Research ArticleEVOLUTIONARY BIOLOGY

Dosage analysis of the 7q11.23 Williams region identifies BAZ1B as a major human gene patterning the modern human face and underlying self-domestication

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Science Advances  04 Dec 2019:
Vol. 5, no. 12, eaaw7908
DOI: 10.1126/sciadv.aaw7908
  • Fig. 1 BAZ1B KD impairs migration and induction of patient-specific iPSC-derived NCSCs.

    (A) Schematic representation of the KD strategy on our iPSC-derived NCSC cohort. (B) BAZ1B mRNA levels in all the interfered lines (scr, sh1, and sh2) as measured by qPCR. Data represent aggregates of samples with the same number of BAZ1B copies (7dup, CTL + atWBS, and WBS). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is used as a normalizer. (C) Eight- and 16-hour time points from the wound-healing assay analyses performed on a 7dupASD and a WBS NCSC line upon BAZ1B KD. Cells from the same line infected with the scr sh were used as references for the migration (n = 2). (D) Days 7, 10, and 12 of NC differentiation from embryoid bodies (EBs) of an scr-interfered iPSC line and its respective BAZ1B KD (n = 3). (E) mRNA levels of NC markers at day 12 of differentiation in three individual experimental replicates [bright-field images are reported in (D)]. An iPSC line is included as a negative control. Student’s t test was used (ns, not significant; *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0. 0001).

  • Fig. 2 BAZ1B KD is responsible for transcriptional alterations in NC-related pathways.

    (A) Overlap between genes directly or inversely following BAZ1B levels identified in the pairwise comparative analysis (scr versus shBAZ1B) and in the regression analysis on BAZ1B-level sensitive genes on iPSC-derived NCSCs (FDR < 0.1). (B) Volcano plot reporting DEGs identified in the RNA-seq analysis on iPSC-derived NCSCs [fold change (FC) > 1.25; FDR < 0.1]. (C) Top most specific enrichments for GO biological processes among the DEGs in the RNA-seq analysis on iPSC-derived NCSCs. (D) Heat map representing DEGs that are dysregulated in genetic disorders involving mental retardation (“Mental”), intellectual disability (“Intellectual”), and/or facial dysmorphisms (“Face”) according to OMIM database classification. cnv, copy number variant. (E) Putative regulators of genes that follow BAZ1B levels identified by a master regulator analysis. Regulators were divided in four different groups based on their main functions.

  • Fig. 3 BAZ1B preferentially binds its targets at their enhancer regions and its KD causes a redistribution of enhancer histone marks.

    (A) Schematic representation of the strategy for CRISPR-Cas9–mediated tagging of endogenous BAZ1B. Briefly, iPSCs from the four genotypes were electroporated with the donor plasmid and the Cas9/single-guide RNA ribonucleoprotein complex; clones were selected via hygromycin and PCR, differentiated to NCSCs, and then subjected to ChIP-seq. (B) PCA showing the distribution of the four BAZ1B-tagged NCSC lines according to their chromatin profiles. (C) Overlap between genes expressed in our NCSC lines (purple) and genes bound by BAZ1B at their enhancer (red) or promoter (blue) regions. (D) Top most specific enrichments for GO biological processes among the genes that are bound by BAZ1B and expressed in our NCSC cohort. (E) Most represented BAZ1B DNA binding motifs identified by HOMER show high similarity to neural and NCSC-specific transcription factors motifs. (F) BAZ1B differentially bound regions according to its copy number (FDR < 0.1; n = 2). (G) Overlap between genes that are differentially expressed have their enhancers differentially marked concordantly (H3K27ac, H3K4me1, and H3K27me3) and are bound by BAZ1B at enhancers.

  • Fig. 4 Exploration of paleogenomic datasets supports a key evolutionary role for BAZ1B and validates the self-domestication hypothesis.

    (A) Archaic (Neanderthal) and modern skulls, illustrating the target domesticated phenotype that was captured by our analysis. Skull images were derived from work under a CC BY-SA 2.0 license (https://creativecommons.org/licenses/by-sa/2.0/deed.en) by hairymuseummatt. (B) Overlap between BAZ1B level–sensitive genes and datasets, which bring out differences between AMHs and archaics, as well as genes under positive selection in modern humans and domesticates. (C) Barplots showing the occurrence of high-frequency changes, missense mutations, and mutations in regulatory regions in genes from the AMH (nearly) fixed mutation dataset (pink Venn in B). (D) Heat map representing the amount of overlaps for each list selected from (B). Gene overlaps and detailed list descriptions are reported in table S2. (E) Rendering of a typical WBS face (left) against the background of a typical modern face (right). Red segments indicate areas of the lower face where the two faces most sharply depart (nose, philtrum, and lower front of the mandible). The lower midface region is most often associated with mutations in genes figuring prominently in our intersections, as discussed in the text and table S3.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/12/eaaw7908/DC1

    Fig. S1. BAZ1B KD validation in iPSC-derived NCSCs and evaluation of its impact on NCSC migration.

    Fig. S2. BAZ1B KD affects the transcriptome of iPSC-derived NCSCs.

    Fig. S3. Generation of BAZ1B-FLAG iPSC lines and differentiation to NCSCs.

    Fig. S4. BAZ1B KD induces a significant chromatin remodeling at distal regions.

    Text S1A. Detailed description of HOMER motif enrichments performed on BAZ1B ChIP-seq data.

    Text S1B. List of key direct targets of BAZ1B involved in neural- and NC-related development and relevant associated literature.

    Table S1. Genes relevant for NC and NC-derived features whose expression follows BAZ1B levels.

    Table S2A. Significant genes in human evolution.

    Table S2B. Regulatory excess in archaic humans, overlap with BAZ1B targets.

    Table S2C. Mutation excess in archaic humans, overlap with BAZ1B targets.

    Table S2D. Regulatory changes (exclusive) in archaic humans, overlap with BAZ1B targets.

    Table S2E. Missense mutations in archaic humans, overlap with BAZ1B targets.

    Table S2F. Mutation excess in archaic humans corrected for length, overlap with BAZ1B targets.

    Table S2G. Regulatory excess in modern humans, overlap with BAZ1B targets.

    Table S2H. Mutation excess in modern humans, overlap with BAZ1B targets.

    Table S2I. Regulatory changes (exclusive) in modern humans, overlap with BAZ1B targets.

    Table S2J. Missense mutations in modern humans, overlap with BAZ1B targets.

    Table S2K. Mutation excess in modern humans corrected for length, overlap with BAZ1B targets.

    Table S2L. Genes under positive selection in domesticated animals, overlap with BAZ1B targets.

    Table S2M. Genes under positive selection from Peyrégne et al. (13) in modern humans, overlap with BAZ1B targets.

    Table S2N. Genes under positive selection from Racimo (14) in modern humans, overlap with BAZ1B targets.

    Table S3. Crucial genes identified in the overlap between BAZ1B datasets and archaic versus modern human datasets reported in this study.

    Table S4. Alternative differential expression analysis functions tested with iPSCpower to assess the efficacy of our design matrix (~individual+BAZ1B). R code provided.

    Table S5. Number of genes differentially expressed following BAZ1B data in our numerical analysis compared to an analysis conducted on randomized HipSci data, using Edg2 function (see table S4).

    References (6189)

  • Supplementary Materials

    The PDFset includes:

    • Fig. S1. BAZ1B KD validation in iPSC-derived NCSCs and evaluation of its impact on NCSC migration.
    • Fig. S2. BAZ1B KD affects the transcriptome of iPSC-derived NCSCs.
    • Fig. S3. Generation of BAZ1B-FLAG iPSC lines and differentiation to NCSCs.
    • Fig. S4. BAZ1B KD induces a significant chromatin remodeling at distal regions.
    • Text S1A. Detailed description of HOMER motif enrichments performed on BAZ1B ChIP-seq data.
    • Text S1B. List of key direct targets of BAZ1B involved in neural- and NC-related development and relevant associated literature.
    • Table S1. Genes relevant for NC and NC-derived features whose expression follows BAZ1B levels.
    • Table S3. Crucial genes identified in the overlap between BAZ1B datasets and archaic versus modern human datasets reported in this study.
    • Table S4. Alternative differential expression analysis functions tested with iPSCpower to assess the efficacy of our design matrix (~individual+BAZ1B). R code provided.
    • Table S5. Number of genes differentially expressed following BAZ1B data in our numerical analysis compared to an analysis conducted on randomized HipSci data, using Edg2 function (see table S4).
    • References (6189)

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    Other Supplementary Material for this manuscript includes the following:

    • Table S2A (Microsoft Excel format). Significant genes in human evolution.
    • Table S2B (Microsoft Excel format). Regulatory excess in archaic humans, overlap with BAZ1B targets.
    • Table S2C (Microsoft Excel format). Mutation excess in archaic humans, overlap with BAZ1B targets.
    • Table S2D (Microsoft Excel format). Regulatory changes (exclusive) in archaic humans, overlap with BAZ1B targets.
    • Table S2E (Microsoft Excel format). Missense mutations in archaic humans, overlap with BAZ1B targets.
    • Table S2F (Microsoft Excel format). Mutation excess in archaic humans corrected for length, overlap with BAZ1B targets.
    • Table S2G (Microsoft Excel format). Regulatory excess in modern humans, overlap with BAZ1B targets.
    • Table S2H (Microsoft Excel format). Mutation excess in modern humans, overlap with BAZ1B targets.
    • Table S2I (Microsoft Excel format). Regulatory changes (exclusive) in modern humans, overlap with BAZ1B targets.
    • Table S2J (Microsoft Excel format). Missense mutations in modern humans, overlap with BAZ1B targets.
    • Table S2K (Microsoft Excel format). Mutation excess in modern humans corrected for length, overlap with BAZ1B targets.
    • Table S2L (Microsoft Excel format). Genes under positive selection in domesticated animals, overlap with BAZ1B targets.
    • Table S2M (Microsoft Excel format). Genes under positive selection from Peyrégne et al. (13) in modern humans, overlap with BAZ1B targets.
    • Table S2N (Microsoft Excel format). Genes under positive selection from Racimo (14) in modern humans, overlap with BAZ1B targets.

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