Assembly-independent measurement of human Yq12 heterochromatin reveals population-scale variation and Yq12-derived RNA in testis

Abstract

Despite advances in long-read sequencing and telomere-to-telomere assembly, population-scale analysis of large satellite arrays remains challenging. Human Yq12 is a megabase-scale heterochromatic region composed primarily of DYZ1/HSat3A6 and DYZ2/HSat1B repeats, whose near-equal proportions are maintained despite extensive variation in total array length. Here, using complete Y-chromosome assemblies, we developed k-mer-based tools to estimate Yq12 size directly from short- and long-read sequencing data without requiring sample-specific complete assemblies. Across 1,597 males from the 1000 Genomes Project, Yq12 length ranged from ~1 to 52 Mb (mean 26.5 Mb) and varied substantially within and between Y haplogroups. Phylogenetic analyses supported long-term constraint of total Yq12 length near an equilibrium of 26 Mb and strongly coupled evolution of DYZ1 and DYZ2 around near-parity composition. In 303 father-son pairs, fathers had longer estimated Yq12 regions in 70.6% of pairs, while sons more often showed DYZ1:DYZ2 ratios closer to 1:1. Testis Iso-Seq identified transcripts containing DYZ1 and DYZ2 satellite sequences, while single-cell RNA-seq revealed their expression in haploid spermatids. These results establish a scalable framework for studying Yq12 population variation, intergenerational dynamics, and transcription, providing new opportunities to investigate how extreme satellite-array variation relates to Y-chromosome evolution and male germ-cell biology.