Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Small RNA sequencing in maize hybrids and inbreds

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, 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 to 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 downregulation following hybridization. Surprisingly, hybrid vigor is fully maintained when 24-nt siRNAs are globally reduced by mutation of the RNA-dependent RNA polymerase2 (RDR2) encoded by modifier of paramutation1 (mop1). We also discovered that 21-22nt siRNAs derived from a number of distinct retrotransposon families differentially accumulate between B73 and Mo17 as well as their hybrid. Thus, maize possesses a novel source of genetic variation for regulating both transposons and genes at a genomic scale, which may contribute to its high degree of observed heterosis. sRNA libraries were derived from RNA isolated from the seedling shoot apex and developing ear tissues from B73, Mo17, B73xMo17 and Mo17xB73. The shoot apex was chosen because it is enriched for meristematic tissue where cell proliferation occurs, rates of organ initiation are determined, and organ size is specified. The developing ear was examined because it is enriched in meristematic tissue and is undergoing rapid growth, and also because the mature ear shows the highest degree of heterosis. Total RNA was isolated and separated on a 15% TBE-Urea polyacrylamide gel. Using a 10-bp ladder, the sRNA fraction representing 10-40-bp was excised. sRNA libraries were prepared according to Lu et al. (2007) or manufacturer's instructitions (Illumina). A combination of Perl scripts and FASTX toolkit scripts were used to remove adapters, collapse identical sequences and count reads per sequence. Supplementary processed data text files contain the distinct sRNA sequences for all of the genotypes analyzed in that experiment. Abundance (reads per million) was calculated for each distinct sequence by dividing the number of reads of distinct sRNA in a library by the total number of sRNA reads for that library and multiplying this by 1 million. Genome builds: B73 genome, maizesequence.org release 4a.53 (October, 2009); Mo17 whole genome shotgun clones.

ORGANISM(S): Zea mays

SUBMITTER: Wesley Barber

PROVIDER: E-GEOD-37411 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

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

Barber Wesley T WT Zhang Wei W Win Hlaing H Varala Kranthi K KK Dorweiler Jane E JE Hudson Matthew E ME Moose Stephen P SP

Proceedings of the National Academy of Sciences of the United States of America 20120611 26

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 o ...[more]

PMID: 22689990

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Project description:Combining dissimilar parents often leads to increased vigor in the hybrid offspring. “Heterosis” describes both this behavior and its underlying Mendelian and non-Mendelian interactions [1], although its molecular basis remains largely unknown. Recent comparisons of small RNA (sRNA) profiles from parents and their heterotic progeny identified correlations between interparental 24-nucleotide (24-nt) RNA variation and non-additive 24-nt RNA changes in the resulting hybrid [2,3]. 24-nt RNAs guide de novo cytosine methylation, and several proteins are required for their biogenesis, including a Snf2-like ATPase: required to maintain repression1 (RMR1) [4]. We found height variation between heterotic hybrids +/- RMR1 activity, implicating a role for RMR1 in heterosis. Based on the published correlations mentioned above [2,3], we hypothesized that RMR1-loss reduces parental sRNAs, altering their relative ratios and changing the sRNA profiles in the resulting hybrid from those of a standard hybrid (from +RMR1 parents). To probe this hypothesis, we profiled sRNAs from parents and hybrids +/- RMR1 function, limiting the parental diversity to only portions of chromosomes 6 and 9. Our analysis will address how RMR1 loss changes hybrid sRNAs in the presence and absence of underlying genetic variation and help to determine how this loss results in different phenotypic outcomes from heterotic crosses. 1. Shull (1948) Genetics 2. Groszmann et al (2011) PNAS 3. Barber et al (2012) PNAS 4. Hale et al (2007) PLoS Biology We used an interchange chromosome (T6-9 043-1) introgressed into B73 (~95% B73) to create a local region of genetic diversity when crossed as a female parent to 100% B73 individual. Small RNAs from immature cobs were sequenced from both of these parents and their resulting hybrid (2 samples each). Additionally, a similar cross where the T6-9 containing female parent lacked RMR1 activity (rmr1-1 / rmr1-1) was used to generate a counterpart hybrid whose small RNAs were also profiled (2 samples) from immature cobs. Finally, small RNAs from 2 immature cobs of an unrelated rmr1 heterozygote (again highly introgressed into B73) were sequenced as well.

Project description:Transposable elements have the potential to act as controlling elements to influence the expression of genes. The current paradigm suggests that heterochromatic silencing can spread beyond the borders of the transposable element and influence the chromatin state of neighboring low-copy sequences. This would allow transposable elements to condition obligatory or facilitated epialleles and act as controlling elements. The maize genome contains numerous families of class I transposable elements (retrotransposons) that are present in moderate to high copy numbers and many are found in regions near genes which provides an opportunity to test whether the spreading of heterochromatin from retrotransposons is prevalent. We have investigated the extent of heterochromatin spreading into flanking DNA around each family of retrotransposons through profiling of DNA methylation and di-methylation of lysine 9 of histone 3 (H3K9me2) in low-copy regions of the maize genome. The effects of different retrotransposon families on local chromatin are highly variable. Some LTR families exhibit enrichment of heterochromatic marks within 800-1200 base pairs of the insertion site while other families have very little evidence for spreading of heterochromatic marks. The analysis of chromatin state in genotypes that lack specific insertions suggests that the adjacent heterochromatin results from spreading of silencing rather than insertion-site preferences. Genes that are located near elements that exhibit spreading of heterochromatin tend to be expressed at lower levels than other genes. Our findings suggest that a subset of LTR retrotransposon families may act as controlling elements influencing neighboring sequences while the majority of elements have little effect on flanking sequences Transposable elements comprise a substantial portion of many eukaryotic genomes. These mobile fragments of DNA can result in mutations through insertions into genes but may also affect the regulation of genes they insert near. There is evidence that the majority of transposable elements are epigenetically silenced and in some cases this silencing may affect neighboring sequences. However, evolutionary theory would predict that there would be selective pressures for transposable elements to limit their effects on neighboring genes. The maize genome has a complex organization with many genes flanked by retrotransposons. We profiled the spread of heterochromatin from the retrotransposons into nearby low copy sequences for 150 high copy retrotransposon families. While the manymajority of retrotransposons exhibit little to no spreading of heterochromatin there are a small number ofsome retrotransposon families that influence the chromatin state of surrounding regions. The families may represent bad “neighbors” that spread heterochromatin and influence nearby genes. 6 total samples: 3 replicates of B73 H3k9 and 3 replicates of Mo17 H3k9 3 total samples: mop1 mutant, b73.zmet2 (zmet2.m1 mutant in B73 background) and mo17.zmet2 (zmet2.m1 mutant in Mo17 background)