A long-read RNA sequencing and polysome profiling framework reveals transposable element–driven transcript diversity and translational rewiring in glioblastoma [LR]

Transposable elements (TEs) account for over half of the human genome and are often derepressed in cancer. TEs can add cryptic splice sites, undergo exonization, and generate gene–TE fusion transcripts, but the combined effects of TEs on RNA processing and translation in glioblastoma stem cells (GSCs) remains incompletely elucidated. We combined long-read RNA sequencing with polysome profiling in four patient-derived GSCs and two neural stem cell (NSC) controls to resolve TE-associated transcript diversity and its relationship to ribosomal engagement. Across GSCs, we identified 13,421 alternative splicing (AS) events, 3,077 of which contained TEs within 150 bp of splice junctions. AS sites proximal to TEs were associated with increased isoform switching compared to non–TE-associated AS sites (odds ratio 2.9 - 4.3). Moreover, AS isoforms generated from TE-proximal sites were more likely to exhibit altered ribosomal association (odds ratio 2.54). Directional shifts were observed, with shorter isoforms associating with monosome fractions and longer isoforms with polysome fractions. To enable systematic detection of gene - TE chimeric transcripts, we developed FuTER (Fusion TE Reporter), a long-read–based framework for identifying TE-associated fusions. Application to GSC datasets identified 78 GSC enriched fusion transcripts, several supported by breakpoint-spanning reads in polysome fractions, consistent with ribosome association. Our data suggest that TEs correlate with abnormal splicing activity and altered ribosome engagement in glioblastoma stem cells. By integrating long-read sequencing with polysome profiling and fusion detection, we establish a framework for analysis of TE-induced transcript diversity and its effects on cancer evolution and plasticity.