A 10,000-year transect of Roman populations: Rome wasn't built (or settled) in a day. Antonio et al. performed an ancestral DNA analysis to investigate the genetic changes that occurred in Rome and central Italy from the Mesolithic into modern times. By examining 127 Roman genomes and their archaeological context, the authors demonstrate a major ancestry shift in the Neolithic between hunter gatherers and farmers. A second ancestry shift is observed in the Bronze Age, likely coinciding with trade and an increased movement of populations. Genetic changes track the historical changes occurring in Rome and reflect gene flow from across the Mediterranean, Europe, and North Africa over time. Science , this issue p. 708

Significance: More than three-fourths of human germline mutations are paternal in origin and their total number increases with the father’s age at conception. These observations are thought to support the textbook view that germline point mutations stem mostly from DNA replication errors. Analyzing large germline mutation datasets for humans, we find that this understanding cannot explain the observed patterns of new mutations. Instead, we show that the male mutation bias is not driven by spermatogenesis. We further find evidence that a substantial fraction of mutations are not replicative in origin and uncover a potential effect of a mother’s age on the number of mutations that happen early in the development of the embryo.

Mutations can originate from the chance misincorporation of nucleotides during DNA replication or from DNA lesions that arise between replication cycles and are not repaired correctly. We introduce a model that relates the source of mutations to their accumulation with cell divisions, providing a framework for understanding how mutation rates depend on sex, age, and cell division rate. We show that the accrual of mutations should track cell divisions not only when mutations are replicative in origin but also when they are non-replicative and repaired efficiently. One implication is that observations from diverse fields that to date have been interpreted as pointing to a replicative origin of most mutations could instead reflect the accumulation of mutations arising from endogenous reactions or exogenous mutagens. We further find that only mutations that arise from inefficiently repaired lesions will accrue according to absolute time; thus, unless life history traits co-vary, the phylogenetic “molecular clock” should not be expected to run steadily across species. We relate how mutations arise to how they accumulate in different sexes, with age and with cell division. This model provides a single framework within which to interpret emerging results from evolutionary biology, human genetics, and cancer genetics. We show that the accrual of mutations should track cell divisions not only when mutations originate during DNA replication but also when they arise through non-replicative mechanisms and are repaired efficiently. This realization means that previous observations of correlations between mutation and cell division rates actually provide little support to the commonly held belief that most germline and somatic mutations arise from replication errors. We further find that only mutations that arise from inefficiently repaired lesions will accrue according to absolute time; thus, without covariation in life history traits, the phylogenetic “molecular clock” should not be expected to run at constant rates across species.

The number of de novo mutations (DNMs) found in an offspring's genome increases with both paternal and maternal age. But does the rate of mutation accumulation in human gametes differ across families? Using sequencing data from 33 large, three-generation CEPH families, we observed significant variability in parental age effects on DNM counts across families, ranging from 0.19 to 3.24 DNMs per year. Additionally, we found that ~3% of DNMs originated following primordial germ cell specification in a parent, and differed from non-mosaic germline DNMs in their mutational spectra. We also discovered that nearly 10% of candidate DNMs in the second generation were post-zygotic, and present in both somatic and germ cells; these gonosomal mutations occurred at equivalent frequencies on both parental haplotypes. Our results demonstrate that rates of germline mutation accumulation vary among families with similar ancestry, and confirm that post-zygotic mosaicism is a substantial source of human DNM.

Our understanding of the chronology of human evolution relies on the “molecular clock” provided by the steady accumulation of substitutions on an evolutionary lineage. Recent analyses of human pedigrees have called this understanding into question by revealing unexpectedly low germline mutation rates, which imply that substitutions accrue more slowly than previously believed. Translating mutation rates estimated from pedigrees into substitution rates is not as straightforward as it may seem, however. We dissect the steps involved, emphasizing that dating evolutionary events requires not “a mutation rate” but a precise characterization of how mutations accumulate in development in males and females—knowledge that remains elusive.

Gene expression variation is a major contributor to phenotypic variation in human complex traits. Selection on complex traits may therefore be reflected in constraint on gene expression. Here, we explore the effects of stabilizing selection on cis-regulatory genetic variation in humans. We analyze patterns of expression variation at copy number variants and find evidence for selection against large increases in gene expression. Using allele-specific expression (ASE) data, we further show evidence of selection against smaller-effect variants. We estimate that, across all genes, singletons in a sample of 122 individuals have ∼2.2× greater effects on expression variation than the average variant across allele frequencies. Despite their increased effect size relative to common variants, we estimate that singletons in the sample studied explain, on average, only 5% of the heritability of gene expression from cis-regulatory variants. Finally, we show that genes depleted for loss-of-function variants are also depleted for cis-eQTLs and have low levels of allelic imbalance, confirming tighter constraint on the expression levels of these genes. We conclude that constraint on gene expression is present, but has relatively weak effects on most cis-regulatory variants, thus permitting high levels of gene-regulatory genetic variation.