Methanogens implicated by DNA evidence
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Apaf-1 is an evolutionarily conserved DNA sensor that switches the cell fate between apoptosis and inflammation
Apoptotic protease activating factor 1 (Apaf-1) was traditionally defined as a scaffold protein in mammalian cells for assembling a caspase activation platform known as the ‘apoptosome’ after its binding to cytochrome c. Although Apaf-1 structurally resembles animal NOD-like receptor (NLR) and plant resistance (R) proteins, whether it is directly involved in innate immunity is still largely unknown. Here, we found that Apaf-1-like molecules from lancelets, fruit flies, mice, and humans have conserved DNA sensing functionality. Mechanistically, mammalian Apaf-1 recruits receptor-interacting protein 2 (RIP2, also known as RIPK2) via its WD40 repeat domain and promotes RIP2 oligomerization to initiate NF-κB-driven inflammation upon cytoplasmic DNA recognition. Furthermore, DNA binding of Apaf-1 determines cell fate by switching the cellular processes between intrinsic stimuli-activated apoptosis and inflammation. These findings suggest that Apaf-1 is an evolutionarily conserved DNA sensor and may serve as a cell fate checkpoint, which determines whether cells initiate inflammation or undergo apoptosis by distinct ligand binding.
KorB switching from DNA-sliding clamp to repressor mediates long-range gene silencing in a multi-drug resistance plasmid
Examples of long-range gene regulation in bacteria are rare and generally thought to involve DNA looping. Here, using a combination of biophysical approaches including X-ray crystallography and single-molecule analysis for the KorB–KorA system in Escherichia coli, we show that long-range gene silencing on the plasmid RK2, a source of multi-drug resistance across diverse Gram-negative bacteria, is achieved cooperatively by a DNA-sliding clamp, KorB, and a clamp-locking protein, KorA. We show that KorB is a CTPase clamp that can entrap and slide along DNA to reach distal target promoters up to 1.5 kb away. We resolved the tripartite crystal structure of a KorB–KorA–DNA co-complex, revealing that KorA latches KorB into a closed clamp state. DNA-bound KorA thus stimulates repression by stalling KorB sliding at target promoters to occlude RNA polymerase holoenzymes. Together, our findings explain the mechanistic basis for KorB role switching from a DNA-sliding clamp to a co-repressor and provide an alternative mechanism for long-range regulation of gene expression in bacteria.
EV DNA from pancreatic cancer patient-derived cells harbors molecular, coding, non-coding signatures and mutational hotspots
DNA packaged into cancer cell-derived EV is not well appreciated. Here, we uncovered signatures of EV DNA secreted by pancreatic cancer cells. The cancer cells and non-cancer counterparts exhibit distinct low vs. high molecular weight (LMW vs. HMW) EV DNA fragments distribution, respectively. Genome sequencing and Single Nucleotide Variants analysis revealed that 95% of reads and 94% of SNVs map to noncoding regions of the genome. Given that ~1% of the human genome represents coding regions, the 5% mapping rate to coding regions suggests a non-random enrichment of certain coding regions and mutations. The LMW DNA fragments not only set cancer cells apart, but also harbor cancer specific enrichment of unique coding regions, the top nine being FAM135B, COL22A1, TSNARE1, KCNK9, ZFAT, JRK, MROH5, GSDMD, and MIR3667HG. Additionally, the cancer cells’ LMW DNA fragments exhibit dense centromeric mapping more strikingly on chromosomes 3, 7, 9, 10, 11, 13, 17, and 20. Mutational profiling turned up close to 200 mutations specific for the cancer cells. Altogether, our analyses suggest that centromeric regions might hold clues to EV DNA content from pancreatic cancer, the molecular, mutational signatures thereof, and rationalizes the need for a new approach to DNA biomarker research.
Humic acid-dependent respiratory growth of Methanosarcina acetivorans involves pyrroloquinoline quinone
Although microbial humus respiration plays a critical role in organic matter decomposition and biogeochemical cycling of elements in diverse anoxic environments, the role of methane-producing species (methanogens) is not well defined. Here we report that a major fraction of humus, humic acid reduction enhanced the growth of Methanosarcina acetivorans above that attributed to methanogenesis when utilizing the energy sources methanol or acetate, results which showed both respiratory and fermentative modes of energy conservation. Growth characteristics with methanol were the same for an identically cultured mutant deleted for the gene encoding a multi-heme cytochrome c (MmcA), results indicating MmcA is not essential for respiratory electron transport to humic acid. Transcriptomic analyses revealed that growth with humic acid promoted the upregulation of genes annotated as cell surface pyrroloquinoline quinone (PQQ)-binding proteins. Furthermore, PQQ isolated from the membrane fraction was more abundant in humic acid-respiring cells, and the addition of PQQ improved efficiency of the extracellular electron transport. Given that the PQQ-binding proteins are widely distributed in methanogens, the findings extend current understanding of microbial humus respiration in the context of global methane dynamics.
Histone H3 lysine 4 methylation recruits DNA demethylases to enforce gene expression in Arabidopsis
Patterning of DNA methylation in eukaryotic genomes is controlled by de novo methylation, maintenance mechanisms and demethylation pathways. In Arabidopsis thaliana, DNA demethylation enzymes are clearly important for shaping methylation patterns, but how they are regulated is poorly understood. Here we show that the targeting of histone H3 lysine four trimethylation (H3K4me3) with the catalytic domain of the SDG2 histone methyltransferase potently erased DNA methylation and gene silencing at FWA and also erased CG DNA methylation in many other regions of the Arabidopsis genome. This methylation erasure was completely blocked in the ros1 dml2 dml3 triple mutant lacking DNA demethylation enzymes, showing that H3K4me3 promotes the active removal of DNA methylation. Conversely, we found that the targeted removal of H3K4me3 increased the efficiency of targeted DNA methylation. These results highlight H3K4me3 as a potent anti-DNA methylation mark and also pave the way for development of more powerful epigenome engineering tools.
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