Functional profiling of long intergenic non-coding RNAs in fission yeast.
Functional profiling of long intergenic non-coding RNAs in fission yeast.
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DOI：
10.7554/elife.76000
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发表时间：
2022-01-05
期刊：
影响因子：
7.7
通讯作者：
Bähler J
中科院分区：
文献类型：
--
作者：
Rodriguez-Lopez M;Anver S;Cotobal C;Kamrad S;Malecki M;Correia-Melo C;Hoti M;Townsend S;Marguerat S;Pong SK;Wu MY;Montemayor L;Howell M;Ralser M;Bähler J
真核生物基因组表达许多不与任何编码基因重叠的长基因间非编码RNA（lincRNA）。一些lincRNA在基因调控的各个方面发挥作用，但一般不清楚lincRNA在多大程度上有助于从基因型到表型的信息流。为了探索这个问题，我们系统地分析了lincRNA在粟酒裂殖酵母中的细胞作用。使用基于CRISPR/Cas9的无缝基因组编辑，我们删除了141个lincRNA基因以广泛表型这些突变体，以及238个不同的编码基因突变体用于功能背景。我们应用高通量基于菌落的测定来确定突变体在良性条件下的生长和生存能力，以及对145种不同的营养、药物和应激条件的响应。这些分析揭示了47.5%的lincRNA和96%的蛋白质编码基因的表型。对于110个lincRNA突变体，我们还进行了高通量显微镜和流式细胞术测定，将这些lincRNA中的37%与细胞大小和/或细胞周期控制联系起来。结合所有测定，我们检测到84个（59.6%）测试的所有lincRNA缺失突变体的表型。对于互补功能推断，我们分析了在47种不同条件下异位过表达113个lincRNA基因的菌株的菌落生长。在这些过表达菌株中，102株（90.3%）在某些条件下显示出生长改变。聚类分析为一些lincRNA提供了进一步的功能线索和关系。这些丰富的表型组学数据集将lincRNA突变体与数百种表型相关联，表明所分析的大多数lincRNA在特定的环境或生理背景下发挥细胞功能。这项研究为进一步剖析这些lincRNA在相关条件下的作用提供了基础。
Eukaryotic genomes express numerous long intergenic non-coding RNAs (lincRNAs) that do not overlap any coding genes. Some lincRNAs function in various aspects of gene regulation, but it is not clear in general to what extent lincRNAs contribute to the information flow from genotype to phenotype. To explore this question, we systematically analysed cellular roles of lincRNAs in Schizosaccharomyces pombe. Using seamless CRISPR/Cas9-based genome editing, we deleted 141 lincRNA genes to broadly phenotype these mutants, together with 238 diverse coding-gene mutants for functional context. We applied high-throughput colony-based assays to determine mutant growth and viability in benign conditions and in response to 145 different nutrient, drug, and stress conditions. These analyses uncovered phenotypes for 47.5% of the lincRNAs and 96% of the protein-coding genes. For 110 lincRNA mutants, we also performed high-throughput microscopy and flow cytometry assays, linking 37% of these lincRNAs with cell-size and/or cell-cycle control. With all assays combined, we detected phenotypes for 84 (59.6%) of all lincRNA deletion mutants tested. For complementary functional inference, we analysed colony growth of strains ectopically overexpressing 113 lincRNA genes under 47 different conditions. Of these overexpression strains, 102 (90.3%) showed altered growth under certain conditions. Clustering analyses provided further functional clues and relationships for some of the lincRNAs. These rich phenomics datasets associate lincRNA mutants with hundreds of phenotypes, indicating that most of the lincRNAs analysed exert cellular functions in specific environmental or physiological contexts. This study provides groundwork to further dissect the roles of these lincRNAs in the relevant conditions.
影响因子：
14.9
作者：
Ard R;Allshire RC
通讯作者：
Allshire RC