1. Identification of epigenetically regulated genes involved in plant-virus interaction and their role in virus-triggered induced resistanceRégis L. Corrêa, Denis Kutnjak, Silvia Ambrós, Mónica Bustos, Santiago F. Elena, 2024, original scientific article Abstract: Background: Plant responses to a wide range of stresses are known to be regulated by epigenetic mechanisms. Path-ogen-related investigations, particularly against RNA viruses, are however scarce. It has been demonstrated that Arabi-dopsis thaliana plants defective in some members of the RNA-directed DNA methylation (RdDM) or histone modi-fication pathways presented differential susceptibility to the turnip mosaic virus. In order to identify genes directly targeted by the RdDM-related RNA Polymerase V (POLV ) complex and the histone demethylase protein JUMONJI14 (JMJ14) during infection, the transcriptomes of infected mutant and control plants were obtained and integrated with available chromatin occupancy data for various epigenetic proteins and marks. Results: A comprehensive list of virus-responsive gene candidates to be regulated by the two proteins was obtained. Twelve genes were selected for further characterization, confirming their dynamic regulation during the course of infection. Several epigenetic marks on their promoter sequences were found using in silico data, raising confidence that the identified genes are actually regulated by epigenetic mechanisms. The altered expression of six of these genes in mutants of the methyltransferase gene CURLY LEAF and the histone deacetylase gene HISTONE DEACETYLASE 19 suggests that some virus-responsive genes may be regulated by multiple coordinated epigenetic complexes. A temporally separated multiple plant virus infection experiment in which plants were transiently infected with one virus and then infected by a second one was designed to investigate the possible roles of the identified POLV- and JMJ14-regulated genes in wild-type (WT ) plants. Plants that had previously been stimulated with viruses were found to be more resistant to subsequent virus challenge than control plants. Several POLV- and JMJ14-regulated genes were found to be regulated in virus induced resistance in WT plants, with some of them poisoned to be expressed in early infection stages. Conclusions: A set of confident candidate genes directly regulated by the POLV and JMJ14 proteins during virus infection was identified, with indications that some of them may be regulated by multiple epigenetic modules. A sub-set of these genes may also play a role in the tolerance of WT plants to repeated, intermittent virus infections.Keywords Biotic stress, Defense priming, Epigenetics, Histone modifications, Induced resistance, Potyvirus, RNA-directed DNA methylation.
Keywords: biotic stress, defense priming, epigenetics, histone modifications, induced resistance, Potyvirus, RNA-directed DNA methylation
Published in DiRROS: 17.05.2024; Views: 20; Downloads: 7
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2. New approach for detection of normal alternative splicing events and aberrant spliceogenic transcripts with long-range PCR and deep RNA sequencingVita Šetrajčič Dragoš, Vida Stegel, Ana Blatnik, Gašper Klančar, Mateja Krajc, Srdjan Novaković, 2021, original scientific article Abstract: RNA sequencing is a promising technique for detecting normal and aberrant RNA isoforms. Here, we present a new single-gene, straightforward 1-day hands-on protocol for detection of splicing alterations with deep RNA sequencing from blood. We have validated our method%s accuracy by detecting previously published normal splicing isoforms of STK11 gene. Additionally, the same technique was used to provide the first comprehensive catalogue of naturally occurring alternative splicing events of the NBN gene in blood. Furthermore, we demonstrate that our approach can be used for detection of splicing impairment caused by genetic variants. Therefore, we were able to reclassify three variants of uncertain significance: NBN:c.584G>A, STK11:c.863-5_863-3delCTC and STK11:c.615G>A. Due to the simplicity of our approach, it can be incorporated into any molecular diagnostics laboratory for determination of variant%s impact on splicing.
Keywords: RNA sequencing, DNA variant, splicing
Published in DiRROS: 21.09.2022; Views: 473; Downloads: 260
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3. Identification of spliceogenic variants beyond canonical GT-AG splice sites in hereditary cancer genesVita Šetrajčič Dragoš, Ksenija Strojnik, Gašper Klančar, Petra Škerl, Vida Stegel, Ana Blatnik, Marta Banjac, Mateja Krajc, Srdjan Novaković, 2022, original scientific article Abstract: Pathogenic/likely pathogenic variants in susceptibility genes that interrupt RNA splicing are a well-documented mechanism of hereditary cancer syndromes development. However, if RNA studies are not performed, most of the variants beyond the canonical GT-AG splice site are characterized as variants of uncertain significance (VUS). To decrease the VUS burden, we have bioinformatically evaluated all novel VUS detected in 732 consecutive patients tested in the routine genetic counseling process. Twelve VUS that were predicted to cause splicing defects were selected for mRNA analysis. Here, we report a functional characterization of 12 variants located beyond the first two intronic nucleotides using RNAseq in APC, ATM, FH, LZTR1, MSH6, PALB2, RAD51C, and TP53 genes. Based on the analysis of mRNA, we have successfully reclassified 50% of investigated variants. 25% of variants were downgraded to likely benign, whereas 25% were upgraded to likely pathogenic leading to improved clinical management of the patient and the family members.
Keywords: hereditary cancer, RNA sequencing, spliceogenic
Published in DiRROS: 07.09.2022; Views: 500; Downloads: 258
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5. RNA-dependent cysteine biosynthesis in bacteria and archaeaTakahito Mukai, Ana Crnković, Takuya Umehara, Natalia N. Ivanova, Nikos C. Kyrpides, Dieter Söll, 2017, original scientific article Keywords: amino acyl-tRNA synthetases, archaea, archaeal proteins, bacteria, computational biology, crystallography, X-ray, genetic code, genome, archaeal, genome, bacterial, phosphoserine, protein binding, protein biosynthesis, RNA, archaeal, RNA, bacterial, RNA, transfer, amino acyl, RNA, transfer, cys, sulfur/metabolism, biochemistry, bioinformatics, cysteine biosynthesis, genetic code, translation
Published in DiRROS: 07.05.2020; Views: 1740; Downloads: 1322
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