Repeat associated small RNAs vary among parents and following hybridization in maize

Abstract

Small RNAs (sRNAs) are hypothesized to contribute to hybrid vigor because they maintain genome integrity, contribute to genetic diversity, and control gene expression. We used Illumina sequencing to assess how sRNA populations vary between two maize inbred lines (B73 and Mo17) and their hybrid. We sampled sRNAs from the seedling shoot apex and the developing ear, two rapidly growing tissues that program the greater growth of maize hybrids. We found that parental differences in siRNAs primarily originate from repeat regions. Although the maize genome contains greater number and complexity of repeats compared with Arabidopsis or rice, we confirmed that, like these simpler plant genomes, 24-nt siRNAs whose abundance differs between maize parents also show a trend of down-regulation following hybridization. Surprisingly, hybrid vigor is fully maintained when 24-nt siRNAs are globally reduced by mutation of the RNA-dependent RNA polymerase 2 encoded by modifier of paramutation1 (mop1). We also discovered that 21-22-nt siRNAs derived from a number of distinct retrotransposon families differentially accumulate between B73 and Mo17 as well as their hybrid. Thus, maize possesses a unique source of genetic variation for regulating transposons and genes at a genomic scale, which may contribute to its high degree of observed heterosis.

Fig. 1.

Summary of actively growing and highly proliferative tissues investigated for maize inbred and hybrid sRNA sequencing experiment. (A) At 11 DAS, tissues enriched for the shoot apex were collected by removing emerged leaves and sampling the bottom 1 cm of the remaining leaf tissue. (B) When the shoot had elongated 12 fully-expanded leaves (i.e., at V12), the top developing ear was excised.

Fig. 2.

Global differences in sRNAs between parents and hybrids result from parents passing on different populations of distinct siRNAs. Venn diagrams show percentage of total 21- to 24-nt siRNA abundance accounted for by each genotypic group for shoot apex (A) and developing ear (B).

Fig. 3.

Parental differences in siRNA regions primarily originate from repeats and deviate less from midparent in the shoot apex compared with the developing ear. Shown are the 22-nt and 24-nt siRNA clusters with at least 5 rpm-repnorm for shoot apex (A–C; n = 1,306) and developing ear (D–F; n = 5,110). Clusters are arranged in ascending order of parental fold change. (A and D) Classification of clusters based on type and genetic feature. (B and E) Degree of parental difference for siRNA clusters (log10 of high parent abundance divided by low parent abundance). Clusters below the horizontal gray line have parental differences that fall within the top 10% of the values for all of the clusters (shoot apex, 8.9-fold; ear, 9.5-fold). (C and F) Deviation from midparent values for siRNA clusters (log2 of F1 abundance divided by midparent abundance).

Fig. 4.

Parental differences in retrotransposon siRNA activity are driven by 21-nt and 22-nt siRNAs. siRNA profiles are displayed for shoot apex (A) and developing ear (B). The total abundance column displays the relative abundance of 21-, 22-, and 24-nt siRNAs matching maize characterized retrotransposon families (at most 1-bp mismatch). For each family, abundance is partitioned by genotype and siRNA length.

Fig. 5.

Loss of mop1-1 reduces 24-nt siRNAs in the developing ear but does not suppress hybrid vigor for B73×Mo17. (A) ΔΔCt values for siRNAs 24-A and 24-B using microRNA172 as the reference. RNAs were assayed from developing tissue (top ear from V10–V12 plant growth stage) from WT and mop1-1 field grown plants. Error bars represent ±2 SEM of the ΔΔCt values for four individual ears. (B) Vegetative and reproductive growth for WT and mop1-1 mutant B73×Mo17 hybrids compared with their parents. (C) Hybrid-to-inbred ratio for mean agronomic trait values. For each genotype, the height of 20 plants and cob weight of 25 plants were measured, and five replicate rows were measured for flowering traits and stover biomass.