Joint Segmental Duplication Co-option Drives Human-specific Transcriptional Readthrough and Expression Fine-tuning of NPEPPS - TBC1D3

Abstract Segmental duplication (SD) is a major driver of functional changes in evolution and disease. Many genes embedded within SDs, such as NPEPPS and TBC1D3 , display substantial copy number variation (CNV) across individuals. Yet, the precise identification of the functional copies and their transcriptional outputs remains largely unstudied. Focusing on NPEPPS and TBC1D3 , we illustrate human-specific expression fine-tuning mechanisms associated with readthrough transcripts. We identified a human-specific NPEPPS - TBC1D3 digenic genomic structure that originated from a joint SD pair and became fixed across populations. Experiments demonstrate that this structure generates NPEPPS - TBC1D3 readthrough transcripts, which are the predominant isoforms of TBC1D3 expression in various cell types, fine-tuning its protein level. Furthermore, a human-specific hypomethylation signal within an upstream CpG island of NPEPPS precisely pinpoints the expressed TBC1D3 paralog. Moreover, we reveal transcriptional readthrough events are ∼3-fold enriched for joint-SD-associated transcriptional readthrough (JSDTR) and identify 109 JSDTR gene pairs, including neurodevelopmentally important pairs and clinically interesting SERF1A/B-SMN1/2 . Taken together, our findings comprehensively describe an example of how a joint SD event shaped evolution and suggest that JSDTR is a broad mechanism for the emergence of new functions.