Paramecium bursaria 內含子：內共生期間的剪接調控與纖毛蟲物種中短內含子的演化

Abstract

在內共生過程中，共生體整合進宿主細胞會顯著改變基因表現與細胞身分。雖然選擇性剪接已知可透過快速調節基因表現與蛋白質同功異構體的多樣性來促進細胞適應，但其在內共生期間扮演的調控角色仍所知甚少。Paramecium bursaria 是研究此現象的理想模式生物，因其細胞質中攜帶數百個 Chlorella variabilis 藻類共生體。此外，P. bursaria 及其相關的 Paramecium 物種具有極短的內含子（中位數約為 24 個核苷酸），因此為探討短內含子動態剪接、內共生期間的調控模式，以及纖毛蟲內含子演化提供了寶貴的系統。在本研究中，我們對含共生體（綠色）與無共生體（白色）的 P. bursaria 細胞進行時間進程RNA 定序，以分析內共生期間的內含子剪接模式。一般而言，5’端近端內含子的剪接，有助於提高基因表現量。我們發現綠色與白色的 P. bursaria細胞之間存在差異性剪接的內含子，且在跨膜轉運蛋白基因中有顯著富集，這些基因對於內共生期間宿主與藻類共生體間的養分交換至關重要。此外，我們鑑定出一系列剪接增強子與抑制子，位於保守的剪接體基因中，其表現量與內共生期間的差異性剪接內含子密切相關。透過跨纖毛蟲的內含子直系同源性分析，我們發現保留的內含子具有較高的剪接效率、較低的 GC 含量以及一致的內含子長度，顯示新演化出的內含子會經過剪接與表現上的優化。我們的研究結果提供了選擇性剪接在宿主於內共生過程中適應角色的新見解，並解釋了短內含子在真核基因體中的演化動態。

The integration of symbionts into host cells during endosymbiosis significantly alters gene expression and cell identity. While alternative splicing facilitates cellular adaptation through rapid modulation of gene expression and protein isoform diversity, its regulatory role during endosymbiosis remains poorly understood. Paramecium bursaria represents an ideal model for investigating this, as it harbors hundreds of Chlorella variabilis algae within its cytoplasm. Furthermore, P. bursaria and related Paramecium species possess exceptionally short introns (median ~24 nucleotides), providing a valuable system for examining short intron splicing dynamics, regulatory patterns during endosymbiosis, and ciliate intron evolution. In this study, we conducted time-course RNA sequencing on symbiont-bearing (green) and symbiont-free (white) P. bursaria cells to analyze intron splicing patterns during endosymbiosis. While in general, splicing, especially 5’ proximal introns, enhances gene expression, we identified differentially spliced introns between green and white, with an enrichment in transmembrane transporter genes, which are crucial for establishing nutrient transport between the host cell and its algal symbionts during endosymbiosis. Additionally, we identified splicing enhancers and repressors among conserved spliceosome genes, whose expression was closely linked to differentially spliced introns during endosymbiosis. Through intron orthology analysis across ciliates, we found that conserved introns are spliced more efficiently, with lower GC content and uniform length, suggesting that newly evolved introns undergo refinement to optimize splicing and expression. Our findings provide insight into the role of alternative splicing in host adaptation during endosymbiosis while shedding light on the evolutionary dynamics of short introns in eukaryotic genomes.