Supplemental Material for Heft et al., 2020 Ilea Heft Yulia Mostovoy Walfred Ma Aaron J. Stevens Steven Pastor Jennifer McCaffery Dario Boffelli David Martin Ming Xiao Martin Kennedy Pui-Yan Kwok James M. Sikela 10.25386/genetics.10673792.v1 https://gsajournals.figshare.com/articles/dataset/Supplemental_Material_for_Heft_et_al_2020/10673792 <p>Sequences encoding Olduvai protein domains (formerly DUF1220) show the greatest human lineage-specific increase in copy number of any coding region in the genome and have been associated, in a dosage-dependent manner, with brain size, cognitive aptitude, autism, and schizophrenia. Tandem intragenic duplications of a three-domain block, termed the Olduvai triplet, in four <i>NBPF</i> genes in the chromosomal 1q21.1-.2 region are primarily responsible for the striking human-specific copy number increase. Interestingly, most of the Olduvai triplets are adjacent to, and transcriptionally co-regulated with, three human-specific <i>NOTCH2NL</i> genes that have been shown to promote cortical neurogenesis. Until now, the underlying genomic events that drove the Olduvai hyper-amplification in humans have remained unexplained. Here, we show that the presence or absence of an alternative first exon of the Olduvai triplet perfectly discriminates between amplified (58/58) and unamplified (0/12) triplets. We provide sequence and breakpoint analyses that suggest the alternative exon was produced by an NAHR-based mechanism involving the duplicative transposition of an existing Olduvai exon found in the CON3 domain that typically occurs at the carboxy end of <i>NBPF</i> genes. We also provide suggestive <i>in vitro</i> evidence that the alternative exon may promote instability through a putative G-quadraplex-based (pG4) mechanism. Lastly, we use single-molecule optical mapping to characterize the intragenic structural variation observed in <i>NBPF</i> genes in 154 unrelated individuals and 52 related individuals from 16 families and show that the presence of pG4-containing Olduvai triplets is strongly correlated with high levels of Olduvai copy number variation. These results suggest that the same driver of genomic instability that allowed the evolutionarily recent, rapid, and extreme human-specific Olduvai expansion remains highly active in the human genome.</p> 2019-11-21 18:46:58 Genome Evolution Protein Domain Olduvai DUF1220 Gene Duplication Human Brain Evolution Genome Structural Variation Population, Ecological and Evolutionary Genetics Quantitative Genetics (incl. Disease and Trait Mapping Genetics) Neurogenetics Molecular Evolution Genomics Genome Structure and Regulation Genetics Evolutionary Biology