Biology subjects

Bradamante, G.

Publications and source records attributed to Bradamante, G..

6 recordsLinked to original sources

bioRxiv2025

AGO5 restricts virus vertical transmission in plant gametophytes

The propagation of a viral infection from a host parent to its progeny is known as vertical transmission, or seed transmission in plants. It allows viral infections to rapidly spread locally via pollen and worldwide through seeds. To be vertically transmitted to each progeny, a virus must pass through the tight bottleneck of at least one cell per parent - the gametes. Therefore, stopping infection during sexual reproduction is of vital importance to generate healthy offspring. Accordingly, vertical transmission of plant viruses often occurs at very low rates, if at all, suggesting the existence of highly effective - yet unknown - antiviral defenses in pre-meiotic cells, gametes and/or embryos. In this study, we show that AGO5, an RNA interference factor expressed specifically in shoot apical meristem stem cells and the germline of Arabidopsis thaliana, drastically reduces the vertical transmission of Turnip yellow mosaic virus (TYMV). Through a series of controlled pollination experiments leveraging different zygosity of ago5 knock-out, cell type-specific rescue of ago5 and TYMV detection in whole-mount reproductive tissues, we provide evidence that AGO5 acts in pollen and sperm cells to restrict virus transmission to progeny. We further show that triggering antiviral RNA interference specifically in sperm cells leads to a significant reduction in TYMV vertical transmission. In summary, this study provides the first description of a gamete-specific antiviral defense mechanism restricting virus vertical transmission, paving the way for new strategies to prevent the spread of pollen- and seed-borne viral epidemics.

plant biology↗

bioRxiv2023

A versatile CRISPR-based system for lineage tracing in living plants

Individual cells give rise to diverse cell lineages during the development of multicellular organisms. Understanding the contribution of these lineages to mature organisms is a central question of developmental biology. Several techniques to document cell lineages have been used, from marking single cells with mutations that express a visible marker to generating molecular bar codes by CRISPR-induced mutations and subsequent single-cell analysis. Here, we exploit the mutagenic activity of CRISPR to allow lineage tracing within living plants. Cas9-induced mutations are directed to correct a frameshift mutation that restores expression of a nuclear fluorescent protein, labelling the initial cell and all progenitor cells with a strong signal without modifying other phenotypes of the plants. Spatial and temporal control of Cas9 activity can be achieved using tissue-specific and/or inducible promoters. We provide proof of principle for the function of lineage tracing in two model plants. The conserved features of the components and the versatile cloning system, allowing for easy exchange of promoters, are expected to make the system widely applicable. SIGNIFICANCE STATEMENTBy targeting Cas9 in a tissue- and time-specific way to correct a frameshift mutation, resulting in fluorescence labelling of nuclei, we generated a method for in vivo visual lineage tracing in two model plants. The versatile cloning system makes the system widely applicable in other plants.

cell biology↗

bioRxiv2022

A hormone-activated mobile RNAi pathway defends plant stem cells from virus infection

Stem cells are essential for the development and organ regeneration of multicellular organisms, so their infection by pathogenic viruses must be prevented. Accordingly, mammalian stem cells are highly resistant to viral infection due to dedicated antiviral pathways including RNA interference (RNAi) (1, 2). In plants, a small group of stem cells harbored within the shoot apical meristem (SAM) generates all postembryonic above-ground tissues, including the germline cells. Many viruses do not proliferate in these cells, yet the molecular bases of this exclusion remain only partially understood (3, 4). Here we show that a plant-encoded RNA-dependent RNA polymerase, after activation by the plant hormone salicylic acid, amplifies antiviral RNAi in infected tissues. This provides stem cells with RNA-based virus sequence information, which prevents virus proliferation. Furthermore, we find RNAi to be necessary for stem cell exclusion of several unrelated RNA viruses, despite their ability to efficiently suppress RNAi in the rest of the plant. This work elucidates a molecular pathway of great biological and economic relevance and lays the foundations for our future understanding of the unique systems underlying stem cell immunity.

plant biology↗

bioRxiv2022

Two AGO proteins with transposon-derived sRNA cargo mark the germline in Arabidopsis

Summary paragraphIn sexually propagating organisms, genetic and epigenetic mutations are evolutionarily relevant only if they occur in the germline and provide inherited information to the next generation. In contrast to most animals, plants are thought to lack an early segregating germline, implying that somatic cells can contribute genetic information to the progeny. Here we demonstrate that two ARGONAUTE proteins, AGO5 and AGO9, mark an early-segregating germline. Both AGOs are loaded with dynamically changing populations of small RNAs derived from highly methylated, pericentromeric, long transposons. Sequencing single nuclei revealed that many of these transposons are co-expressed within an AGO5/9 expression domain of the shoot apical meristem (SAM). This indicates a host-parasite tug of war and specific silencing pathways along the plant germline throughout development. Our results open the path to investigate transposon biology and epigenome dynamics at cellular resolution in the SAM stem cell niche.

plant biology↗

bioRxiv2019

Dual-sensing genetically encoded fluorescent indicators resolve the spatiotemporal coordination of cytosolic abscisic acid and second messenger dynamics in Arabidopsis

Deciphering signal transduction processes is crucial for understanding how plants sense and respond to environmental changes. Various chemical compounds function as central messengers within deeply intertwined signaling networks. How such compounds act in concert remains to be elucidated. We have developed dual-sensing genetically encoded fluorescent indicators (2-In-1-GEFIs) for multiparametric in vivo analyses of the phytohormone abscisic acid (ABA), Ca2+, protons (H+), chloride (anions), the glutathione redox potential (EGSH) and hydrogen peroxide (H2O2). Simultaneous analyses of two signaling compounds in Arabidopsis (Arabidopsis thaliana) roots revealed that ABA treatment and uptake did not trigger rapid cytosolic Ca2+ or H+ fluxes. Glutamate, ATP, Arabidopsis PLANT ELICITOR PEPTIDE (AtPEP1) and glutathione disulfide (GSSG) treatments induced rapid spatiotemporally overlapping cytosolic Ca2+, H+ and anion fluxes, but except for GSSG only weakly affected the cytosolic redox state. Overall, 2-In-1-GEFIs enable complementary high-resolution in vivo analyses of signaling compound dynamics and facilitate an advanced understanding of the spatiotemporal coordination of signal transduction processes in Arabidopsis.

plant biology↗

bioRxiv2018

Stage-specific transcriptomes and DNA methylomes indicate an early and transient loss of transposon control in Arabidopsis shoot stem cells

In contrast to animals, postembryonic development in plants is modular, and aerial organs originate from stem cells in the center of the shoot apical meristem (SAM) throughout life. Descendants of SAM stem cells in the subepidermal layer (L2) give also rise to male and female gametes (reviewed in 1) and are therefore considered primordial germ cells. In these cells, transmission of somatic mutations including virus and TE insertions must be avoided. Despite their essential role for plant development and intergenerational continuity, no comprehensive molecular analysis of SAM stem cells exists, due to their low number, deep embedding among non-stem cells, and difficult isolation. Here we present a comprehensive analysis of stage-specific gene expression and DNA methylation dynamics in Arabidopsis SAM stem cells. Stem cell expression signatures are mostly defined by development, but we also identified a core set of differentially expressed stemness genes. Surprisingly, vegetative SAM stem cells showed increased expression of transposable elements (TEs) relative to surrounding cells, despite high expression of genes connected to epigenetic silencing. We also find increasing methylation at CHG and a drop in CHH methylation at TEs before stem cells enter the reproductive lineage, indicating an onset of epigenetic reprogramming at an early stage. Transiently elevated TE expression is reminiscent of that in animal primordial germ cells (PGCs) 2 and demonstrates commonality of transposon biology. Our results connect SAM stem cells with germline development and transposon evolution and will allow future experiments to determine the degree of epigenetic heritability between generations.

developmental biology↗