Sunflower centromeres consist of a centromere-specific LINE and a chromosome-specific tandem repeat.

Kiyotaka Nagaki,Naoki Yamaji,Keisuke Tanaka,Hisato Kobayashi,Minoru Murata

doi:10.3389/fpls.2015.00912

Kiyotaka Nagaki, Naoki Yamaji + Show 3 more

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https://doi.org/10.3389/fpls.2015.00912

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Abstract

The kinetochore is a protein complex including kinetochore-specific proteins that plays a role in chromatid segregation during mitosis and meiosis. The complex associates with centromeric DNA sequences that are usually species-specific. In plant species, tandem repeats including satellite DNA sequences and retrotransposons have been reported as centromeric DNA sequences. In this study on sunflowers, a cDNA-encoding centromere-specific histone H3 (CENH3) was isolated from a cDNA pool from a seedling, and an antibody was raised against a peptide synthesized from the deduced cDNA. The antibody specifically recognized the sunflower CENH3 (HaCENH3) and showed centromeric signals by immunostaining and immunohistochemical staining analysis. The antibody was also applied in chromatin immunoprecipitation (ChIP)-Seq to isolate centromeric DNA sequences and two different types of repetitive DNA sequences were identified. One was a long interspersed nuclear element (LINE)-like sequence, which showed centromere-specific signals on almost all chromosomes in sunflowers. This is the first report of a centromeric LINE sequence, suggesting possible centromere targeting ability. Another type of identified repetitive DNA was a tandem repeat sequence with a 187-bp unit that was found only on a pair of chromosomes. The HaCENH3 content of the tandem repeats was estimated to be much higher than that of the LINE, which implies centromere evolution from LINE-based centromeres to more stable tandem-repeat-based centromeres. In addition, the epigenetic status of the sunflower centromeres was investigated by immunohistochemical staining and ChIP, and it was found that centromeres were heterochromatic.

Highlights

Asterales is the most diverged order of dicots and includes 11 families and 27,000 species
The epigenetic status at the centromeric DNA sequences was investigated by chromatin immunoprecipitation (ChIP)-qPCR with antibodies against modified histones
Centromeric DNA sequences in plant species consist of species-specific tandem repeats and retrotransposons, and these sequences are located on all centromeric regions (Zhong et al, 2002; Nagaki et al, 2003, 2009, 2011; Nagaki and Murata, 2005; Houben et al, 2007; Tek et al, 2010, 2011; Wang et al, 2011; Neumann et al, 2012; He et al, 2015)

Summary

Introduction

Asterales is the most diverged order of dicots and includes 11 families and 27,000 species. Among the 11 families, Asteraceae is the largest and includes the sunflower and daisy. Sunflowers (Helianthus annuus L., 2n = 2x = 34, genome size = 2.43 Gb/haploid) are one of the most important crops in Asterales because their seeds can be used for oil production (Bennett et al, 1982). Sunflowers have been genetically and cytogenetically investigated (Feng et al, 2013), and a genome sequencing project is in progress (http://sunflowergenome.org/). At present, there is no genetic and cytogenetic marker for sunflower centromeres

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