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1. Evolutionarily New Solanaceous DCL 2 Family Members Acquire Functions in Tomato

   https://doi.org/10.1111/mpp.70173  

   Hao, Jie; Ma, Junfei; Liu, Bin; Jakše, Jernej; Wang, Ying (November 2025, Molecular Plant Pathology)

   ABSTRACT RNA silencing is one of the major defence activities against viral pathogens in plants. Silencing signals are initiated by Dicer‐like proteins (DCLs) to generate viral‐derived small RNAs (sRNAs). Viral sRNAs are then loaded into Argonaute proteins to form an RNA‐induced silencing complex to guide cleavage of target RNAs based on sequence homology. While the model regarding RNA silencing‐mediated defence against viral pathogens is largely established based on extensive studies using the model plantArabidopsis thaliana, there are diverse sets of silencing components in other plants, especially in domesticated crops. Here, we tracked the expansion of solanaceous‐specific DCL2 genes during the course of evolution. We found that theDCL2agene in tomato chromosome 6 is likely an evolutionarily new gene copy. We also found thatDCL2bis more prone to be induced by viral pathogens in tomato plants, which is dependent on the combinations of cultivar and viral pathogen. Both DCL2a and DCL2b are critical to suppress the accumulation titre of a subviral agent, potato spindle tuber viroid (PSTVd). We noticed an unusually high accumulation of viral sRNAs shorter than 20 nt (16‐ to 19‐nt in length) in viroid‐infected tomato cv. Heinz 1706. Using synthetic small interfering RNAs, we demonstrated that shorter size sRNAs may also play a role in suppressing target RNAs, which can be interfered with by a viral suppressor of silencing, P19. Altogether, we provided further insights into the expansion of functional DCL2 family members in theSolanaceaefamily and their roles in combating viral and subviral agents. 

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2. Phylogenetic analyses of seven protein families refine the evolution of small RNA pathways in green plants

   https://doi.org/10.1093/plphys/kiad141  

   Bélanger, Sébastien; Zhan, Junpeng; Meyers, Blake C (March 2023, Plant Physiology)

   Abstract Several protein families participate in the biogenesis and function of small RNAs (sRNAs) in plants. Those with primary roles include Dicer-like (DCL), RNA-dependent RNA polymerase (RDR), and Argonaute (AGO) proteins. Protein families such as double-stranded RNA-binding (DRB), SERRATE (SE), and SUPPRESSION OF SILENCING 3 (SGS3) act as partners of DCL or RDR proteins. Here, we present curated annotations and phylogenetic analyses of seven sRNA pathway protein families performed on 196 species in the Viridiplantae (aka green plants) lineage. Our results suggest that the RDR3 proteins emerged earlier than RDR1/2/6. RDR6 is found in filamentous green algae and all land plants, suggesting that the evolution of RDR6 proteins coincides with the evolution of phased small interfering RNAs (siRNAs). We traced the origin of the 24-nt reproductive phased siRNA-associated DCL5 protein back to the American sweet flag (Acorus americanus), the earliest diverged, extant monocot species. Our analyses of AGOs identified multiple duplication events of AGO genes that were lost, retained, or further duplicated in subgroups, indicating that the evolution of AGOs is complex in monocots. The results also refine the evolution of several clades of AGO proteins, such as AGO4, AGO6, AGO17, and AGO18. Analyses of nuclear localization signal sequences and catalytic triads of AGO proteins shed light on the regulatory roles of diverse AGOs. Collectively, this work generates a curated and evolutionarily coherent annotation for gene families involved in plant sRNA biogenesis/function and provides insights into the evolution of major sRNA pathways. 

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3. Small RNA biogenesis and degradation

   https://doi.org/10.1007/978-3-319-55520-1_6  

   Yu, Q.; Liu, Y.; Li, M.; Yu, B. (January 2017, RNA technologies)

   Small RNAs (sRNAs), ~20–25 nucleotide (nt) in size, regulate various biological processes in plants through directing sequence-specific gene silencing. sRNAs are derived from either single- or double-stranded precursor RNAs. Proper levels of sRNAs are crucial for plant growth, development, genomic stability, and adaptation to abiotic and biotic stresses. Studies have identified the machineries controlling sRNA levels through biogenesis and degradation. This chapter covers recent progresses related to mechanisms governing small RNA biogenesis and degradation. 

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4. Disruption of a ∼23–24 nucleotide small RNA pathway elevates DNA damage responses in Tetrahymena thermophila

   https://doi.org/10.1091/mbc.E20-10-0631  

   Lee, Suzanne R.; Pollard, Daniel A.; Galati, Domenico F.; Kelly, Megan L.; Miller, Brian; Mong, Christina; Morris, Megan N.; Roberts-Nygren, Kerry; Kapler, Geoffrey M.; Zinkgraf, Matthew; et al (July 2021, Molecular Biology of the Cell)

   Misteli, Tom (Ed.)

   Endogenous RNA interference (RNAi) pathways regulate a wide range of cellular processes in diverse eukaryotes, yet in the ciliated eukaryote, Tetrahymena thermophila, the cellular purpose of RNAi pathways that generate ∼23–24 nucleotide (nt) small (s)RNAs has remained unknown. Here, we investigated the phenotypic and gene expression impacts on vegetatively growing cells when genes involved in ∼23–24 nt sRNA biogenesis are disrupted. We observed slower proliferation and increased expression of genes involved in DNA metabolism and chromosome organization and maintenance in sRNA biogenesis mutants RSP1Δ, RDN2Δ, and RDF2Δ. In addition, RSP1Δ and RDN2Δ cells frequently exhibited enlarged chromatin extrusion bodies, which are nonnuclear, DNA-containing structures that may be akin to mammalian micronuclei. Expression of homologous recombination factor Rad51 was specifically elevated in RSP1Δ and RDN2Δ strains, with Rad51 and double-stranded DNA break marker γ-H2A.X localized to discrete macronuclear foci. In addition, an increase in Rad51 and γ-H2A.X foci was also found in knockouts of TWI8, a macronucleus-localized PIWI protein. Together, our findings suggest that an evolutionarily conserved role for RNAi pathways in maintaining genome integrity may be extended even to the early branching eukaryotic lineage that gave rise to Tetrahymena thermophila. 

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

   https://doi.org/10.1105/tpc.18.00872  

   Baldrich, Patricia; Rutter, Brian D.; Zand Karimi, Hana; Podicheti, Ram; Meyers, Blake C.; Innes, Roger W. (January 2019, The Plant Cell)

   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. 

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