Table_3_Exploring the Relationship Among Divergence Time and Coding and Non-coding Elements in the Shaping of Fungal Mitochondrial Genomes.docx

The order Hypocreales (Ascomycota) is composed of ubiquitous and ecologically diverse fungi such as saprobes, biotrophs, and pathogens. Despite their phylogenetic relationship, these species exhibit high variability in biomolecules production, lifestyle, and fitness. The mitochondria play an important role in the fungal biology, providing energy to the cells and regulating diverse processes, such as immune response. In spite of its importance, the mechanisms that shape fungal mitogenomes are still poorly understood. Herein, we investigated the variability and evolution of mitogenomes and its relationship with the divergence time using the order Hypocreales as a study model. We sequenced and annotated for the first time Trichoderma harzianum mitochondrial genome (mtDNA), which was compared to other 34 mtDNAs species that were publicly available. Comparative analysis revealed a substantial structural and size variation on non-coding mtDNA regions, despite the conservation of copy number, length, and structure of protein-coding elements. Interestingly, we observed a highly significant correlation between mitogenome length, and the number and size of non-coding sequences in mitochondrial genome. Among the non-coding elements, group I and II introns and homing endonucleases genes (HEGs) were the main contributors to discrepancies in mitogenomes structure and length. Several intronic sequences displayed sequence similarity among species, and some of them are conserved even at gene position, and were present in the majority of mitogenomes, indicating its origin in a common ancestor. On the other hand, we also identified species-specific introns that advocate for the origin by different mechanisms. Investigation of mitochondrial gene transfer to the nuclear genome revealed that nuclear copies of the nad5 are the most frequent while atp8, atp9, and cox3 could not be identified in any of the nuclear genomes analyzed. Moreover, we also estimated the divergence time of each species and investigated its relationship with coding and non-coding elements as well as with the length of mitogenomes. Altogether, our results demonstrated that introns and HEGs are key elements on mitogenome shaping and its presence on fast-evolving mtDNAs could be mostly explained by its divergence time, although the intron sharing profile suggests the involvement of other mechanisms on the mitochondrial genome evolution, such as horizontal transference.

Hypocreales（子囊菌亚门）这一类群由广泛分布且生态多样性丰富的真菌组成，包括腐生菌、生物寄生菌和病原菌。尽管这些物种在系统发育上存在关联，但它们在生物分子生产、生活方式和适应度方面表现出显著的多样性。线粒体在真菌生物学中发挥着至关重要的作用，为细胞提供能量并调节免疫反应等多样化的生命过程。尽管其重要性不言而喻，但塑造真菌线粒体基因组（mitogenome）的机制仍处于认识不足的状态。在本研究中，我们以Hypocreales类群为研究对象，探究了线粒体基因组的变异性和演化，并分析了其与物种分化时间的关系。我们首次对Trichoderma harzianum的线粒体基因组（mtDNA）进行了测序和注释，并将其与公开发表的34种mtDNA物种进行比较。比较分析显示，尽管蛋白编码区段的复制数、长度和结构得到保守，但在非编码的mtDNA区域却存在着显著的结构性及大小差异。有趣的是，我们观察到线粒体基因组长度与线粒体基因组中非编码序列的数量和大小之间存在高度显著的相关性。在非编码元件中，I组和II组内含子以及同源内切酶基因（HEGs）是造成线粒体基因组结构和长度差异的主要因素。一些内含子序列在物种间显示出序列相似性，其中一些甚至在基因位置上保守，并且存在于大多数线粒体基因组中，这表明其起源于共同祖先。另一方面，我们也发现了物种特异性的内含子，这表明它们可能通过不同的机制起源。对线粒体基因向核基因组转移的研究表明，nad5的核拷贝是最常见的，而atp8、atp9和cox3在所分析的任何核基因组中均未发现。此外，我们还估计了每个物种的分化时间，并探讨了其与编码和非编码元件以及线粒体基因组长度的关系。总之，我们的研究结果揭示了内含子和HEGs是塑造线粒体基因组的关键要素，其在快速演化的mtDNAs中的存在大多可由其分化时间解释，尽管内含子共享模式暗示了其他机制在线粒体基因组演化中的参与，例如水平转移。