Correction: Functional profiling of long intergenic non-coding RNAs in fission yeast

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 codinggene 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

XAP5 CIRCADIAN TIME KEEPER (XCT): A Conserved Chromatin Regulator that Affects Growth, Development and Stress Response

Circadian systems are widespread in nature and enable the organisms that posses these endogenous systems to anticipate environmental changes and therefore allow them to take full advantage of their phenotypic/developmental plasticity. Since current plant clock models are not complete enough to explain the roles of the circadian clock, novel clock components need to be identified. XAP5 CIRCADIAN TIME KEEPER (XCT) is potentially one such protein. I show that XCT is required for a fully functional clock, proper growth and development, and stress responses especially under temperature stress. XCT is well conserved in eukaryotes and I demonstrate that it is functionally conserved between Arabidopsis thaliana and Schizosaccharomyces pombe at the physiological and molecular levels. I therefore functionally characterized the XCT ortholog Xap5 in fission yeast and demonstrate that it is a novel chromatin regulatory protein functioning with multiple known chromatin regulators including the H2A variant H2A.Z. Chromatin regulation plays an important role in regulating the expression of stimulus-responsive genes to which category many developmental, circadian and temperatureresponsive genes belong. Because of XCT’s potential involvement in the circadian clock, chromatin modifications and plant growth-development-stress responses, I conclude that XCT is a conserved novel protein which connects the worlds of clock and chromatin modifications both to each other and to growth, development and stress response pathways

XAP5 CIRCADIAN TIMEKEEPER regulates RNA splicing and the circadian clock by genetically separable pathways

The circadian oscillator allows organisms to synchronize their cellular and physiological activities with diurnal environmental changes. In plants, the circadian clock is primarily composed of multiple transcriptional-translational feedback loops. Regulators of post-transcriptional events, such as pre-mRNA splicing factors, are also involved in controlling the pace of the clock. However, in most cases the underlying mechanisms remain unclear. We have previously identified XAP5 CIRCADIAN TIMEKEEPER (XCT) as an Arabidopsis thaliana circadian clock regulator with uncharacterized molecular functions. Here, we report that XCT physically interacts with components of the spliceosome, including members of the Nineteen Complex (NTC). PacBio Iso-Seq data show that xct mutants have transcriptome-wide pre-mRNA splicing defects, predominantly aberrant 3' splice site selection. Expression of a genomic copy of XCT fully rescues those splicing defects, demonstrating that functional XCT is important for splicing. Dawn-expressed genes are significantly enriched among those aberrantly spliced in xct mutants, suggesting that the splicing activity of XCT may be circadian regulated. Furthermore, we show that loss-of-function mutations in PRP19A or PRP19B, two homologous core NTC components, suppress the short circadian period phenotype of xct-2. However, we do not see rescue of the splicing defects of core clock genes in prp19 xct mutants. Therefore, our results suggest that XCT may regulate splicing and the clock function through genetically separable pathways

XAP5 CIRCADIAN TIMEKEEPER regulates RNA splicing and the circadian clock by genetically separable pathways.

The circadian oscillator allows organisms to synchronize their cellular and physiological activities with diurnal environmental changes. In plants, the circadian clock is primarily composed of multiple transcriptional-translational feedback loops. Regulators of post-transcriptional events, such as precursor messenger RNAs (pre-mRNA) splicing factors, are also involved in controlling the pace of the clock. However, in most cases the underlying mechanisms remain unclear. We have previously identified XAP5 CIRCADIAN TIMEKEEPER (XCT) as an Arabidopsis thaliana circadian clock regulator with uncharacterized molecular functions. Here, we report that XCT physically interacts with components of the spliceosome, including members of the Nineteen Complex (NTC). PacBio Iso-Seq data show that xct mutants have transcriptome-wide pre-mRNA splicing defects, predominantly aberrant 3' splice site selection. Expression of a genomic copy of XCT fully rescues those splicing defects, demonstrating that functional XCT is important for splicing. Dawn-expressed genes are significantly enriched among those aberrantly spliced in xct mutants, suggesting that the splicing activity of XCT may be circadian regulated. Furthermore, we show that loss-of-function mutations in PRP19A or PRP19B, 2 homologous core NTC components, suppress the short circadian period phenotype of xct-2. However, we do not see rescue of the splicing defects of core clock genes in prp19 xct mutants. Therefore, our results suggest that XCT may regulate splicing and the clock function through genetically separable pathways

Yeast X‐chromosome‐associated protein 5 (Xap5) functions with H2A.Z to suppress aberrant transcripts

Chromatin regulatory proteins affect diverse developmental and environmental response pathways via their influence on nuclear processes such as the regulation of gene expression. Through a genome-wide genetic screen, we implicate a novel protein called X-chromosome-associated protein 5 (Xap5) in chromatin regulation. We show that Xap5 is a chromatin-associated protein acting in a similar manner as the histone variant H2A.Z to suppress expression of antisense and repeat element transcripts throughout the fission yeast genome. Xap5 is highly conserved across eukaryotes, and a plant homolog rescues xap5 mutant yeast. We propose that Xap5 likely functions as a chromatin regulator in diverse organisms

An incoherent feed‐forward loop mediates robustness and tunability in a plant immune network

Immune signaling networks must be tunable to alleviate fitness costs associated with immunity and, at the same time, robust against pathogen interferences. How these properties mechanistically emerge in plant immune signaling networks is poorly understood. Here, we discovered a molecular mechanism by which the model plant species Arabidopsis thaliana achieves robust and tunable immunity triggered by the microbe-associated molecular pattern, flg22. Salicylic acid (SA) is a major plant immune signal molecule. Another signal molecule jasmonate (JA) induced expression of a gene essential for SA accumulation, EDS5 Paradoxically, JA inhibited expression of PAD4, a positive regulator of EDS5 expression. This incoherent type-4 feed-forward loop (I4-FFL) enabled JA to mitigate SA accumulation in the intact network but to support it under perturbation of PAD4, thereby minimizing the negative impact of SA on fitness as well as conferring robust SA-mediated immunity. We also present evidence for evolutionary conservation of these gene regulations in the family Brassicaceae Our results highlight an I4-FFL that simultaneously provides the immune network with robustness and tunability in A. thaliana and possibly in its relatives