Fig. 7. scATAC-trained convolutional neural network accurately predicts cell type-specific accessibility status and human mutation effects in a transiently developing cell type.

a Neural net predicted chromatin accessibility profiles (red) compared to actual scATAC sequencing coverage (black) for a region of mouse chromosome 6 in three cell types (cMN7 e10.5, cMN7 e11.5, and cMN12 e11.5). The gray box highlights a transient 678 bp peak (cRE2) that is accessible in cMN7 e10.5, but not cMN7 e11.5 or cMN12 e11.5. SNVs within the human orthologous peak cRE2 cause congenital facial weakness, a disorder of cMN7. b Neural net-trained in silico saturation mutagenesis predictions for specific nucleotide changes in human cRE2 for cMN7 e10.5, cMN7 e11.5, and cMN12 e11.5. Predicted loss-of-function nucleotide changes are colored in blue and gain-of-function in red. Predictions for four known loss-of-function pathogenic variants (chr3:128178260 G > C, chr3:128178261 G > A, chr3:128178262 T > C, chr3:128178262 T > G) are boxed. All four pathogenic variants are predicted loss-of-function for cMN7 e10.5, but not cMN7 e11.5 or cMN12 e11.5. c Pseudobulk accessibility profiles of cRE2 (red box) CN7 e10.5 for wildtype and two CRISPR-mutagenized mouse lines (cRE2^Fam4/Fam4 and cRE2^Fam5/Fam5) show a qualitative reduction in cRE2 scATAC sequencing coverage, consistent with in silico saturation mutagenesis predictions. Each pseudobulk profile represents normalized sequencing coverage across two biological replicates. d Locus-specific footprinting evidence overlapping cRE2. A 792 bp window showing sequencing coverage for cMN7 e10.5 after correcting for Tn5 insertion bias. The NR2F1 transcription factor binding site is mutated in individuals with HCFP1-CFP and overlaps a local minimum in scATAC coverage. TOBIAS footprinting scores for cRE2 wildtype, cRE2^Fam4/Fam4, and cRE2 ^Fam5/Fam5 are depicted in solid, dashed, and dotted lines, respectively. e. Stacked barplot depicting wildtype (gray) versus mutant (blue) scATAC read counts over a 7.7 kb window for cMN7 e10.5 in cRE2^WT/Fam5 heterozygote embryos. cRE2 mutant alleles are consistently depleted across two biological replicates (counts_WT / counts_MUTANT = 4.21; p value = 2.4 × 10⁻¹⁴, binomial test). The mean expected value under the null is depicted by a solid black line for each experiment. Scale bars in f and i = 500 μm and are approximate measurements based on E11.5 embryo average crown-rump length of 6 mm. Source data are provided as a Source Data file.