Pre‐mRNA splicing is a highly conserved eukaryotic process, but our understanding of it is limited by a historical focus on well‐studied organisms such as humans and yeast. There is considerable diversity in mechanisms and components of pre‐mRNA splicing, especially in lineages that have evolved under the pressures of genome reduction. The ancestor of red algae is thought to have undergone genome reduction prior to the lineage's radiation, resulting in overall gene and intron loss in extant groups. Previous studies on the extremophilic red alga Cyanidioschyzon merolae revealed an intron‐sparse genome with a highly reduced spliceosome. To determine whether these features applied to other red algae, we investigated multiple aspects of pre‐mRNA splicing in the mesophilic red alga Porphyridium purpureum. Through strand‐specific RNA‐Seq, we observed high levels of intron retention across a large number of its introns, and nearly half of the transcripts for these genes are not spliced at all. We also discovered a relationship between variability of 5′ splice site sequences and levels of splicing. To further investigate the connections between intron retention and splicing machinery, we bioinformatically assembled the P. purpureum spliceosome, and biochemically verified the presence of snRNAs. While most other core spliceosomal components are present, our results suggest highly divergent or missing U1 snRNP proteins, despite the presence of an uncharacteristically long U1 snRNA. These unusual aspects highlight the diverse nature of pre‐mRNA splicing that can be seen in lesser‐studied eukaryotes, raising the importance of investigating fundamental eukaryotic processes outside of model organisms.

中文翻译：

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Porphyridium purpureum 中前 mRNA 剪接和剪接体机制的表征以及红藻的进化意义

Pre-mRNA 剪接是一个高度保守的真核过程，但我们对它的理解受限于历史上对人类和酵母等经过充分研究的生物体的关注。mRNA 前体剪接的机制和成分存在相当大的多样性，特别是在基因组减少压力下进化的谱系中。红藻的祖先被认为在谱系辐射之前经历了基因组减少，导致现存群体的整体基因和内含子丢失。先前对嗜极红藻Cyanidioschyzon merolae 的研究揭示了具有高度减少的剪接体的内含子稀疏基因组。为了确定这些特征是否适用于其他红藻，我们研究了嗜温红藻中前 mRNA 剪接的多个方面紫菜 紫菜。通过链特异性 RNA-Seq，我们观察到其大量内含子的高水平内含子保留，并且这些基因的近一半转录本根本没有剪接。我们还发现了 5' 剪接位点序列的可变性与剪接水平之间的关系。为了进一步研究内含子保留和剪接机制之间的联系，我们通过生物信息学组装了P。 紫癜剪接体，并通过生化验证了 snRNA 的存在。虽然存在大多数其他核心剪接体成分，但我们的结果表明存在高度不同或缺失的 U1 snRNP 蛋白，尽管存在异常长的 U1 snRNA。这些不寻常的方面突出了前 mRNA 剪接的多样性，这可以在研究较少的真核生物中看到，从而提高了研究模式生物之外的基本真核过程的重要性。