SUMMARYHow mutations in gene regulatory elements lead to evolutionary changes remains largely unknown. Human accelerated regions (HARs) are ideal for exploring this question, because they are associated with human-specific traits and contain multiple human-specific variants at sites conserved across mammals, suggesting that they alter or compensate to preserve function. We performed massively parallel reporter assays on all human and chimpanzee HAR sequences in human and chimpanzee iPSC-derived neural progenitors at two differentiation stages. Forty-three percent (306/714) of HARs function as neuronal enhancers, with two-thirds (204/306) showing consistent changes in activity between human and chimpanzee sequences.These changes were almost all sequence dependent and not affected by cell species or differentiation stage. We tested all evolutionary intermediates between human and chimpanzee sequences of seven HARs, finding variants that interact both positively and negatively. This study shows that variants acquired during human evolution interact to buffer and amplify changes to enhancer function.

31Dynamic changes in chromatin accessibility coincide with important aspects of neuronal differentiation, such as 32 fate specification and arealization and confer cell type-specific associations to neurodevelopmental disorders. 33However, studies of the epigenomic landscape of the developing human brain have yet to be performed at single-34 cell resolution. Here, we profiled chromatin accessibility of >75,000 cells from eight distinct areas of developing 35human forebrain using single cell ATAC-seq (scATACseq). We identified thousands of loci that undergo 36 extensive cell type-specific changes in accessibility during corticogenesis. Chromatin state profiling also reveals 37 novel distinctions between neural progenitor cells from different cortical areas not seen in transcriptomic profiles 38 and suggests a role for retinoic acid signaling in cortical arealization. Comparison of the cell type-specific 39 chromatin landscape of cerebral organoids to primary developing cortex found that organoids establish broad 40 cell type-specific enhancer accessibility patterns similar to the developing cortex, but lack many putative 41 regulatory elements identified in homologous primary cell types. Together, our results reveal the important 42 contribution of chromatin state to the emerging patterns of cell type diversity and cell fate specification and 43 provide a blueprint for evaluating the fidelity and robustness of cerebral organoids as a model for cortical 44 development. 45 46Main text 47The diverse cell types of the human cerebral cortex (Fig. 1a) have been mostly classified based on a handful of 48 morphological, anatomical, and physiological features. Recent innovations in single cell genomics, such as single 49 cell mRNA sequencing (scRNA-seq), have enabled massively parallel profiling of thousands of molecular 50 features in every cell, uncovering the remarkable molecular diversity of cell types previously considered 51 homologous, such as excitatory neurons located in different areas of the cerebral cortex 1-6 . However, the 52 developmental mechanisms underlying the emergence of distinct cellular identities are largely unknown, as most 53 cortical neurons are generated at stages that are inaccessible to experimentation 5 . 54 55Over 60 years ago, Conrad Waddington introduced the concept of an epigenomic landscape to account for the 56 emergence of distinct cell fates 7 . In particular, chromatin state defines the functional architecture of the genome 57

16During meiotic prophase, chromosomes organise into a series of chromatin loops emanating from 17 a proteinaceous axis, but the mechanisms of assembly remain unclear. Here we elucidate how 18 this elaborate three-dimensional chromosome organisation is underpinned by genomic sequence 19 in Saccharomyces cerevisiae. Entering meiosis, strong cohesin-dependent grid-like Hi-C 20 interaction patterns emerge, reminiscent of mammalian interphase organisation, but with distinct 21 regulation. Meiotic patterns agree with simulations of loop extrusion limited by barriers, yet are 22 patterned by convergent transcription rather than binding of the mammalian interphase factor, 23 CTCF, which is absent in S. cerevisiae-thereby both challenging and extending current 24 paradigms of local chromosome organisation. While grid-like interactions emerge independently 25 of meiotic chromosome synapsis, synapsis itself generates additional compaction that matures 26 differentially according to telomere proximity and chromosome size. Collectively, our results 27 elucidate fundamental principles of chromosome assembly and demonstrate the essential role of 28 cohesin within this evolutionarily conserved process. 29 Schalbetter, Fudenberg et al. 2018 Introduction and Results 30 During meiosis, eukaryotic chromosomes are broken, repaired and paired with their homologs 31 followed by two rounds of segregation-a series of events accompanied by dynamic structural 32 changes of the chromosomes. Most prominent is the paired arrangement of pachytene 33 chromosomes into a dense array of chromatin loops emanating from proteinaceous axes linked 34 by a central core, the synaptonemal complex (SC), which is highly conserved across 35 eukaryotes 1,2 . In S. cerevisiae, structural components include the meiotic cohesin kleisin subunit, 36 Rec8 3 , the transverse filament, Zip1 4 , the axial/lateral elements, Hop1 and Red1 5,6 , and the pro-37 DSB factors Rec114-Mei4-Mer2 (RMM) 7,8 . Much of our understanding of meiotic chromosome 38 structure has been deduced from a combination of electron microscopy, immunofluorescence 39 microscopy, and the genome-wide patterns of protein localisation determined by ChIP. However, 40 the link between key meiotic protein complexes, chromosome conformation, and genomic 41 sequence remains uncharacterized. 42 43 Chromosome conformation capture (3C) techniques generate maps of pairwise contact 44 frequencies that are snapshots of chromosome organisation. 3C methods were originally applied 45 to assay chromosome conformation in S. cerevisiae, including during meiosis 9 . Now they are 46 widely used across a range of organisms and cellular contexts to link 3D organisation directly with 47 53 54Starting with a synchronized G1 population we analysed timepoints encompassing DNA 55 replication, meiotic prophase and both meiotic divisions ( Fig. 1a,b,c, Extended Data Fig. 1a,b,c). 56 In G1, we detect strong centromere clustering ( Fig. 1a,d) and folding back of the arms at the 57 centromeres ( Fig. 1a, Supplementary Fig...

40We present the Metagenomic Intra-species Diversity Analysis System (MIDAS), which is an 41 integrated computational pipeline for quantifying bacterial species abundance and strain-42 level genomic variation, including gene content and single nucleotide polymorphisms, from 43 shotgun metagenomes. Our method leverages a database of >30,000 bacterial reference 44 genomes which we clustered into species groups. These cover the majority of abundant 45 species in the human microbiome but only a small proportion of microbes in other 46 environments, including soil and seawater. We applied MIDAS to stool metagenomes from 4798 Swedish mothers and their infants over one year and used rare single nucleotide 48 variants to reveal extensive vertical transmission of strains at birth but colonization with 49 strains unlikely to derive from the mother at later time points. This pattern was missed 50with species-level analysis, because the infant gut microbiome composition converges 51 towards that of an adult over time. We also applied MIDAS to 198 globally distributed 52 marine metagenomes and used gene content to show that many prevalent bacterial species 53 have population structure that correlates with geographic location. Strain-level genetic 54 variants present in metagenomes clearly reveal extensive structure and dynamics that are 55 obscured when data is analyzed at a higher taxonomic resolution. 56 57