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Human-specific segmental duplications (HSDs) contain millions of base pairs of sequence unique to the human genome, including genes that shape neurodevelopment. Despite their young age (<6 million years), HSD genes exhibit widespread regulatory divergence, with paralog-specific expression patterns documented across a variety of tissues and cell types. Using long-read expression and epigenomic data, we show that human-specific paralogs tend to have lower activity than the shared, ancestral ones. To systematically characterize the cis-regulatory elements (CREs) within HSDs and understand patterns of regulatory change in recently evolved gene families, we conducted a massively parallel reporter assay of 7,160 human duplicated and chimpanzee orthologous sequences in lymphoblastoid (GM12878) and neuroblastoma (SH-SY5Y) cell lines. A large proportion (14–24%) of sequences exhibited differential activity relative to the chimpanzee ortholog (or between human paralogs), mostly with small fold-differences. Combining measured activity levels across all assayed sequences, predicted differences in cis-regulatory activity correlated with mRNA levels in SH-SY5Y. Differentially active CREs were validated for CHRFAM7A, HYDIN2, and SRGAP2C that may contribute to paralog-specific expression patterns and thereby to human-specific traits. While we find some changes in CRE activity shared between duplicate paralogs likely driving regulatory divergence in gene expression, consideration of non-shared adjacent sequences to duplications suggests a larger role for altered genome positional effects. In all, this work suggests that functional divergence of duplicated CREs contributes moderately to regulatory divergence of HSD genes and uncovers enhancers that are candidate drivers of human-specific regulatory patterns.
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This content will become publicly available on April 4, 2026
Quantifying the influence of genetic context on duplicated mammalian genes
Abstract Gene duplication is a fundamental part of evolutionary innovation. While single-gene duplications frequently exhibit asymmetric evolutionary rates between paralogs, the extent to which this applies to multi-gene duplications remains unclear. In this study, we investigate the role of genetic context in shaping evolutionary divergence within multi-gene duplications, leveraging microsynteny to differentiate source and target copies. Using a dataset of 193 mammalian genome assemblies and a bird outgroup, we systematically analyze patterns of sequence divergence between duplicated genes and reference orthologs. We find that target copies, those relocated to new genomic environments, exhibit elevated evolutionary rates compared to source copies in the ancestral location. This asymmetry is influenced by the distance between copies and the size of the target copy. We also demonstrate that the polarization of rate asymmetry in paralogs, the “choice” of the slowly evolving copy, is biased towards collective, block-wise polarization in multi-gene duplications. Our findings highlight the importance of genetic context in modulating post-duplication divergence, where differences in cis-regulatory elements and co-expressed gene clusters between source and target copies may be responsible. This study presents a large-scale test of asymmetric evolution in multi-gene duplications, offering new insight into how genome architecture shapes functional diversification of paralogs. Significance statementAfter a gene is duplicated, reduced selective constraints can lead the two copies to rapidly diverge, with one copy often evolving faster and occasionally gaining a new function. We quantify the influence of genetic context in choosing which copy of a duplicated gene has an elevated substitution rate. In a representative dataset of 193 mammalian genomes, we found strong evidence that gene copies pasted into new genomic locations tend to evolve faster than the corresponding copies in ancestral locations, suggesting an important role for the regulatory environment. The asymmetry in evolutionary rates of duplicated genes persists even for very large multigenic duplications, up to the scale of megabases, indicating that regulatory interactions frequently reach farther than previously thought.
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- Award ID(s):
- 2019745
- PAR ID:
- 10595434
- Publisher / Repository:
- bioRxiv
- Date Published:
- Format(s):
- Medium: X
- Institution:
- bioRxiv
- Sponsoring Org:
- National Science Foundation
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