Biology subjects

Nasfi, S.

Publications and source records attributed to Nasfi, S..

4 recordsLinked to original sources

bioRxiv2025

Cross-kingdom degradomics identifies natural plant small RNAs with disease protecting activity

RNA interference (RNAi) is a highly specific process regulating genes transcriptionally or post-transcriptionally via silencing. small regulatory RNAs (sRNAs) provide the specificity of post-transcriptional silencing by binding to complementary mRNAs leading to their degradation or translational repression. sRNAs are promising non-chemical control agents against plant pests and diseases. Their application requires, however, a deep understanding of the determinants of functional specificity and efficiency. In nature, interacting plants and pathogens follow the manipulative strategy of cross-kingdom RNAi (ckRNAi) based on the targeted transfer of sRNAs. In this study, we employed a genome-wide approach to explore the diversity of natural ck-sRNAs exchanged in the pathogenic interaction of Brachypodium distachyon with the fungal pathogen Fusarium graminearum. By deep-sequencing of the cross-kingdom sRNAome, transcriptome and degradome 258 ckRNAi-mediating sRNAs were discovered. In a simulation-based approach, specific sequence characteristics allowed categorizing sRNAs into functionality classes. Subsequent genome annotation analyses revealed the organization of plant ck-sRNAs in long non-coding RNAs. Moreover, comparative sequence analyses revealed an evolution of ck-sRNAs towards invariant mRNA target regions. Functional analyses with sRNA candidates confirmed an antipathogenic activity at exceptionally low application doses. It indicates natural sRNA as untapped resource and potential blueprint for the development of highly specific and effective plant-derived bioprotectants.

plant biology↗

bioRxiv2025

High Precision Quantification of small RNA Slicing Activity - Native Index Ligation-based Targeted Degradome Sequencing (NIL-TDS)

RNA interference (RNAi) is an effective and precise regulatory mechanism in eukaryotes in which small RNAs mediate endonucleolytic slicing of complementary target mRNAs. Despite the potential of RNAi for human therapeutics and crop bio-protection, analytical platforms to quantitatively validate the slicing activities of small RNAs remain limited. Here, we present NIL-TDS, a cost-effective method that combines RNA ligase-mediated PCR with Nanopore Sequencing for direct, rapid, and high-resolution detection and quantification of sRNA-mediated slicing events. Using NIL-TDS, we quantitatively detected minute changes in Ath-mir400 mediated slicing of PPR1 in heat and salt stressed Arabidopsis plants. We further demonstrate the broader applicability of NIL-TDS by detecting rare slicing events in a mammalian system. Of relevance for malignancy of certain cancers, and tumor progression and metastasis, NIL-TDS further confirmed miR-196-HOXB8 interactions in lung cancer cells and discovered a novel miR-7162-HOXA10-AS slicing site. These findings demonstrate the sensitivity of NIL-TDS for uncovering small RNA mediated regulatory mechanisms of gene expression and disease progression in eukaryotes. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=83 SRC="FIGDIR/small/679503v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@adef35org.highwire.dtl.DTLVardef@1339570org.highwire.dtl.DTLVardef@1980652org.highwire.dtl.DTLVardef@1c732ef_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Broad-scale phenotyping in Arabidopsis reveals varied involvement of RNA interference across diverse plant-microbe interactions

RNA interference (RNAi) is a crucial mechanism that can contribute to immunity against infectious microbes through the action of DICER-LIKE (DCL) and ARGONAUTE (AGO) proteins. In the case of the fungal pathogen Botrytis cinerea and the oomycete Hyaloperonospora arabidopsidis, plant DCL and AGO proteins have proven roles as negative regulators of immunity, suggesting functional specialization of these proteins. To address this aspect in a broader taxonomic context, we characterized the colonization pattern of an informative set of DCL and AGO loss-of-function mutants in Arabidopsis thaliana upon infection with a panel of pathogenic microbes with different lifestyles, and a fungal mutualist. Our results revealed that AGO1 and AGO4 function as positive regulators of immunity to a bacterial and a fungal pathogen, respectively. Additionally, AGO2 and AGO10 positively modulated the colonization by a fungal mutualist. Therefore, analysing the role of RNAi across a broader range of plant-microbe interactions has identified previously unknown functions for AGO proteins. For some pathogen interactions, however, all tested mutants exhibited wild type-like infection phenotypes, suggesting that the roles of AGO and DCL proteins in these interactions may be more complex to elucidate.

plant biology↗

bioRxiv2023

A robust pipeline for the systematic validation of sRNA effectors suggests cross-kingdom communication in the symbiosis of Serendipita indica with Arabidopsis thaliana

Bidirectional communication between pathogenic microbes and their plant hosts via small (s)RNA-mediated cross-kingdom RNA interference (ckRNAi) is one key element for successful host colonisation. However, whether mutualistic fungi from the Serendipitaceae family, known for their extremely broad host range, employ sRNAs to colonize plant roots is still under discussion. To address this question, we developed a pipeline to validate the accumulation, translocation, and activity of fungal sRNAs in post-transcriptional silencing of Arabidopsis thaliana genes. Using stem-loop PCR, we detected the expression of a specific set of Serendipita indica (Si)sRNAs, targeting host genes involved in cell wall organization, hormonal signalling regulation, immunity, and gene regulation. To confirm the gene silencing activity of these sRNA in plant cells, SisRNAs were transiently expressed in protoplasts. Stem-loop PCR proved the expression of sRNAs, while qPCR validated post-transcriptional gene silencing of their predicted target genes. Furthermore, ARGONAUTE 1 immunoprecipitation (AtAGO1-IP) revealed the loading of fungal SisRNAs into the plant RNAi machinery, suggesting the translocation of SisRNA from the fungus into root cells. In conclusion, this study provides a blueprint for rapid selection and analysis of sRNA effectors in plant-microbe interactions and further suggests cross-kingdom communication in the Sebacinoid symbiosis. HighlightSmall RNAs of the beneficial fungus Serendipita indica are translocated and silence Arabidopsis genes at the onset of the interaction, revealing cross-kingdom communication in sebacinoid symbiosis.

plant biology↗