Light-up of nuclear G-quadruplex DNA in cells by an aggregating and red/NIR emitting dye.

The herpes simplex virus 1 (HSV-1) genome is extremely rich in guanine tracts that fold into G-quadruplexes (G4s), nucleic acid secondary structures implicated in key biological functions. Viral G4s were visualized in HSV-1 infected cells, with massive virus cycle-dependent G4-formation peaking during viral DNA replication. Small molecules that specifically interact with G4s have been shown to inhibit HSV-1 DNA replication. We here investigated the antiviral activity of TMPyP4, a porphyrin known to interact with G4s. The analogue TMPyP2, with lower G4 affinity, was used as control. We showed by biophysical analysis that TMPyP4 interacts with HSV-1 G4s, and inhibits polymerase progression in vitro; in infected cells, it displayed good antiviral activity which, however, was independent of inhibition of virus DNA replication or entry. At low TMPyP4 concentration, the virus released by the cells was almost null, while inside the cell virus amounts were at control levels. TEM analysis showed that virus particles were trapped inside cytoplasmatic vesicles, which could not be ascribed to autophagy, as proven by RT-qPCR, western blot, and immunofluorescence analysis. Our data indicate a unique mechanism of action of TMPyP4 against HSV-1, and suggest the unprecedented involvement of currently unknown G4s in viral or antiviral cellular defense pathways.

Summary: Decline in tissue NAD levels during aging is linked to aging and its associated diseases. However, the mechanism for aging-associated NAD decline remains unclear. Here we report that pro-inflammatory M1-like macrophages, but not naïve or M2 macrophages, accumulate in metabolic tissues including visceral white adipose tissue and the liver during aging. Remarkably, these M1-like macrophages highly express the NAD consuming enzyme CD38 and have enhanced CD38-dependent NADase activity. We also find that senescent cells progressively accumulate in visceral white adipose tissue during aging and that inflammatory cytokines found in the supernatant from senescent cells (Senescence associated secretory proteins, SASP) induce macrophages to proliferate and to express CD38. These results highlight a new causal link between visceral tissue senescence and tissue NAD decline during aging and represent a novel therapeutic opportunity targeting maintenance of NAD levels during aging.