Biology subjects

de Wit, E.

Publications and source records attributed to de Wit, E..

3 recordsLinked to original sources

bioRxiv2018

Mini viral RNAs act as innate immune agonists during influenza virus infection

Influenza A virus infection usually causes a mild to moderately severe respiratory disease in humans. However, infection with the 1918 H1N1 pandemic or highly pathogenic avian influenza viruses (HPAIV) of the H5N1 subtype, can lead to viral pneumonia, systemic disease and death. The molecular processes that determine the outcome of influenza virus infection are multifactorial and involve a complex interplay between host, viral, and bacterial factors1. However, it is generally accepted that a strong innate immune dysregulation known as cytokine storm contributes to the pathology of pandemic and avian influenza virus infections2-4. The RNA sensor Retinoic acid-inducible gene I (RIG-I) plays an important role in sensing viral infection and initiating a signalling cascade that leads to interferon (IFN) expression5. Here we show that short aberrant RNAs (mini viral RNAs; mvRNAs), produced by the viral RNA polymerase during the replication of the viral RNA genome, bind and activate the intracellular pathogen sensor RIG-I, and lead to the expression of interferon-{beta}. We find that erroneous polymerase activity, dysregulation of viral RNA replication, or the presence of avian-specific amino acids underlie mvRNA generation and cytokine expression in mammalian cells and propose an intramolecular copy-choice mechanism for mvRNA generation. By deep-sequencing RNA samples from lungs of ferrets infected with influenza viruses we show that mvRNAs are generated during infection of animal models. We propose that mvRNAs act as main agonists of RIG-I during influenza virus infection and the ability of influenza virus strains to generate mvRNAs should be considered when assessing their virulence potential.

microbiology↗

bioRxiv2017

Condensin’s ATPase Machinery Drives and Dampens Mitotic Chromosome Condensation

Chromosome condensation by condensin is essential for faithful chromosome segregation. Metazoans have two complexes, named condensin I and II. Both are thought to act by creating looped structures in DNA, but how they do so is unknown. Condensins SMC subunits together form a composite ATPase with two pseudo-symmetric ATPase sites. We reveal that these sites have opposite functions in the condensation process. One site drives condensation, while the other site rather has a dampening function. Mutation of this dampener site hyperactivates both condensin I and II complexes. We find that hyperactive condensin I efficiently shortens chromosomes in the total absence of condensin II. The two complexes form loops with different lengths, and specifically condensin II is key to the decatenation of sister chromatids and the formation of a straight chromosomal axis.

molecular biology↗

bioRxiv2017

Locus-Specific Enhancer Hubs And Architectural Loop Collisions Uncovered From Single Allele DNA Topologies

Chromatin folding is increasingly recognized as a regulator of genomic processes such as gene activity. Chromosome conformation capture (3C) methods have been developed to unravel genome topology through the analysis of pair-wise chromatin contacts and have identified many genes and regulatory sequences that, in populations of cells, are engaged in multiple DNA interactions. However, pair-wise methods cannot discern whether contacts occur simultaneously or in competition on the individual chromosome. We present a novel 3C method, Multi-Contact 4C (MC-4C), that applies Nanopore sequencing to study multi-way DNA conformations of tens of thousands individual alleles for distinction between cooperative, random and competing interactions. MC-4C can uncover previously missed structures in sub-populations of cells. It reveals unanticipated cooperative clustering between regulatory chromatin loops, anchored by enhancers and gene promoters, and CTCF and cohesin-bound architectural loops. For example, we show that the constituents of the active b-globin super-enhancer cooperatively form an enhancer hub that can host two genes at a time. We also find cooperative interactions between further dispersed regulatory sequences of the active proto-cadherin locus. When applied to CTCF-bound domain boundaries, we find evidence that chromatin loops can collide, a process that is negatively regulated by the cohesin release factor WAPL. Loop collision is further pronounced in WAPL knockout cells, suggestive of a \"cohesin traffic jam\". In summary, single molecule multi-contact analysis methods can reveal how the myriad of regulatory sequences spatially coordinate their actions on individual chromosomes. Insight into these single allele higher-order topological features will facilitate interpreting the consequences of natural and induced genetic variation and help uncovering the mechanisms shaping our genome.

genetics↗