Innovative Tools for DNA Topology Probing in Human Cells Reveal a Build-Up of Positive Supercoils Following Replication Stress at Telomeres and at the FRA3B Fragile Site.

Linear unconstrained DNA cannot harbor supercoils since these supercoils can diffuse and be eliminated by free rotation of the DNA strands at the end of the molecule. Mammalian telomeres, despite constituting the ends of linear chromosomes, can hold supercoils and be subjected to topological stress. While negative supercoiling was previously observed, thus proving the existence of telomeric topological constraints, positive supercoils were never probed due to the lack of an appropriate tool. Indeed, the few tools available currently could only investigate unwound (Trioxsalen) or overwound (GapR) DNA topology (variations in twist) but not the variations in writhe (supercoils and plectonemes). To address this question, we have designed innovative tools aimed at analyzing both positive and negative DNA writhe in cells. Using them, we could observe the build-up of positive supercoils following replication stress and inhibition of Topoisomerase 2 on telomeres. TRF2 depletion caused both telomere relaxation and an increase in positive supercoils while the inhibition of Histone Deacetylase I and II by TSA only caused telomere relaxation. Moving outside telomeres, we also observed a build-up of positive supercoils on the FRA3B fragile site following replication stress, suggesting a topological model of DNA fragility for this site.

Tissue Adaptation to Environmental Cues by Symmetric and Asymmetric Division Modes of Intestinal Stem Cells.

Tissues must adapt to the different external stimuli so that organisms can survive in their environments. The intestine is a vital organ involved in food processing and absorption, as well as in innate immune response. Its adaptation to environmental cues such as diet and biotic/abiotic stress involves regulation of the proliferative rate and a switch of division mode (asymmetric versus symmetric) of intestinal stem cells (ISC). In this review, we outline the current comprehension of the physiological and molecular mechanisms implicated in stem cell division modes in the adult Drosophila midgut. We present the signaling pathways and polarity cues that control the mitotic spindle orientation, which is the terminal determinant ensuring execution of the division mode. We review these events during gut homeostasis, as well as during its response to nutrient availability, bacterial infection, chemical damage, and aging. JNK signaling acts as a central player, being involved in each of these conditions as a direct regulator of spindle orientation. The studies of the mechanisms regulating ISC divisions allow a better understanding of how adult stem cells integrate different signals to control tissue plasticity, and of how various diseases, notably cancers, arise from their alterations.