The miRNome function transitions from regulating developmental genes to transposable elements during pollen maturation

Abstract

Animal and plant microRNAs (miRNAs) are essential for the spatio-temporal regulation of development. Together with this role, plant miRNAs have been proposed to target transposable elements (TEs) and stimulate the production of epigenetically active small interfering RNAs. This activity is evident in the plant male gamete containing structure, the male gametophyte or pollen grain. How the dual role of plant miRNAs, regulating both genes and TEs, is integrated during pollen development and which mRNAs are regulated by miRNAs in this cell type at a genome-wide scale are unknown. Here, we provide a detailed analysis of miRNA dynamics and activity during pollen development in Arabidopsis thaliana using small RNA and degradome parallel analysis of RNA end high-throughput sequencing. Furthermore, we uncover miRNAs loaded into the two main active Argonaute (AGO) proteins in the uninuclear and mature pollen grain, AGO1 and AGO5. Our results indicate that the developmental progression from microspore to mature pollen grain is characterized by a transition from miRNAs targeting developmental genes to miRNAs regulating TE activity.

Single-nucleus RNA-seq reveals that MBD5, MBD6, and SILENZIO maintain silencing in the vegetative cell of developing pollen

“…We also looked at the expression patterns of the small RNA machinery, given its prominent role in pollen biology. 45 , 46 As previously described, 45 , 46 we found that AGO5 was expressed in GN and AGO1 in microspores and VN ( Figure S5K ).…”

Single-nucleus RNA-seq reveals that MBD5, MBD6, and SILENZIO maintain silencing during epigenetic reprogramming in pollen

“…We also looked at the expression patterns of the small RNA machinery, given its prominent role in pollen biology (Borges et al, 2011;Oliver et al, 2022). As previously described, we found that AGO5 was expressed in the GN and AGO1 in microspores and VN (Figure S5K).…”

“…Of note, the H3K27 methyltransferases CURLY LEAF ( CLF ) and SWINGER ( SWN ) were enriched in the VN and depleted from SN, which is consistent with a recent report of the erasure of H3K27 in sperm nuclei (Borg et al, 2020). We also looked at the expression patterns of the small RNA machinery, given its prominent role in pollen biology (Borges et al, 2011; Oliver et al, 2022). As previously described, we found that AGO5 was expressed in the GN and AGO1 in microspores and VN (Figure S5K).…”

Epigenetic Dynamics and Regulation of Plant Male Reproduction

“…AGOs can form RNA-induced silencing complexes (RISC) by binding 21–22 nt siRNAs and then mediating RNA cleavage leading to translational inhibition or participating in the RdDM pathway by binding 24 nt siRNAs [ 95 ]. In Arabidopsis , AtAGO1 is located in the pollen VC nucleus [ 96 ], and AtAGO5 accumulates in the SC cytoplasm [ 97 ], which makes the major AGO protein substances active in the mature pollen [ 86 ]. Weak mutants of ago1 and dcl1 showed pollen developmental defects and led to aborted or shrunken mature pollen grains, while strong ago1 and dcl1 mutants failed to develop gametes [ 98 , 99 ], indicating their essential role in gametophyte development.…”