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Multi-OMICs landscape of SARS-CoV-2-induced host responses in human lung epithelial cells.
Pinto, SM, Subbannayya, Y, Kim, H, Hagen, L, Górna, MW, Nieminen, AI, Bjørås, M, Espevik, T, Kainov, D, Kandasamy, RK
iScience. 2023;(1):105895
Abstract
COVID-19 pandemic continues to remain a global health concern owing to the emergence of newer variants. Several multi-Omics studies have produced extensive evidence on host-pathogen interactions and potential therapeutic targets. Nonetheless, an increased understanding of host signaling networks regulated by post-translational modifications and their ensuing effect on the cellular dynamics is critical to expanding the current knowledge on SARS-CoV-2 infections. Through an unbiased transcriptomics, proteomics, acetylomics, phosphoproteomics, and exometabolome analysis of a lung-derived human cell line, we show that SARS-CoV-2 Norway/Trondheim-S15 strain induces time-dependent alterations in the induction of type I IFN response, activation of DNA damage response, dysregulated Hippo signaling, among others. We identified interplay of phosphorylation and acetylation dynamics on host proteins and its effect on the altered release of metabolites, especially organic acids and ketone bodies. Together, our findings serve as a resource of potential targets that can aid in designing novel host-directed therapeutic strategies.
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2.
Insight into ALKBH8-related intellectual developmental disability based on the first pathogenic missense variant.
Maddirevula, S, Alameer, S, Ewida, N, de Sousa, MML, Bjørås, M, Vågbø, CB, Alkuraya, FS
Human genetics. 2022;(2):209-215
Abstract
ALKBH8 is a methyltransferase that modifies tRNAs by methylating the anticodon wobble uridine residue. The syndrome of ALKBH8-related intellectual developmental disability (MRT71) has thus far been reported solely in the context of homozygous truncating variants that cluster in the last exon. This raises interesting questions about the disease mechanism, because these variants are predicted to escape nonsense mediated decay and yet they appear to be loss of function. Furthermore, the limited class of reported variants complicates the future interpretation of missense variants in ALKBH8. Here, we report a consanguineous family in which two children with MRT71-compatible phenotype are homozygous for a novel missense variant in the methyltransferase domain. We confirm the pathogenicity of this variant by demonstrating complete absence of ALKBH8-dependent modifications in patient cells. Targeted proteomics analysis of ALKBH8 indicates that the variant does not lead to loss of ALKBH8 protein expression. This report adds to the clinical delineation of MRT71, confirms loss of function of ALKBH8 as the disease mechanism and expands the repertoire of its molecular lesions.
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3.
Sculpting of DNA at abasic sites by DNA glycosylase homolog mag2.
Dalhus, B, Nilsen, L, Korvald, H, Huffman, J, Forstrøm, RJ, McMurray, CT, Alseth, I, Tainer, JA, Bjørås, M
Structure (London, England : 1993). 2013;(1):154-166
Abstract
Modifications and loss of bases are frequent types of DNA lesions, often handled by the base excision repair (BER) pathway. BER is initiated by DNA glycosylases, generating abasic (AP) sites that are subsequently cleaved by AP endonucleases, which further pass on nicked DNA to downstream DNA polymerases and ligases. The coordinated handover of cytotoxic intermediates between different BER enzymes is most likely facilitated by the DNA conformation. Here, we present the atomic structure of Schizosaccharomyces pombe Mag2 in complex with DNA to reveal an unexpected structural basis for nonenzymatic AP site recognition with an unflipped AP site. Two surface-exposed loops intercalate and widen the DNA minor groove to generate a DNA conformation previously only found in the mismatch repair MutS-DNA complex. Consequently, the molecular role of Mag2 appears to be AP site recognition and protection, while possibly facilitating damage signaling by structurally sculpting the DNA substrate.
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4.
The human homolog of Escherichia coli endonuclease V is a nucleolar protein with affinity for branched DNA structures.
Fladeby, C, Vik, ES, Laerdahl, JK, Gran Neurauter, C, Heggelund, JE, Thorgaard, E, Strøm-Andersen, P, Bjørås, M, Dalhus, B, Alseth, I
PloS one. 2012;(11):e47466
Abstract
Loss of amino groups from adenines in DNA results in the formation of hypoxanthine (Hx) bases with miscoding properties. The primary enzyme in Escherichia coli for DNA repair initiation at deaminated adenine is endonuclease V (endoV), encoded by the nfi gene, which cleaves the second phosphodiester bond 3' of an Hx lesion. Endonuclease V orthologs are widespread in nature and belong to a family of highly conserved proteins. Whereas prokaryotic endoV enzymes are well characterized, the function of the eukaryotic homologs remains obscure. Here we describe the human endoV ortholog and show with bioinformatics and experimental analysis that a large number of transcript variants exist for the human endonuclease V gene (ENDOV), many of which are unlikely to be translated into functional protein. Full-length ENDOV is encoded by 8 evolutionary conserved exons covering the core region of the enzyme, in addition to one or more 3'-exons encoding an unstructured and poorly conserved C-terminus. In contrast to the E. coli enzyme, we find recombinant ENDOV neither to incise nor bind Hx-containing DNA. While both enzymes have strong affinity for several branched DNA substrates, cleavage is observed only with E. coli endoV. We find that ENDOV is localized in the cytoplasm and nucleoli of human cells. As nucleoli harbor the rRNA genes, this may suggest a role for the protein in rRNA gene transactions such as DNA replication or RNA transcription.
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5.
Human ABH3 structure and key residues for oxidative demethylation to reverse DNA/RNA damage.
Sundheim, O, Vågbø, CB, Bjørås, M, Sousa, MM, Talstad, V, Aas, PA, Drabløs, F, Krokan, HE, Tainer, JA, Slupphaug, G
The EMBO journal. 2006;(14):3389-97
Abstract
Methylating agents are ubiquitous in the environment, and central in cancer therapy. The 1-methyladenine and 3-methylcytosine lesions in DNA/RNA contribute to the cytotoxicity of such agents. These lesions are directly reversed by ABH3 (hABH3) in humans and AlkB in Escherichia coli. Here, we report the structure of the hABH3 catalytic core in complex with iron and 2-oxoglutarate (2OG) at 1.5 A resolution and analyse key site-directed mutants. The hABH3 structure reveals the beta-strand jelly-roll fold that coordinates a catalytically active iron centre by a conserved His1-X-Asp/Glu-X(n)-His2 motif. This experimentally establishes hABH3 as a structural member of the Fe(II)/2OG-dependent dioxygenase superfamily, which couples substrate oxidation to conversion of 2OG into succinate and CO2. A positively charged DNA/RNA binding groove indicates a distinct nucleic acid binding conformation different from that predicted in the AlkB structure with three nucleotides. These results uncover previously unassigned key catalytic residues, identify a flexible hairpin involved in nucleotide flipping and ss/ds-DNA discrimination, and reveal self-hydroxylation of an active site leucine that may protect against uncoupled generation of dangerous oxygen radicals.