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Spatiotemporally dynamic, cell-type–dependent premeiotic and meiotic phasiRNAs in maize anthers
Saturday, 2015/03/14 | 07:04:31
  1. Jixian Zhaia,1,2,
  2. Han Zhangb,1,
  3. Siwaret Arikita,3,
  4. Kun Huanga,
  5. Guo-Ling Nanb,
  6. Virginia Walbotb,4, and
  7. Blake C. Meyersa,4

Significance

By RNA profiling of 10 stages of maize anthers plus mature pollen, we found two distinct classes of phased small-interfering RNAs (phasiRNAs): 21-nt premeiotic phasiRNAs, after germinal and somatic cell specification, and 24-nt meiotic phasiRNAs coordinately accumulated during meiosis and persist into pollen. Sequencing of RNA from five male-sterile, anther developmental mutants—ocl4, mac1, ms23, msca1, and ameiotic1—demonstrated the involvement of specific somatic layers. Premeiotic phasiRNAs require a functional epidermis, whereas meiotic phasiRNAs require a normal tapetum. Mammalian germ cells express “prepachytene” or “pachytene” PIWI-interacting RNAs (piRNAs). Whereas differences in biogenesis indicate independent origins, grass phasiRNAs and mammalian piRNAs share developmental timing, a lack of obvious targets, and an impact on male fertility, suggesting a possible evolutionary convergence.

Abstract

Maize anthers, the male reproductive floral organs, express two classes of phased small-interfering RNAs (phasiRNAs). PhasiRNA precursors are transcribed by RNA polymerase II and map to low-copy, intergenic regions similar to PIWI-interacting RNAs (piRNAs) in mammalian testis. From 10 sequential cohorts of staged maize anthers plus mature pollen we find that 21-nt phased siRNAs from 463 loci appear abruptly after germinal and initial somatic cell fate specification and then diminish, whereas 24-nt phasiRNAs from 176 loci coordinately accumulate during meiosis and persist as anther somatic cells mature and haploid gametophytes differentiate into pollen. Male-sterile ocl4 anthers defective in epidermal signaling lack 21-nt phasiRNAs. Male-sterile mutants with subepidermal defects—mac1 (excess meiocytes), ms23 (defective pretapetal cells), and msca1 (no normal soma or meiocytes)—lack 24-nt phasiRNAs. ameiotic1 mutants (normal soma, no meiosis) accumulate both 21-nt and 24-nt phasiRNAs, ruling out meiotic cells as a source or regulator of phasiRNA biogenesis. By in situ hybridization, miR2118 triggers of 21-nt phasiRNA biogenesis localize to epidermis; however, 21-PHAS precursors and 21-nt phasiRNAs are abundant subepidermally. The miR2275 trigger, 24-PHAS precursors, and 24-nt phasiRNAs all accumulate preferentially in tapetum and meiocytes. Therefore, each phasiRNA type exhibits independent spatiotemporal regulation with 21-nt premeiotic phasiRNAs dependent on epidermal and 24-nt meiotic phasiRNAs dependent on tapetal cell differentiation. Maize phasiRNAs and mammalian piRNAs illustrate putative convergent evolution of small RNAs in male reproduction.

 

See: http://www.pnas.org/content/112/10/3146.abstract.html?etoc

PNAS March 10, 2015 vol. 112 no. 10 3146-3151  

 

Fig. 2.

Fig. 2.

The 21-nt premeiotic and 24-nt meiotic phasiRNAs are developmentally regulated. (A) Anthers at 10 different lengths (developmental stages) were analyzed plus pollen. Above, a schematic of cell patterns in a single lobe of the anther; cell types in colors shown in key (Right). (B) Heat maps depicting the abundances of 21-nt premeiotic phasiRNAs from 463 loci (Left) and 24-nt meiotic phasiRNAs from 176 loci (Right) at each stage. Hierarchical clustering was based on similarity of expression pattern. (Left) Solid bubbles represent the total abundance of phasiRNAs; blue pie charts represent the proportion of 21-nt phasiRNAs from all 21-nt sRNAs at each stage; striped blue bubbles represent miR2118 abundances (trigger of premeiotic phasiRNA). (Right) The solid and striped orange bubbles represent 24-nt phasiRNA and miR2275 abundances, respectively; orange pie charts represent 24-nt phasiRNAs from all 24-nt sRNAs. Controls (Far Right): green represents TAS3-derived ta-siRNAs, and pink represents all TE-associated siRNAs mapped to the first 100 Mb of maize chromosome 1 (as a proxy for the whole genome). (C) Quantification of 21-PHAS and 24-PHAS precursor transcripts by RNA-seq.

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