Long terminal repeat (LTR) -retrotransposons constitute a significant part of eukaryotic genomes and influence their function and evolution . Like other RNA viruses, LTR-retrotransposons efficiently utilize their RNA genome to interact with host cell machinery during replication . Here, we provide the first genome-wide RNA secondary structure model for a LTR-retrotransposon in living cells . Using SHAPE probing, we explore the secondary structure of the yeast Ty1 retrotransposon RNA genome in its native in vivo state and under defined in vitro conditions . Comparative analyses reveal the strong impact of the cellular environment on folding of Ty1 RNA . In vivo, Ty1 genome RNA is significantly less structured and more dynamic but retains specific well-structured regions harboring functional cis-acting sequences . Ribosomes participate in the unfolding and remodeling of Ty1 RNA, and inhibition of translation initiation stabilizes Ty1 RNA structure . Together, our findings support the dual role of Ty1 genomic RNA as a template for protein synthesis and reverse transcription . This study also contributes to understanding how a complex multifunctional RNA genome folds in vivo, and strengthens the need for studying RNA structure in its natural cellular context.