ABSTRACT: The extent to which low-frequency (minor allele frequency (MAF) between 1-5%) and rare (MAF???1%) variants contribute to complex traits and disease in the general population is mainly unknown. Bone mineral density (BMD) is highly heritable, a major predictor of osteoporotic fractures, and has been previously associated with common genetic variants, as well as rare, population-specific, coding variants. Here we identify novel non-coding genetic variants with large effects on BMD (ntotal?=?53,236) and fracture (ntotal?=?508,253) in individuals of European ancestry from the general population. Associations for BMD were derived from whole-genome sequencing (n?=?2,882 from UK10K (ref. 10); a population-based genome sequencing consortium), whole-exome sequencing (n?=?3,549), deep imputation of genotyped samples using a combined UK10K/1000 Genomes reference panel (n?=?26,534), and de novo replication genotyping (n?=?20,271). We identified a low-frequency non-coding variant near a novel locus, EN1, with an effect size fourfold larger than the mean of previously reported common variants for lumbar spine BMD (rs11692564(T), MAF?=?1.6%, replication effect size?=?+0.20 s.d., Pmeta?=?2?×?10(-14)), which was also associated with a decreased risk of fracture (odds ratio?=?0.85; P?=?2?×?10(-11); ncases?=?98,742 and ncontrols?=?409,511). Using an En1(cre/flox) mouse model, we observed that conditional loss of En1 results in low bone mass, probably as a consequence of high bone turnover. We also identified a novel low-frequency non-coding variant with large effects on BMD near WNT16 (rs148771817(T), MAF?=?1.2%, replication effect size?=?+0.41 s.d., Pmeta?=?1?×?10(-11)). In general, there was an excess of association signals arising from deleterious coding and conserved non-coding variants. These findings provide evidence that low-frequency non-coding variants have large effects on BMD and fracture, thereby providing rationale for whole-genome sequencing and improved imputation reference panels to study the genetic architecture of complex traits and disease in the general population.
