Table_5_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（子囊菌门）这一类群包含着广泛分布且生态多样性丰富的真菌，诸如腐生菌、生物寄主和病原菌。尽管这些物种在系统发育上存在关联，但在生物大分子合成、生活方式和适应度等方面却展现出显著差异。线粒体在真菌生物学中扮演着至关重要的角色，为细胞提供能量并调节多种生理过程，例如免疫反应。尽管其重要性不言而喻，但塑造真菌线粒体基因组的机制仍处于模糊不清的状态。在本研究中，我们以Hypocreales类群为研究对象，探讨了线粒体基因组（mitogenome）的变异性和进化，以及其与物种分化时间的关联。我们首次对Trichoderma harzianum的线粒体基因组（mtDNA）进行了测序和注释，并将其与其他34种公开可用的mtDNAs物种进行了比较。比较分析显示，尽管蛋白编码元件的拷贝数、长度和结构得以保持，但在非编码的mtDNA区域中仍存在显著的构造和大小变异。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关联。在非编码元件中，I组和II组内含子和同源内切酶基因（HEGs）是导致线粒体基因组结构和长度差异的主要贡献者。一些内含子序列在不同物种间表现出序列相似性，其中一些甚至在基因位置上保持保守，存在于大多数线粒体基因组中，这表明它们起源于共同的祖先。另一方面，我们也发现了具有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的转移进行的研究揭示了nad5基因在核基因组中的拷贝最为常见，而atp8、atp9和cox3基因在任何分析的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。总体而言，我们的研究结果揭示了内含子和HEGs是线粒体基因组塑造的关键要素，而其在快速进化的mtDNAs中的存在，大多可以由其分化时间来解释，尽管内含子共享模式暗示了其他机制，如水平转移，在线粒体基因组进化中的参与。