# Family C of Short Interspersed Elements in the Genomes of Lagomorphs: Structure, Evolution, Transcription and Transcript Polyadenylation

## TL;DR

This study explores the C SINE family in lagomorphs, revealing their structure, evolutionary history, and role in transcription and polyadenylation.

## Contribution

The study identifies and characterizes the C SINE family in lagomorphs, including their polyadenylation mechanism and evolutionary activity.

## Key findings

- C SINEs are present in over a million copies in lagomorph genomes and have been active for at least 60 million years.
- C1 subfamily retains functional AATAAA motifs and is involved in polyadenylation, while C2 is active in hares and rabbits but absent in pikas.
- Transcription of C SINEs is activated at the 16-cell stage of rabbit embryos.

## Abstract

Short Interspersed Elements (SINEs) are small pieces of DNA that can move around in the genetic material of living things. They can modulate the genome function, e.g., induce hereditary diseases in humans and animals. In mammals, some SINEs have specific signals that help them make more copies of themselves. Researchers found a type of these genetic mobile elements called C SINEs in rabbits, hares, and pikas. More than one million copies of C SINE have been detected in the genomes of each of the five studied species of the order Lagomorpha. This SINE first appeared at least 60 million years ago. We discovered that C SINEs have different versions with unique features. By studying these features, we could see how these SINEs have been active in the evolution of these mammals. In particular, they found certain sequences in C SINEs that are important for making more copies of themselves. The study showed that these elements become transcriptionally active at the 16-cell stage of rabbit embryos.

Short Interspersed Elements (SINEs) are eukaryotic non-autonomous retrotransposons that rely on RNA polymerase III (pol III) for transcription. A subset of mammalian SINEs—designated T+ SINEs—harbors a canonical polyadenylation signal (AATAAA), a pol III terminator, and an A-rich tail at their 3′ end, thereby acquiring the unusual ability to undergo AAUAAA-dependent polyadenylation. Here, we delineate the genomic architecture, evolutionary history, and polyadenylation behavior of the C SINE family in Lagomorpha. Comprehensive bioinformatics searches identified 1.2–1.6 million C copies distributed across Leporidae (hares and rabbits) and Ochotonidae (pikas) genomes. Phylogenetic reconstruction resolved two diverged leporid subfamilies, C1 and C2, with C1 predating C2 and comprising five-fold more copies. Only C1 qualifies as a T+ SINE, retaining functional or rudimentary AATAAA motifs and pol III terminators. In contrast, C2 is absent from pika genomes, yet remains retrotranspositionally competent in hares and rabbits. Lineage-specific analyses further reveal episodic activity of certain C1 variants throughout the last 10 million years of pika evolution. Functional assays in transfected HeLa cells demonstrate that AATAAA and an upstream polypyrimidine tract constitute the minimal cis-determinant for efficient C1 transcript polyadenylation. Finally, transcriptome profiling of pre-implantation rabbit embryos indicates that pol III-driven SINE C transcription is activated at the 16-cell stage.