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Avena sativa/Zea mays addition line

ABSTRACT: ChIP-seq in oat-maize additional lines using CenH3 antibody

PROVIDER: PRJNA205183 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR867049_1.fastq.gz	Fastqsanger.gz
	SRR867049_2.fastq.gz	Fastqsanger.gz
	SRR867050_1.fastq.gz	Fastqsanger.gz
	SRR867050_2.fastq.gz	Fastqsanger.gz
	SRR867051_1.fastq.gz	Fastqsanger.gz

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ChIP-seq in oat-maize additional lines using CenH3 antibody

Project description:Many existing centromeres may have originated as neocentromeres that activated de novo from non-centromeric regions. However, the evolutionary path from a neocentromere to a mature centromere has been elusive. Here we analyzed the centromeres of six chromosomes that were transferred from maize into oat as the result of an inter-species cross. Centromere size and location were assayed by chromatin immunoprecipitation for the histone variant CENH3, which is a defining feature of functional centromeres. Maize and oat are highly divergent and differ in genome size by four fold. Two isolates of maize chromosome proved to contain neocentromeres in the sense that they had moved from the original site, whereas the remaining seven centromeres (1, 2, 5, 6, 8, 9 and 10) were retained in the same area in both species. In all cases the CENH3-binding domains were dramatically expanded to encompass a larger area in the oat background (~4 Mb) than the average centromere size in maize (~2 Mb). The expansion of maize centromeres appeared to be restricted by the transcription of genes located in regions flanking the original centromeres. The results from the current study provide evidence that (1) centromere size is regulated; (2) centromere sizes tend to be uniform within a species regardless of chromosome size or origin of the centromere; and (3) neocentromeres emerge and expand preferentially in gene poor regions. Our results, together with data from several animal species, suggest that centromere size expansion may be a key factor in the survival of neocentric chromosomes in natural populations.

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Project description:The centromere, as an essential element to control chromosome segregation, is epigenetically determined by CENH3-containing nucleosomes as a functional marker, therefore the accurate deposition of CENH3 is crucial to chromosome transmission. We characterized the deposition of CENH3 in maize by over-expression and mutational analysis. Our results revealed that over-expressing CENH3 in callus is lethal while over-expressing GFP-CENH3 and CENH3-YFP in callus and plants is not and can be partly deposited normally. Different mutations of GFP-CENH3 demonstrated that CENH3-Thr4 in the N terminus was needed for the deposition as a positive phosphorylation site and the last five amino acids in the C terminus are necessary for deposition. The C terminal tail of CENH3 is confirmed to be responsible for the interaction of CENH3 and histone H4, which indicates that CENH3 maintains deposition in centromeres via interacting with H4 to form stable nucleosomes. For GFP-CENH3 and CENH3-YFP, the fused tags at the termini probably affect the structure of CENH3 and reduce its interaction with other proteins, which in turn could decrease proper deposition. Taken together, multiple amino acids or motifs were shown to play essential roles in CENH3 deposition, which is suggested to be affected by numerous factors in maize.

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Project description:To determine the centromere of the maize B chromosome, we used previously published anti-CENH3-ChIP-seq data from TB-9Sb, which contain a complete functional B centromere. Distribution of centromere-specific DNA repeats, including CentC, CRM element and B-repeat, were observed in the proximal end of the assembled maize B chromosome, and this region was shown to be associated with CENH3 nucleosomes. Furthermore, six small scaffolds with sizes ranging from 10 to 174 kilobase display CENH3 enrichment, also with the distribution of these repeat sequences. These results were consistent with previously cytogenetic observation. Therefore, approximately 520 kb centromeric regions were determined in the assembled maize B chromosome.

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