Modeling uniquely human gene regulatory function via targeted humanization of the mouse genome

Dutrow, Emily V.; Emera, Deena; Yim, Kristina; Uebbing, Severin; Kocher, Acadia A.; Krenzer, Martina; Nottoli, Timothy; Burkhardt, Daniel; Krishnaswamy, Smita; Louvi, Angeliki; Noonan, James P.

doi:10.1038/s41467-021-27899-w

Cited by 25 publications

(19 citation statements)

References 80 publications

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“…This sequence is located in an intron of the gene, and actually lies within a human craniofacial superenhancer region ( Figure 5A ), suggesting its role in the craniofacial apparatus. Despite its proximity to known craniofacial regulatory regions, HACNS1 has been proposed to regulate Gbx2 in the mouse limb ( 113 ). However, this gene is expressed at relatively low levels in human craniofacial tissues ( 97 ) and the mouse molar compared to Agap1 .…”

Section: Resultsmentioning

confidence: 99%

Integration of multimodal data in the developing tooth reveals candidate regulatory loci driving human odontogenic phenotypes

Winchester

Cotney

2022

Front. Dent. Med

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Human odontogenic aberrations such as abnormal tooth number and delayed tooth eruption can occur as a symptom of rare syndromes or, more commonly, as nonsyndromic phenotypes. These phenotypes can require extensive and expensive dental treatment, posing a significant burden. While many dental phenotypes are heritable, most nonsyndromic cases have not been linked to causal genes. We demonstrate the novel finding that common sequence variants associated with human odontogenic phenotypes are enriched in developmental craniofacial enhancers conserved between human and mouse. However, the bulk nature of these samples obscures if this finding is due to the tooth itself or the surrounding tissues. We therefore sought to identify enhancers specifically active in the tooth anlagen and quantify their contribution to the observed genetic enrichments. We systematically identified 22,001 conserved enhancers active in E13.5 mouse incisors using ChIP-seq and machine learning pipelines and demonstrated biologically relevant enrichments in putative target genes, transcription factor binding motifs, and in vivo activity. Multi-tissue comparisons of human and mouse enhancers revealed that these putative tooth enhancers had the strongest enrichment of odontogenic phenotype-associated variants, suggesting a role for dysregulation of tooth developmental enhancers in human dental phenotypes. The large number of these regions genome-wide necessitated prioritization of enhancer loci for future investigations. As enhancers modulate gene expression, we prioritized regions based on enhancers' putative target genes. We predicted these target genes and prioritized loci by integrating chromatin state, bulk gene expression and coexpression, GWAS variants, and cell type resolved gene expression to generate a prioritized list of putative odontogenic phenotype-driving loci active in the developing tooth. These genomic regions are of particular interest for downstream experiments determining the role of specific dental enhancer:gene pairs in odontogenesis.

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Section: Resultsmentioning

confidence: 99%

Integration of multimodal data in the developing tooth reveals candidate regulatory loci driving human odontogenic phenotypes

Winchester

Cotney

2022

Front. Dent. Med

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“…This sequence is located in an intron of the gene, and actually lies within a human craniofacial superenhancer region (Figure 5A), suggesting its role in the craniofacial apparatus. Despite its proximity to known craniofacial regulatory regions, HACNS1 has been proposed to regulate Gbx2 in the mouse limb [98]. However, this gene is expressed at relatively low levels in human craniofacial tissues [70] and the mouse molar compared to Agap1 (Supplemental Figure 9).…”

Section: Resultsmentioning

confidence: 99%

“Integration of multimodal data in the developing tooth reveals candidate dental disease genes”

Winchester

Cotney

2022

Preprint

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Dental malformations and diseases affect a large number of people in their lifetime. Their widespread occurrence and the documented influence of oral health on systemic health make understanding their risk factors a public health priority. Previous studies have shown that common dental phenotypes are heritable, but the genetic causes of most cases have not been identified. Loci that harbor risk for human craniofacial morphology and related diseases are enriched genome-wide in gene regulatory sequences, specifically enhancers, active during human embryonic craniofacial development. We demonstrated here that these enhancers also show enrichment of dental phenotype-associated variants, which is conserved in embryonic mouse craniofacial enhancers. Given these findings in bulk craniofacial tissues, we looked to determine the role of tooth enhancers in this phenomenon. We performed ChIP-seq and functionally annotated the genome of E13.5 mouse incisors, identifying enhancers. Multi-tissue comparisons of human and mouse enhancers revealed that putative tooth enhancers had the strongest enrichment of variants associated with dental phenotypes. These findings suggested a role for dysregulation of tooth development in dental phenotypes and diseases. To uncover novel dental phenotype-driving genes in the developing tooth we performed coexpression analysis on E12.5-E17.5 mouse incisors and molars, and annotated the contributing cell types of gene modules using scRNAseq of E14 molars. Through integration of chromatin state, bulk gene coexpression, and cell type resolved gene expression we prioritized a list of candidate novel dental disease genes for future investigations in mouse models and human studies.

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“…Sometimes termed ‘humanization’, this process narrowly refers to engineering human variants in a single locus and should not be construed as general humanization of an animal model. Using this approach, human HACNS1 variants were shown to increase Gbx2 expression in distal limbs as predicted by reporter assays, but morphological changes could not be detected using current techniques 159 . Similarly, introduction of mutations that evolved in the common ancestor of humans and chimpanzees into a regulatory region of the mouse Cbln2 gene increased the expression of Cbln2 in cortical excitatory neurons.…”

Section: Models For Functional Studiesmentioning

confidence: 97%

Human-specific genetics: new tools to explore the molecular and cellular basis of human evolution

et al. 2023

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Our ancestors acquired morphological, cognitive and metabolic modifications that enabled humans to colonize diverse habitats, develop extraordinary technologies and reshape the biosphere. Understanding the genetic, developmental and molecular bases for these changes will provide insights into how we became human. Connecting human-specific genetic changes to species differences has been challenging owing to an abundance of low-effect size genetic changes, limited descriptions of phenotypic differences across development at the level of cell types and lack of experimental models. Emerging approaches for single-cell sequencing, genetic manipulation and stem cell culture now support descriptive and functional studies in defined cell types with a human or ape genetic background. In this Review, we describe how the sequencing of genomes from modern and archaic hominins, great apes and other primates is revealing human-specific genetic changes and how new molecular and cellular approaches — including cell atlases and organoids — are enabling exploration of the candidate causal factors that underlie human-specific traits.

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Modeling uniquely human gene regulatory function via targeted humanization of the mouse genome

Cited by 25 publications

References 80 publications

Integration of multimodal data in the developing tooth reveals candidate regulatory loci driving human odontogenic phenotypes

Integration of multimodal data in the developing tooth reveals candidate regulatory loci driving human odontogenic phenotypes

“Integration of multimodal data in the developing tooth reveals candidate dental disease genes”

Human-specific genetics: new tools to explore the molecular and cellular basis of human evolution

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