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DNA polymerase theta (Polθ)-mediated end-joining (TMEJ) repairs DNA double-strand breaks and confers resistance to genotoxic agents. How Polθ is regulated at the molecular level to exert TMEJ remains poorly characterized. We find that Polθ interacts with and is PARylated by PARP1 in a HPF1-independent manner. PARP1 recruits Polθ to the vicinity of DNA damage via PARylation dependent liquid demixing, however, PARylated Polθ cannot perform TMEJ due to its inability to bind DNA. PARG-mediated de-PARylation of Polθ reactivates its DNA binding and end-joining activities. Consistent with this, PARG is essential for TMEJ and the temporal recruitment of PARG to DNA damage corresponds with TMEJ activation and dissipation of PARP1 and PAR. In conclusion, we show a two-step spatiotemporal mechanism of TMEJ regulation. First, PARP1 PARylates Polθ and facilitates its recruitment to DNA damage sites in an inactivated state. PARG subsequently activates TMEJ by removing repressive PAR marks on Polθ. Here the authors reveal a two-step spatiotemporal regulation of (Polθ)-mediated end-joining (TMEJ). First, PARP1 PARylates Polθ and facilitates its recruitment to the vicinity of DNA damage sites in an inactivated state. PARG subsequently activates TMEJ by removing repressive PAR marks on Polθ. [ABSTRACT FROM AUTHOR]

The DNA damage response (DDR) protein DNA Polymerase θ (Polθ) is synthetic lethal with homologous recombination (HR) factors and is therefore a promising drug target in BRCA1/2 mutant cancers. We discover an allosteric Polθ inhibitor (Polθi) class with 4-6 nM IC50 that selectively kills HR-deficient cells and acts synergistically with PARP inhibitors (PARPi) in multiple genetic backgrounds. X-ray crystallography and biochemistry reveal that Polθi selectively inhibits Polθ polymerase (Polθ-pol) in the closed conformation on B-form DNA/DNA via an induced fit mechanism. In contrast, Polθi fails to inhibit Polθ-pol catalytic activity on A-form DNA/RNA in which the enzyme binds in the open configuration. Remarkably, Polθi binding to the Polθ-pol:DNA/DNA closed complex traps the polymerase on DNA for more than forty minutes which elucidates the inhibitory mechanism of action. These data reveal a unique small-molecule DNA polymerase:DNA trapping mechanism that induces synthetic lethality in HR-deficient cells and potentiates the activity of PARPi. [ABSTRACT FROM AUTHOR]

Cellular temperature and pH govern many cellular physiologies, especially of cancer cells. Besides, attaining higher cellular temperature plays key role in therapeutic efficacy of hyperthermia treatment of cancer. This requires bio-compatible, non-toxic and sensitive probe with dual sensing ability to detect temperature and pH variations. In this regard, fluorescence based nano-sensors for cancer studies play an important role. Therefore, a facile green synthesis of orange carbon nano-dots (CND) with high quantum yield of 90% was achieved and its application as dual nano-sensor for imaging intracellular temperature and pH was explored. CND was synthesized from readily available, bio-compatible citric acid and rhodamine 6G hydrazide using solvent-free and simple heating technique requiring purification by dialysis. Although the particle size of 19 nm (which is quite large for CND) was observed yet CND exhibits no surface defects leading to decrease in photoluminescence (PL). On the contrary, very high fluorescence was observed along with good photo-stability. Temperature and pH dependent fluorescence studies show linearity in fluorescence intensity which was replicated in breast cancer cells. In addition, molecular nature of PL of CND was established using pH dependent fluorescence study. Together, the current investigation showed synthesis of highly fluorescent orange CND, which acts as a sensitive bio-imaging probe: an optical nano-thermal or nano-pH sensor for cancer-related studies. [ABSTRACT FROM AUTHOR]

The cytoprotective action of the synthetic resveratrol (Resv) congener, E-3,3′,5,5′-tetrahydroxystilbene (designated as HST-1) against indomethacin (IND)-induced stomach ulceration has been established using a mice model. HST-1 reversed the adverse effects of IND on several inflammatory (myeloperoxidase, cytokines, adhesion molecules etc.) and ulcer-healing (cyclooxygenases, prostaglandin, growth factors and their receptors etc.) parameters in mice. More importantly, HST-1 down-regulated TNF-α and the TNF-α-mediated activation of NF-κB and JNK/MAPK pathways that are the key determinants in the IND-gastropathy. The effect of HST-1 on all these factors was significantly better than that of Resv, misoprostol, and omeprazole. HST-1 also did not induce small intestinal mucosal injury, unlike some of the proton pump inhibitors. On the other hand, Resv reduced activation of the prosurvival ERK1/2 pathway that may explain its contraindicative property in the gastrointestinal tract. [ABSTRACT FROM AUTHOR]

Three boradiazaindacene (BODIPY) dyes with different-coloured (greenish-yellow, orange and red) fluorescence and good Stokes shifts were synthesised starting from the greenish-yellow BODIPY dye PM546. The high Stokes shifts of the dyes are due to the release of the steric strain in their excited states relative to that in the highly twisted ground states. One of these compounds might be a useful water-soluble fluorophore, whereas the other two are promising H+ sensors. [ABSTRACT FROM AUTHOR]