Biology subjects

Baldrich, P.

Publications and source records attributed to Baldrich, P..

17 recordsLinked to original sources

bioRxiv2026

Extracellular Vacuole-derived bodies (EVacs) mediate RNA secretion in plants

Extracellular RNAs are found in the plant extracellular space, but how they are exported from cells remains unclear. We found that the plant vacuole is a major source of extracellular RNA and identified a class of large extracellular vacuole-derived bodies, which we termed EVacs, that are key mediators of this transport. EVacs are marked by the vacuolar membrane (tonoplast) proteins {gamma}-TIP and V-ATPase and originate as intravacuolar structures formed by inward folding of the tonoplast, encapsulating intact cytoplasmic material, including both RNAs and proteins. These intravacuolar bodies then escape the vacuole and are subsequently released from the plasma membrane of mesophyll cells into the apoplast. These findings provide a novel mechanism for the unconventional secretion of macromolecules in plants.

plant biology↗

bioRxiv2026

Reprogramming of auxin and brassinosteroid signaling is an early part of the homeostatic response to a viral movement protein

Plant viruses rely on intercellular trafficking via plasmodesmata (PD) to move between cells in their hosts. This ability is conferred by virus encoded movement proteins (MPs), which can increase plasmodesmal permeability and intercellular trafficking independent of other viral proteins. Callose dynamics in the cell walls surrounding PD have a critical role in determining plasmodesmal flux, with decreased callose levels correlates with increased trafficking. Notably, PD callose levels are both increased and decreased during virus infection, suggesting that there are regulatory responses to the virus. Here, we found that auxin and brassinosteroid (BR) exert opposing effects on PD connectivity. While auxin enhances intercellular trafficking primarily by promoting PD density, BR restricts connectivity by reducing reduced PD biogenesis and increasing callose accumulation. We identified genes involved in auxin and BR signaling as Most of those genes encode membrane-associated proteins. We identified the receptor-like protein RLP15 as a critical upstream regulator of intracellular auxin homeostasis through stabilization of the ER-localized auxin transporter PILS5. In parallel, negative regulators of PD permeability including ERECTA, PPI, CER3, and DEAL2 define a host connectivity restraint network. These findings point to an auxin-BR module as a nexus for determining the degree of changes in plasmodesmal permeability that is elicited by the viral MP. Together with changes in callose dynamics at PD, this regulatory node allows plants to maintain homeostasis of intercellular trafficking, possibly contributing to maintenance of cell and tissue integrity during infection. Significance StatementPlant viruses encode movement proteins (MP) which increase plasmodesmal permeability to allow the local cell-to-cell trafficking of viral entities. This study identifies a non-canonical auxin-BR antagonistic module that regulates plasmodesmal connectivity in response to the changes triggered by viral movement protein early in the infection cycle. We demonstrate that MP30, encoded by the tobacco mosaic virus, rewires BRs and auxins roles in controlling intercellular communication by interfering with membrane proteins associated with these hormonal pathways. By defining this hormonal nexus, our findings reveal a sophisticated host-pathogen interface where plants attempt to maintain intercellular homeostasis during the onset of viral pathogenesis.

plant biology↗

bioRxiv2026

Stage-Specific RNA Turnover Drives Small RNA Dynamics in Arabidopsis-Colletotrichum Interactions

Small RNAs (sRNAs) are key regulators of plant defense and have been implicated in cross-kingdom interactions with pathogens. The hemibiotrophic fungus Colletotrichum higginsianum infects Arabidopsis thaliana through three stages: appressorial penetration, biotrophy, and necrotrophy. However, the dynamics of fungal and plant sRNA populations across these three stages have not been elucidated. Using high-throughput sequencing, we profiled sRNAs from A. thaliana and C. higginsianum during in planta appressorium (PA), biotrophic (BP), and necrotrophic (NP) phases, and compared them to fungal mycelia (MY) and in vitro appressoria (VA). Our analyses revealed stage-specific patterns in sRNA accumulation in both the plant and the pathogen. In C. higginsianum, sRNAs were dominated by 29 nt species in PA, BP, MY, and VA, but shifted to 18 nt in NP, consistent with RNA degradation during host cell death. In A. thaliana, sRNAs transitioned from 30-33-nt in PA/BP to a 21 nt dominant peak in NP. Also, TE-derived siRNAs and other regulatory sRNAs (miRNAs, ncRNA, snoRNAs and snRNAs) declined during NP. A total of 62 host miRNAs showed differential accumulation, including core plant developmental regulators active across infection stages, and stage-specific miRNAs such as miR396, miR170/171, miR472, and miR858b. tRFs displayed opposite trends in host and pathogen: fungal tRFs declined in NP, while host misc-tRFs, 5'-tRFs, and 3'-tRFs increased, suggesting contrasting regulatory roles. These results provide new insights into RNA-mediated plant-fungal interactions.

plant biology↗

bioRxiv2025

Redesigning petal shape, size, and color in soybean reveals unexpected phenotypes for floral organ development

Soybean (Glycine max) has not yet benefited from large-scale hybrid breeding efforts due to its small, self-fertilizing flowers that are difficult to emasculate, and limited attractiveness to pollinators. This study explores targeted floral trait engineering to enhance pollinator attraction, aiming to overcome barriers to soybean hybridization. We generated a high-resolution floral organ expression atlas and H3K4 trimethylation ChIP-Seq dataset to identify candidate genes involved in petal development, nectar sugar content, and petal pigmentation. Using CRISPR-based activation and repression systems, we modified the expression of AINTEGUMENTA (GmANT), BIGPETAL (GmBPE), and SUCROSE TRANSPORTER2 (GmSUC2). Contrary to expectations based on Arabidopsis homologs, transcriptional activation of GmANT_B and repression of GmBPE led to reduced, rather than increased, petal size, highlighting divergent regulatory mechanisms in soybean. Complementation of the W1 gene that controls petal pigmentation successfully converted white petals to purple, with preliminary evidence indicating that this color conversion may increase pollinator visitation. These results underscore the complexity of floral development in soybean and provide foundational tools and resources for future efforts to engineer reproductive traits for hybrid seed production.

plant biology↗

bioRxiv2025

Distinct classes of 21 and 24-nt phasiRNAs suggest diverse mechanisms of biogenesis and function in rice anther development

PhasiRNAs (phased small interfering RNAs) are a major class of plant small RNAs known to be key regulators in male reproductive development of maize (Zea mays) and rice (Oryza sativa), among other plants. Earlier research focused primarily on premeiotic 21-nt phasiRNAs and meiotic 24-nt phasiRNAs, while new studies uncovered a premeiotic class of 24-nt phasiRNAs. The biogenesis and function of these phasiRNAs remain unclear. We conducted an integrative analysis combining small RNA (sRNA) sequencing and transcriptomic profiling of sRNA-associated genes across ten developmental stages of anther in Kitaake rice to map associations between sRNA-related genes and phasiRNA classes. We identified previously undescribed classes of postmeiotic 21-nucleotide (nt) and 24-nt phasiRNA-producing loci and characterized their unique accumulation patterns. Additionally, our findings reveal distinct nucleotide composition and register accumulation among the phasiRNA classes, suggesting the presence of diverse mechanisms of biogenesis and function. Our results provide new insights into the regulatory complexity of phasiRNAs, establishing a foundation for further functional studies and advancing our understanding of their roles in anther development and their underlying mechanisms. Plain Language SummarySmall RNAs play important roles in controlling how plants develop, but their function in plant reproduction is still not well understood. In rice, small RNAs called phasiRNAs help regulate how pollen develops, but past research has mostly focused on just two types: one found before meiosis and one found during meiosis. In this study, we analyzed small RNA activity across ten stages of rice anther development and found new phasiRNA-producing genes that are active after meiosis. We also discovered that different types of phasiRNAs have distinct patterns of accumulation and composition, suggesting that they have specialized roles in reproductive development. These findings reveal new layers of complexity in how small RNAs function and set the stage for future research to understand their exact roles in pollen formation. This work may help improve our understanding of plant fertility and contribute to advancements in crop breeding. Core ideasO_LIStage-specific accumulation patterns suggest more specialized functions of phasiRNAs. C_LIO_LIMeiotic 24-nt phasiRNAs are compositionally distinct from other 24-nt phasiRNAs. C_LIO_LINewly described phasiRNA clusters may differ in mechanism of biogenesis and function. C_LI

plant biology↗

bioRxiv2024

Diverse plant RNAs coat Arabidopsis leaves and are distinct from apoplastic RNAs

Transgenic expression of a double-stranded RNA in plants can induce silencing of homologous mRNAs in fungal pathogens. Although such host-induced gene silencing is well-documented, the molecular mechanisms by which RNAs can move from the cytoplasm of plant cells across the plasma membrane of both the host cell and fungal cell are poorly understood. Indirect evidence suggests that this RNA transfer may occur at a very early stage of the infection process, prior to breach of the host cell wall, suggesting that silencing RNAs might be secreted onto leaf surfaces. To assess whether Arabidopsis plants possess a mechanism for secreting RNA onto leaf surfaces, we developed a protocol for isolating leaf surface RNA separately from intercellular (apoplastic) RNA. This protocol yielded abundant leaf surface RNA that displayed an RNA banding pattern distinct from apoplastic RNA, suggesting that it may be secreted directly from the leaf surface rather than exuded through stomata or hydathodes. Notably, this RNA was not associated with either extracellular vesicles or protein complexes; however, RNA species longer than 100 nucleotides could be pelleted by ultracentrifugation. Pelleting was inhibited by the divalent cation chelator EGTA, suggesting that these RNAs may form condensates on the leaf surface. These leaf surface RNAs are derived almost exclusively from Arabidopsis, but come from diverse genomic sources, including rRNA, tRNA, mRNA, intergenic RNA, microRNAs, and small interfering RNAs, with tRNAs especially enriched. We speculate that endogenous leaf surface RNA plays an important role in the assembly of distinct microbial communities on leaf surfaces. Significance StatementPlant leaves are colonized by a complex community of microbes that is shaped by host genetics. Although secreted metabolites are thought to mediate this effect, we investigated whether plants might also secrete RNA that could potentially structure microbial communities via cross-kingdom RNA interference. Here we report that Arabidopsis leaves are covered with diverse RNAs of plant origin, including abundant tRNAs and tRNA fragments. This leaf surface RNA is not associated with extracellular vesicles or protein complexes; however, it is less degraded than RNA found inside the extracellular spaces of leaves, suggesting that leaf surface RNA is secreted directly rather than exuded through stomata or hydathodes. We propose that this RNA plays a direct role in shaping the leaf microbiome.

plant biology↗

bioRxiv2023

Arabidopsis AGO1 N-terminal Poly-Q domain promotes phase separation and association with stress granules during heat stress

In Arabidopsis thaliana, ARGONAUTE1 (AGO1) plays a central role in microRNA (miRNA) and small interfering RNA (siRNA)-mediated silencing. Nuclear AGO1 is loaded with miRNAs and exported to the cytosol where it associates to the rough ER to conduct miRNA-mediated translational repression, mRNA cleavage and biogenesis of phased siRNAs. These latter, as well as other cytosolic siRNAs, are loaded into cytosolic AGO1, but in which compartment this happens is not known. Moreover, the effect of stress on AGO1 localization is still unclear. Here, we show that a 37{degrees}C heat stress (HS) promotes AGO1 protein accumulation in cytosolic condensates where it co-localizes with components of siRNA bodies and of stress granules (SGs). AGO1 contains a prion-like domain in its poorly characterized N-terminal Poly-Q domain, which, is sufficient to undergo phase separation, independent of the presence or absence of SGS3. HS only moderately affects the small RNA repertoire, the loading of AGO1 by miRNAs and the signatures of target cleavage, suggesting that its localization in condensates protects AGO1 rather than promotes or impairs its activity in reprograming gene expressing during stress. Collectively, our work shed new light on the impact of high temperature on a main effector of RNA silencing in plants.

plant biology↗

bioRxiv2023

The floral development of the allotetraploid Coffea arabica L. correlates with a small RNA dynamic reprogramming.

O_LINon-coding and coding RNAs are key regulators of plant growth, development, and stress responses. To investigate the types of transcripts accumulated during the vegetative to reproductive transition and floral development in the Coffea arabica L., we sequenced small RNA libraries from eight developmental stages, up to anthesis. C_LIO_LIWe combined this data with messenger RNA and PARE sequencing of two important development stages that marks the transition of an apparent latent to a rapid growth stage. In addition, we took advantage of multiple in silico tools to characterize genomic loci producing small RNAs such as phasiRNAs, miRNAs and tRFs. C_LIO_LIOur differential and co-expression analysis showed that some types of small RNAs such as tRNAs, snoRNAs, snRNAs and phasiRNAs preferentially accumulate in a stage- specific manner. C_LIO_LIMembers of the miR482/miR2118 superfamily and their 21-nucleotide phasiRNAs originating from resistance genes show a robust co-expression pattern that is maintained across all the evaluated developmental stages. Finally, the majority of miRNAs accumulate in a family-stage specific manner, related to modulated hormonal responses and transcription factors expression. C_LI Societal Impact StatementThis research holds potential to benefit millions of coffee-producing families in over 60 countries. We uncovered molecular regulatory mechanisms governing flower development, one of the causes for the Coffea arabicas uneven ripening. The absence of uniformity in coffee production, spanning from floral induction to branch senescence, has a detrimental impact on the final products quality. These insights will inform strategies for controlled coffee maturation, leading to improved, uniform harvests.

plant biology↗

bioRxiv2023

Introduction of Barnase/Barstar in soybean produces a rescuable male sterility system for hybrid breeding

Hybrid breeding for increased vigor has been used for over a century to boost agricultural outputs without requiring higher inputs. While this approach has led to some of the most substantial gains in crop productivity, breeding barriers have fundamentally limited soybean (Glycine max) from reaping the benefits of hybrid vigor. Soybean makes inconspicuous flowers that self-fertilize before opening, and thus are not readily amenable to outcrossing. In this study, we demonstrate that the Barnase/Barstar male sterility/male rescue system can be used in soybean to produce hybrid seeds. By expressing the cytotoxic ribonuclease, Barnase, under a tapetum-specific promoter in soybean anthers, we are able to completely block pollen maturation, creating male-sterile plants. We show that fertility can be rescued in the F1 generation of these Barnase-expressing lines when they are crossed with pollen from plants that express the Barnase inhibitor, Barstar. Importantly, we found that the successful rescue of male fertility is dependent on the relative dosage of Barnase and Barstar. When Barnase and Barstar were expressed under the same tapetum-specific promoter, the F1 offspring remained male-sterile. When we expressed Barstar under a relatively stronger promoter than Barnase, we were able to achieve a successful rescue of male fertility in the F1 generation. This work demonstrates the successful implementation of a biotechnology approach to produce fertile hybrid offspring in soybean.

plant biology↗

bioRxiv2022

miR472 deficiency enhances Arabidopsis thaliana defence without reducing seed production.

After having co-existed in plant genomes for at least 200 million years, the products of microRNA (miRNA) and Nucleotide-Binding Leucine Rich Repeat protein (NLR) genes formed a regulatory relationship in the common ancestor of modern gymnosperms and angiosperms. From then on, DNA polymorphisms occurring at miRNA target sequences within NLR transcripts must have been compensated by mutations in the corresponding mature miRNA sequence, therefore maintaining that regulatory relationship. The potential evolutionary advantage of such regulation remains largely unknown and might be related to two mutually non-exclusive scenarios: miRNA-dependent regulation of NLR levels might prevent defence mis-activation with negative effects on plant growth and reproduction; or reduction of active miRNA levels in response to pathogen derived molecules (PAMPS and silencing suppressors) might rapidly release otherwise silent NLR transcripts for rapid translation and thereby enhance defence. Here, we used Arabidopsis thaliana plants deficient for miR472 function to study the impact of releasing its NLR targets on plant growth and reproduction and on defence against the fungal pathogen Plectospharaella cucumerina. We show that miR472 regulation has a dual role, participating both in the tight regulation of plant defence and growth. MIM472 lines, with reduced active miR472, are more resistant to pathogens and, correlatively, have reduced relative growth compared to wild-type plants. However, despite MIM472 lines flower at the same time than their wild-type, the end of their reproductive phase is delayed, and they exhibit higher adult biomass, resulting in similar seed yield as the wild-type. Our study highlights how negative consequences of defence activation might be compensated by changes in phenology and that miR472 reduction is an integral part of plant defence responses.

plant biology↗

bioRxiv2021

Arabidopsis Apoplastic Fluid Contains sRNA- and Circular RNA-Protein Complexes that Are Located Outside Extracellular Vesicles

Previously, we have shown that apoplastic wash fluid purified from Arabidopsis leaves contains small RNAs (sRNAs). To investigate whether these sRNAs are encapsulated inside extracellular vesicles (EVs), we treated EVs isolated from Arabidopsis leaves with the protease trypsin and RNase A, which should degrade RNAs located outside EVs but not those located inside. These analyses revealed that apoplastic RNAs are mostly located outside EVs and are associated with proteins. Further analyses of these extracellular RNAs (exRNAs) revealed that they comprise both sRNAs and long non-coding RNAs (lncRNAs), including circular RNAs (circRNAs). We also found that exRNAs are highly enriched in the post-transcriptional modification N6-methyladenine (m6A). Consistent with this, we identified a putative m6A-binding protein in apoplastic wash fluid, GLYCINE-RICH RNA-BINDING PROTEIN 7 (GRP7), as well as the small RNA-binding protein ARGONAUTE2 (AGO2). These two proteins co-immunoprecipitated with each other, and with lncRNAs, including circRNAs. Mutation of GRP7 or AGO2 caused changes in both the sRNA and lncRNA content of apoplastic wash fluid, suggesting that these proteins contribute to the secretion and/or stabilization of exRNAs. We propose that these extravesicular RNAs mediate host-induced gene silencing, rather than RNA inside EVs. One-sentence summaryThe apoplast of Arabidopsis leaves contains diverse small and long-noncoding RNAs, including circular RNAs, that are bound to protein complexes and are located outside extracellular vesicles.

plant biology↗

bioRxiv2021

The Evolutionary History of Small RNAs in the Solanaceae

The Solanaceae or "nightshade" family is an economically important group that harbors a remarkable amount of diversity. To gain a better understanding of how the unique biology of the Solanaceae relates to the familys small RNA genomic landscape, we downloaded over 255 publicly available small RNA datasets that comprise over 2.6 billion reads of sequence data. We applied a suite of computational tools to predict and annotate two major small RNA classes: (1) microRNAs (miRNAs), typically 20-22 nt RNAs generated from a hairpin precursor and functioning in gene silencing, and (2) short interfering RNAs (siRNAs), including 24-nt heterochromatic siRNAs (hc-siRNAs) typically functioning to repress repetitive regions of the genome via RNA-directed DNA methylation, as well as secondary phased siRNAs (phasiRNAs) and trans-acting siRNAs (tasiRNAs) generated via miRNA-directed cleavage of a Pol II-derived RNA precursor. Our analyses described thousands of small RNA loci, including poorly-understood clusters of 22-nt siRNAs that accumulate during viral infection. The birth, death, expansion, and contraction of these small RNA loci are dynamic evolutionary processes that characterize the Solanaceae family. These analyses indicate that individuals within the same genus share similar small RNA landscapes, whereas comparisons between distinct genera within the Solanaceae reveal relatively few commonalities. ONE-SENTENCE SUMMARYWe use over 255 publicly-available small RNA datasets to characterize the small RNA landscape for the Solanaceae family.

plant biology↗

bioRxiv2021

The Arabidopsis F-box protein FBW2 degrades AGO1 to avoid spurious loading of illegitimate small RNA

RNA silencing is a conserved mechanism in eukaryotes and is involved in development, heterochromatin maintenance and defense against viruses. In plants, ARGONAUTE1 (AGO1) protein plays a central role in both microRNA (miRNA) and small interfering RNA (siRNA)-directed silencing and its expression is regulated at multiple levels. Here, we report that the F-box protein FBW2 targets proteolysis of AGO1 by a CDC48-mediated mechanism. We found that FBW2 assembles an SCF complex that recognizes the MID-PIWI domain of AGO1 and requires its C-terminal domain containing a GW motif for AGO1 turnover. We showed that FBW2 prefers the unloaded and some mutated forms of AGO1 protein. While FBW2 loss of function does not lead to strong growth or developmental defects, it significantly increases RNA silencing activity. Interestingly, under conditions in which small RNA production or accumulation is affected, the failure to degrade AGO1 in fbw2 mutants becomes more deleterious for the plant. Hence, the non-degradable AGO1 protein assembles high molecular weight complexes and binds illegitimate small RNA leading to the cleavage of new target genes that belong to stress responses and cellular metabolic processes. Therefore, the control of AGO1 homeostasis by ubiquitin ligases plays an important role in quality control to avoid off-target cleavage.

cell biology↗

bioRxiv2020

Ligation bias is a major contributor to nonstoichiometric abundances of secondary siRNAs and impacts analyses of microRNAs

Plant small RNAs are a diverse and complex set of molecules, ranging in length from 21 to 24 nt, involved in a wide range of essential biological processes. High-throughput sequencing is used for the discovery and quantification of small RNAs. However, several biases can occur during the preparation of small RNA libraries, especially using low input RNA. We used two stages of maize anthers to evaluate the performance of seven commercially-available methods for small RNA library construction, using different RNA input amounts. We show that when working with plant material, library construction methods have differing capabilities to capture small RNAs, and that different library construction methods provide better results when applied to the detection of microRNAs, phasiRNAs, or tRNA-derived fragment. We also observed that ligation bias occurs at both ends of miRNAs and phasiRNAs, suggesting that the biased compositions observed in small RNA populations, including nonstoichiometric levels of phasiRNAs within a locus, may reflect a combination of biological and technical influences.

plant biology↗

bioRxiv2020

The commitment of barley microspores into embryogenesis involves miRNA-directed regulation of members of the SPL, GRF and HD-ZIPIII transcription factor families

Microspore embryogenesis is a model for developmental plasticity and cell fate decisions. To investigate the role of miRNAs in this development, we sequenced sRNAs and the degradome of barley microspores collected prior to (day 0) and after (days 2 and 5) the application of a stress treatment known to induce embryogenesis. Microspores isolated at these timepoints were uniform in both appearance and in their complements of sRNAs. We detected 68 miRNAs in microspores. The abundance of 51 of these miRNAs differed significantly during microspore development. One group of miRNAs was induced when the stress treatment was applied, prior to being repressed when microspores transitioned to embryogenesis. Another group of miRNAs were up-regulated in day-2 microspores and their abundance remained stable or increased in day-5 microspores, a timepoint at which the first clear indications of the transition towards embryogenesis were visible. Collectively, these miRNAs might play a role in the modulation of the stress response, the repression of gametic development, and/or the gain of embryogenic potential. A degradome analysis allowed us to validate the role of miRNAs in regulating 41 specific transcripts. We showed that the transition of microspores toward the embryogenesis pathway involves miRNA-directed regulation of members of the ARF, SPL, GRF and HD-ZIPIII transcription factor families. We noted that 41.5% of these targets were shared between day-2 and day-5 microspores while 26.8% were unique to day-5 microspores. The former set may act to disrupt transcripts involved in pollen development while the latter set may drive the commitment to embryogenesis.

plant biology↗

bioRxiv2018

Plant Extracellular Vesicles Contain Diverse Small RNA Species and Are Enriched in 10 to 17 Nucleotide \"Tiny\" RNAs

Small RNAs (sRNAs) that are 21 to 24 nucleotides (nt) in length are found in most eukaryotic organisms and regulate numerous biological functions, including transposon silencing, development, reproduction, and stress responses, typically via control of the stability and/or translation of target mRNAs. Major classes of sRNAs in plants include microRNAs (miRNAs) and small interfering RNAs (siRNAs); sRNAs are known to travel as a silencing signal from cell to cell, root to shoot, and even between host and pathogen. In mammals, sRNAs are transported inside extracellular vesicles (EVs), which are mobile lipid compartments that participate in intercellular communication. In addition to sRNAs, EVs carry proteins, lipids, metabolites, and potentially other types of nucleic acids. Here we report that plant EVs also contain diverse species of sRNA. We found that specific miRNAs and siRNAs are preferentially loaded into plant EVs. We also report a previously overlooked class of \"tiny RNAs\" (10 to 17 nt) that are highly enriched in EVs. This new RNA category of unknown function has a broad and very diverse genome origin and might correspond to degradation products.

plant biology↗

bioRxiv2018

Biogenesis of a young, 22-nt microRNA in Phaseoleae species by precursor-programmed uridylation

Phased, secondary siRNAs (phasiRNAs) represent a class of small RNAs in plants generated via distinct biogenesis pathways, predominantly dependent on the activity of 22 nt miRNAs. Most 22 nt miRNAs are processed by DCL1 from miRNA precursors containing an asymmetric bulge, yielding a 22/21 nt miRNA/miRNA* duplex. Here we show that miR1510, a soybean miRNA capable of triggering phasiRNA production from numerous NB-LRRs, previously described as 21 nt in its mature form, primarily accumulates as a 22 nt isoform via monouridylation. We demonstrate that in Arabidopsis, this uridylation is performed by HESO1. Biochemical experiments showed that the 3 terminus of miR1510 is only partially 2-O-methylated, because of the terminal mispairing in the miR1510/miR1510* duplex that inhibits HEN1 activity in soybean. miR1510 emerged in the Phaseoleae ~41 to 42 MYA with a conserved precursor structure yielding a 22 nt monouridylated form, yet a variant in mung bean is processed directly in a 22 nt mature form. This analysis of miR1510 yields two observations: (1) plants can utilize post-processing modification to generate abundant 22 nt miRNA isoforms to more efficiently regulate target mRNA abundances; (2) comparative analysis demonstrates an example of selective optimization of precursor processing of a young plant miRNA.

plant biology↗