Deleterious, protein-altering variants in the transcriptional coregulator ZMYM3 in 27 individuals with a neurodevelopmental delay phenotype

Abstract

Neurodevelopmental disorders (NDDs) result from highly penetrant variation in hundreds of different genes, some of which have not yet been identified. Using the MatchMaker Exchange, we assembled a cohort of 27 individuals with rare, protein-altering variation in the transcriptional coregulator ZMYM3, located on the X chromosome. Most (n = 24) individuals were males, 17 of which have a maternally inherited variant; six individuals (4 male, 2 female) harbor de novo variants. Overlapping features included developmental delay, intellectual disability, behavioral abnormalities, and a specific facial gestalt in a subset of males. Variants in almost all individuals (n = 26) are missense, including six that recurrently affect two residues. Four unrelated probands were identified with inherited variation affecting Arg441, a site at which variation has been previously seen in NDD-affected siblings, and two individuals have de novo variation resulting in p.Arg1294Cys (c.3880C>T). All variants affect evolutionarily conserved sites, and most are predicted to damage protein structure or function. ZMYM3 is relatively intolerant to variation in the general population, is widely expressed across human tissues, and encodes a component of the KDM1A-RCOR1 chromatin-modifying complex. ChIP-seq experiments on one variant, p.Arg1274Trp, indicate dramatically reduced genomic occupancy, supporting a hypomorphic effect. While we are unable to perform statistical evaluations to definitively support a causative role for variation in ZMYM3, the totality of the evidence, including 27 affected individuals, recurrent variation at two codons, overlapping phenotypic features, protein-modeling data, evolutionary constraint, and experimentally confirmed functional effects strongly support ZMYM3 as an NDD-associated gene.

Keywords: X-linked intellectual disability; ZMYM3; chromatin modifiers; neurodevelopmental disorder; transcriptional coregulators.

Figure 1

Observed variation along the length of ZMYM3 The 1,370 aa ZMYM3 (Q14202, GenBank: NP_005087.1) is annotated with MYM-type zinc fingers (1–9, orange) and Cre-like domain (blue) as described by UniProt and InterPro. (A) Hemizygous variants observed in males in this study are noted above the protein model, with de novo variants in red. ^aNote that p.Arg441Gln was observed in three unrelated males. Hemizygous variants that were previously reported in males are shown below the protein.^, (B) Maternally inherited (black) or de novo (red) heterozygous variants observed in females in this study are noted above the protein model.

Figure 2

Facial features of a subset of individuals with ZMYM3 variation Individual ID and protein effect are noted for each. Note deep-set eyes, long palpebral fissures, large/prominent/cupped ears, and tall forehead.

Figure 3

Missense variants in ZMYM3 mainly lie in ordered regions Three disordered regions (red line) were identified (aa 1–72, 90–301, and 759–830), while the remainder of the protein is predicted to be structured (green line). AlphaFold produces a per-residue confidence score (predicted local distance difference test, pLDDT) between 0 and 100, which is plotted along the length of the ZMYM3 protein. Horizontal bars and shading indicate confidence ranges for pLDDT scores. Missense variants observed here are noted on the graph, and while residues 69, 169, 241, and 302 lie in disordered regions, the remainder of residues lie in structured regions.

Figure 4

p.Arg1274Trp is a hypomorphic variant, while p.Arg688His has similar genome occupancy to that of wild type (A and B) Genomic regions called by csaw between experiments, then overlapped with IDR 0.05 peaks called in either experiment. Yellow color indicates regions determined by csaw to have significantly higher differential binding (at FDR < 0.05) in control, orange indicates regions with no differential binding, and red indicates regions with higher differential binding in variant. For p.Arg1274Trp (A), there are 6,631 regions with higher binding in control, 4,625 regions with no differential binding, and 3 regions with higher binding in variant. For p.Arg688His (B), all 6,416 regions had no differential binding. (C) Protein modeling of Arg1274 (left) and p.Arg1274Trp (right). The van der Waals protein surface is depicted in light gray, and residues in position 1,274 are colored by partial charge (blue, positive; red, negative; white, neutral). Magnified squares show zoomed-in view of the side chains. (D). Genome browser track for ZMYM3-p.Arg1274Trp-variant ChIP-seq experiments. Human genome (hg38) chr8:131,561,953–131,925,404 is displayed. Top track is activity-by-contact ("ABC loops") showing predicted interaction between enhancer element on left and TSS for the gene EFR3A on right. "Genes" track is RefSeq gene model. "Control Rep 1″ and "Control Rep 2″ are aligned bam reads from ZMYM3-p.Arg1274Trp-control experiments, "Variant Rep 1″ and "Variant Rep 2″ are aligned bam reads from ZMYM3-p.Arg1274Trp-variant experiments. All bam files are downsampled to an equal number of reads in each replicate, and all four replicate tracks are scaled from 0 to 60 vertically. "ENCODE" tracks are shown below the ChIP-seq tracks: "cCREs" represent candidate cis-regulatory elements colored by ENCODE standards, "H3K27Ac" is layered H3K27Ac signal from seven ENCODE cell lines, and "TFBSs" are ENCODE TF clusters (340 factors, 129 cell types). The putative enhancer element identified as the most significant loss of binding in the variant experiment is highlighted in yellow, showing the ENCODE distal enhancer cCRE call, the ABC loop to the TSS of EFR3A, and the difference in binding with the two control replicates showing strong signal and the two variant replicates showing substantially less binding.