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Title: Coexpression network and trans ‐activation analyses of maize reproductive phasiRNA loci

The anther‐enriched phased, small interfering RNAs (phasiRNAs) play vital roles in sustaining male fertility in grass species. Their long non‐coding precursors are synthesized by RNA polymerase II and are likely regulated by transcription factors (TFs). A few putative transcriptional regulators of the 21‐ or 24‐nucleotide phasiRNA loci (referred to as21‐or24‐PHASloci) have been identified in maize (Zea mays), but whether any of the individual TFs or TF combinations suffice to activate anyPHASlocus is unclear. Here, we identified the temporal gene coexpression networks (modules) associated with maize anther development, including two modules highly enriched for the21‐or24‐PHASloci. Comparisons of these coexpression modules and gene sets dysregulated in several reported male sterile TF mutants provided insights into TF timing with regard to phasiRNA biogenesis, including antagonistic roles for OUTER CELL LAYER4 and MALE STERILE23.Trans‐activation assays in maize protoplasts of individual TFs using bulk‐protoplast RNA‐sequencing showed that two of the TFs coexpressed with21‐PHASloci could activate several 21‐nucleotide phasiRNA pathway genes but not transcription of21‐PHASloci. Screens for combinatorial activities of these TFs and, separately, the recently reported putative transcriptional regulators of24‐PHASloci using single‐cell (protoplast) RNA‐sequencing, did not detect reproducible activation of either21‐PHASor24‐PHASloci. Collectively, our results suggest that the endogenous transcriptional machineries and/or chromatin states in the anthers are necessary to activate reproductivePHASloci.

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Award ID(s):
1754097 1907220
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  
Publisher / Repository:
Date Published:
Journal Name:
The Plant Journal
Page Range / eLocation ID:
p. 160-173
Medium: X
Sponsoring Org:
National Science Foundation
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