The 3D architecture of the pepper genome and its relationship to function and evolution

ARTICLE https://doi.org/10.1038/s41467-022-31112-x OPEN The 3D architecture of the pepper genome and its relationship to function and evolution 1234567890():,; Yi Liao1,2,10, Juntao Wang 1,3,10, Zhangsheng Zhu1,3, Yuanlong Liu4,5,6, Jinfeng Chen7, Yongfeng Zhou Feng Liu9, Jianjun Lei1,3, Brandon S. Gaut 2, Bihao Cao 1,3 ✉, J. J. Emerson 2 ✉ & Changming Chen 8, 1,3 ✉ The organization of chromatin into self-interacting domains is universal among eukaryotic genomes, though how and why they form varies considerably. Here we report a chromosome-scale reference genome assembly of pepper (Capsicum annuum) and explore its 3D organization through integrating high-resolution Hi-C maps with epigenomic, transcriptomic, and genetic variation data. Chromatin folding domains in pepper are as prominent as TADs in mammals but exhibit unique characteristics. They tend to coincide with heterochromatic regions enriched with retrotransposons and are frequently embedded in loops, which may correlate with transcription factories. Their boundaries are hotspots for chromosome rearrangements but are otherwise depleted for genetic variation. While chromatin conformation broadly affects transcription variance, it does not predict differential gene expression between tissues. Our results suggest that pepper genome organization is explained by a model of heterochromatin-driven folding promoted by transcription factories and that such spatial architecture is under structural and functional constraints. 1 Key Laboratory of Biology and Genetic Improvement of Horticultural Crops (South China), Ministry of Agriculture and Rural Affairs, College of Horticulture, South China Agricultural University, Guangzhou 510642, China. 2 Department of Ecology and Evolutionary Biology, University of California, Irvine, CA 92697, USA. 3 Lingnan Guangdong Laboratory of Modern Agriculture, Guangzhou 510642, China. 4 Department of Computational Biology, University of Lausanne, Lausanne, Switzerland. 5 Swiss Cancer Center Leman, Lausanne, Switzerland. 6 Swiss Institute of Bioinformatics, Lausanne, Switzerland. 7 State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China. 8 Shenzhen Branch, Guangdong Laboratory of Lingnan Modern Agriculture, Genome Analysis Laboratory of the Ministry of Agriculture and Rural Affairs, Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen 518120, China. 9 College of Horticulture, Hunan Agricultural University, Changsha 410128, China. 10These authors contributed equally: Yi Liao, Juntao Wang. ✉email: ; ; NATURE COMMUNICATIONS | (2022)13:3479 | https://doi.org/10.1038/s41467-022-31112-x | www.nature.com/naturecommunications 1 ARTICLE T NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-022-31112-x he folding of chromosomes into self-interaction domains1, also known as topologically associating domains (TADs), appears to be conserved in evolution2. TADs and similar structures occur in diverse groups of eukaryotes, from fungi and bacteria to plants and animals3. Many mechanisms have been proposed for their formation, of which loop extrusion and compartmentalization are two leading models in animal systems4–7. While evidence suggests that these mechanisms may operate in tandem to jointly establish or maintain the spatial organization of the genome, the prevalence of each differs across species8–11. Like animals, TAD-like domains have been observed from Hi-C analyses of many plants; however, the mechanisms by which they form (and whether they are shared with animals) are largely unknown2,12. Additionally, TADs organized by different mechanisms may exhibit distinct structural and functional properties8,13–15. Thus, clarifying the formation mechanisms of TADs is necessary for further elucidating their functional specialization. Unlike in animals, where TADs can be readily detected genome-wide, small plant genomes like Arabidopsis thaliana and its close relative Arabidopsis lyrata carry few such domains16. However, other plant species with relatively large genome sizes do exhibit more pronounced chromatin domain architectures17–20. Comparisons between plant species imply that TAD prevalence in plants may be associated with genome size or other sequence properties, like the linear distribution of genes, regulatory elements, and transposable elements12,21,22. Consequently, 3D genome organization appears to exhibit great diversity in plants. This may also be true of the mechanisms that contribute to TADlike folding domain formation. For example, TAD-like domains in maize and tomato are reported to largely coincide with compartments, suggesting their formation is associated with compartmentalization in these species18. Recent studies in wheat19 have reported that a large proportion of chromatin domains are demarcated by gene-to-gene loops, and the genome is organized into regions of relatively high transcription-i.e. transcription factories23. Many other features such as transcription factors are also found to be associated with the formation of plant chromatin domains14,17. Thus, in plants, there appears to be variation not only in the prevalence of topological domains but also in their mechanism of formation. TADs are thought to behave as functional and structural units of the genome in evolution5. In metazoans, chromosomal rearrangement breakpoints rarely occur within TAD bodies, implying that disruption of TAD integrity is unfavorable and subject to purifying selection24–28. Chromatin structures are also found to be associated with patterns of both somatic mutation29 and genomic variants across evolutionary timescales30. Furthermore, long-range promoter-enhancer contacts that form loops are known to constrain large-scale genome evolution31. Given that the spatial organization of the genome affects organismal function, an open question in plant biology is: how does natural selection affect the acquisition and fate of mutations—particularly, structural variants—that alter spatial organization? In plants, even though 3D genome organization is thought to play an important role in the polyploidization process32–35, our understanding of the relationship between chromatin architecture and structural variants remains incomplete. Spatial genome organization is strongly associated with transcription. Numerous studies at the organismal31,36, tissue24, and cell type37–39 levels have established that rearrangement of 3D chromatin organization (i.e. higher-order chromatin structures, such as loops, TADs, and compartments) is associated with changes in gene expression. However, many studies suggest that chromatin conformation is not required for cis-regulatory interactions that activate normal gene expression40–42, and instead it 2 may primarily act as an architectural framework to facilitate gene regulation43. Although many recent attempts have been made to study t (...truncated)