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Abstract

Self-incompatibility (SI) mechanisms in hermaphroditic flowering plants serve as crucial bar- riers to self-fertilization. In the Brassicaceae, the locus governing SI displays remarkable diver- sity, featuring numerous distinct alleles retained over long evolutionary times and organized in a complex dominance hierarchy. Under this hierarchy, the gene controlling SI specificity in pollen exhibits monoallelic expression in heterozygote individuals. This is achieved through the action of sRNAs produced by precursors acting as ”dominance modifiers” resembling miRNAs but compatible with multiple gene silencing pathways. Single sRNA precursor undergoes extensive processing, generating hundreds of sRNA molecules with varying sizes, abundance levels, and ARGONAUTE loading preferences. To study this gene silencing phenomenon, we established a reverse genetic approach in engineered Arabidopsis thaliana lines expressing components of the A. halleri SI system and observed that the transcriptional repression is independent of the canonical RNA-directed DNA Methylation pathway (RdDM). We developed a single-molecule transcript capture protocol, and remarkably we observed that the sRNAs seem to target the transcription start site, possibly indicating an interference with the transcriptional machinery. Overall, the question of the mechanisms by which this repression occurs remains open, espe- cially regarding the role of DNA methylation on target sequences. Unraveling these silencing mechanisms will offer insights into the mechanisms by which dominance/recessivity interactions can evolve.