Novel insights into retrogene evolution and cold adaptation in the alpine plant Crucihimalaya himalaica (Brassicaceae)

BACKGROUND: Retrogenes, which formed via retrotransposon-mediated processes, are important drivers of genetic innovation and adaptive evolution. Although most plant genomes harbor abundant LTR retrotransposons that facilitate retrogene formation, studies on retrogenes in wild plants remain limited. Crucihimalaya himalaica, a close relative of Arabidopsis thaliana endemic to the Qinghai-Tibet Plateau (QTP) region, provides a valuable model for investigating plant adaptation to high-altitude environments. RESULTS: We report an updated high-quality, chromosome-level genome assembly of C. himalaica (scaffold N50: 30.99 Mb) and transcriptomic data under cold stress. We identified 240 retrogenes (90% with complex structures), predominantly located in gene-rich regions, and found that LTR retrotransposons are more abundant in the upstream regions of C. himalaica retrogenes than in those of Arabidopsis thaliana. Under cold stress, retrogenes were significantly more likely to be differentially expressed compared to background genes, with 89.6% of them displaying expression patterns distinct from their parental genes. These differentially expressed retrogenes were evolutionarily older, under stronger purifying selection, and enriched in carbohydrate metabolism, transmembrane transport, and calcium-mediated signaling. Functional assays in A. thaliana demonstrated that mutants of C. himalaica retrogenes homologs exhibited enhanced freezing tolerance, likely via cell wall remodeling. The recently formed retrogene XMJChr05g02504 exhibited structural divergence, cold-induced expression, and signatures of positive selection, suggesting functional innovation of retrogene potentially contributing to C. himalaica's adaptation to the cold environment of QTP region. CONCLUSIONS: Our study provides a high-quality genome assembly of C. himalaica and reveals the evolutionary dynamics and roles of retrogenes, offering new insights into the genetic mechanisms underlying plant adaptation to high-altitude environments.