Institute of Genetics and Biotechnology, Faculty of Biology, University of Warsaw, 02-106, Warsaw, Poland.
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, 02-106, Warsaw, Poland.
BMC Genomics. 2024 Sep 14;25(1):860. doi: 10.1186/s12864-024-10791-4.
Organellar transcriptomes are relatively under-studied systems, with data related to full-length transcripts and posttranscriptional modifications remaining sparse. Direct RNA sequencing presents the possibility of accessing a previously unavailable layer of information pertaining to transcriptomic data, as well as circumventing the biases introduced by second-generation RNA-seq platforms. Direct long-read ONT sequencing allows for the isoform analysis of full-length transcripts and the detection of posttranscriptional modifications. However, there are still relatively few projects employing this method specifically for studying organellar transcriptomes.
Candida albicans is a promising model for investigating nucleo-mitochondrial interactions. This work comprises ONT sequencing of the Candida albicans mitochondrial transcriptome along with the development of a dedicated data analysis pipeline. This approach allowed for the detection of complete transcript isoforms and posttranslational RNA modifications, as well as an analysis of C. albicans deletion mutants in genes coding for the 5' and 3' mitochondrial RNA exonucleases CaPET127 and CaDSS1. It also enabled for corrections to previous studies in terms of 3' and 5' transcript ends. A number of intermediate splicing isoforms was also discovered, along with mature and unspliced transcripts and changes in their abundances resulting from disruption of both 5' and 3' exonucleolytic processing. Multiple putative posttranscriptional modification sites have also been detected.
This preliminary work demonstrates the suitability of direct RNA sequencing for studying yeast mitochondrial transcriptomes in general and provides new insights into the workings of the C. albicans mitochondrial transcriptome in particular. It also provides a general roadmap for analyzing mitochondrial transcriptomic data from other organisms.
细胞器转录组是一个相对研究较少的系统，有关全长转录物和转录后修饰的数据仍然很少。直接 RNA 测序有可能获得以前无法获得的与转录组数据相关的信息层，并且可以避免第二代 RNA-seq 平台带来的偏差。直接的长读长 ONT 测序允许对全长转录物的异构体分析和转录后修饰的检测。然而，仍然相对较少的项目专门使用这种方法来研究细胞器转录组。
白色念珠菌是研究核-线粒体相互作用的有前途的模型。这项工作包括对白色念珠菌线粒体转录组的 ONT 测序以及专门数据分析管道的开发。这种方法允许检测完整的转录物异构体和翻译后 RNA 修饰，以及对编码线粒体 RNA 5' 和 3' 外切酶 CaPET127 和 CaDSS1 的基因的 C. albicans 缺失突变体进行分析。它还可以纠正以前关于 3' 和 5' 转录末端的研究。还发现了一些中间剪接异构体，以及成熟和未剪接的转录物，并且由于 5' 和 3' 外切酶加工的破坏，它们的丰度发生了变化。还检测到了多个假定的转录后修饰位点。
这项初步工作表明，直接 RNA 测序一般适用于研究酵母线粒体转录组，特别是为白色念珠菌线粒体转录组的工作提供了新的见解。它还为分析来自其他生物体的线粒体转录组数据提供了一般路线图。
