restricted expression of flt4 in sprouting ECs contributing to fenestrated choroid plexus (CP) vasculature, identifying an early molecular distinction from adjacent BBB vessels. Mechanistically, flt4 null and cytoplasmic-domain-deletion mutants exhibit CP vascularization defects when combined with vegfr2 zebrafish paralog deletion. Pharmacological results support this co-requirement of Flt4 and Vegfr2 signaling and suggest the PI3K and ERK pathways as downstream effectors. These findings reveal a specialized developmental origin for BBB and fenestrated EC subtypes, and establish Flt4 as a crucial guidance receptor mediating their angiogenic segregation.

developmental biology↗

bioRxiv2023

Autophagy Restricts Tomato Fruit Ripening Via a General Role in Ethylene Repression

Autophagy, a cellular degradation pathway, and the phytohormone ethylene function in plant development, senescence, and stress responses. However, the manner of their interaction is mostly unknown. We reasoned that this may be revealed by studying autophagy in a climacteric fruit ripening context, for which ethylene is crucial. During ripening, fruits undergo softening, color change, toxic compound degradation, volatile production, and sugar assembly by fine-tuning synthesis and degradation of their cellular content. For autophagy activity assessment, we analyzed autophagy-related 8 (ATG8) lipidation and GFP-ATG8-labeled autophagosome flux in tomato fruit cells. Autophagy activity increased sharply from ripening initiation, climaxed at its middle stage, and declined towards its end, resembling ethylene production dynamics. Silencing the core-autophagy genes SlATG2, SlATG7, and SlATG4 separately in mature fruits resulted in early ethylene production and ripening onset, which was abrogated by 1-methylcyclopropene (1-MCP), an ethylene signaling inhibitor. Beyond ripening, Arabidopsis atg5 and atg7 mutant seedlings exhibited elevated ethylene production and sensitivity to 1-Aminocyclopropane 1-carboxylic acid (ACC), ethylenes precursor, which induces autophagy. This research demonstrates that autophagy limits tomato fruit ripening via a general role in ethylene restriction, opening the path for a mechanistic understanding of autophagy-ethylene crosstalk and harnessing autophagy for fruit shelf-life extension.

plant biology↗

bioRxiv2023

ATG8 delipidation is dispensable for plant autophagy

Intracellular recycling via autophagy is governed by post-translational modifications of the autophagy-related (ATG) proteins. One notable example is ATG4-dependent delipidation of ATG8, a process that plays critical but distinct roles in autophagosome formation in yeast and mammals. Here, we aimed to elucidate the specific contribution of this process to autophagosome formation in species representative of evolutionary distant green plant lineages: unicellular green alga Chlamydomonas reinhardtii, with a relatively simple set of ATG genes, and a vascular plant Arabidopsis thaliana, harboring expanded ATG gene families. Remarkably, the more complex autophagy machinery of Arabidopsis rendered ATG8 delipidation entirely dispensable for the maturation of autophagosomes, autophagic flux and related stress tolerance; whereas autophagy in Chlamydomonas strictly depended on the ATG4-mediated delipidation of ATG8. Importantly, we uncovered the distinct impact of different Arabidopsis ATG8 orthologs on autophagosome formation, especially prevalent under nitrogen depletion, providing a new insight into potential drivers behind the expansion of the ATG8 family in higher plants. Our findings underscore the evolutionary diversification of the molecular mechanism governing the maturation of autophagosomes in eukaryotic lineages and highlight how this conserved pathway is tailored to diverse organisms.

plant biology↗

bioRxiv2021

SPIRO-the automated Petri plate imaging platform designed by biologists, for biologists

Phenotyping of model organisms grown on Petri plates is often carried out manually, despite the procedures being time-consuming and laborious. The main reason for this is the limited availability of automated phenotyping facilities, whereas constructing a custom automated solution can be a daunting task for biologists. Here, we describe SPIRO, the Smart Plate Imaging Robot, an automated platform that acquires time-lapse photos of up to four vertically oriented Petri plates in a single experiment, corresponding to 192 seedlings for a typical root growth assay and up to 2500 seeds for a germination assay. SPIRO is catered specifically to biologists needs, requiring no engineering or programming expertise for assembly and operation. Its small footprint is optimized for standard incubators, the inbuilt green LED enables imaging under dark conditions, and remote control provides access to the data without interfering with sample growth. SPIROs excellent image quality is suitable for automated image processing, which we demonstrate on the example of seed germination and root growth assays. Furthermore, the robot can be easily customized for specific uses, as all information about SPIRO is released under open-source licenses. Importantly, uninterrupted imaging allows considerably more precise assessment of seed germination parameters and root growth rates compared to manual assays. Moreover, SPIRO enables previously technically challenging assays such as phenotyping in the dark. We illustrate the benefits of SPIRO in proof-of-concept experiments which yielded a novel insight on the interplay between autophagy, nitrogen sensing and photoblastic response.

plant biology↗