Methods to introduce targeted double-strand breaks (DSBs) into DNA enable precise genome editing by increasing the rate at which externally supplied DNA fragments are incorporated into the genome through homologous recombination. The efficiency of these methods is limited by non-homologous end joining (NHEJ), an alternative DNA repair pathway that competes with homology-directed repair (HDR). To promote HDR at the expense of NHEJ, we targeted DNA ligase IV, a key enzyme in the NHEJ pathway, using the inhibitor Scr7. Scr7 treatment increased the efficiency of HDR-mediated genome editing using Cas9 in mammalian cell lines and in mice for all four genes examined up to 19-fold. This approach should be applicable to other customizable endonucleases, such as zinc finger nucleases and transcription activator like effector nucleases, and to non-mammalian cells with sufficiently conserved mechanisms of NHEJ and HDR.

Neuropilin-1 surface expression discriminates between nT reg cells with stable expression and Nrp1 low iT reg cells showing inducible expression under inflammatory conditions.

Regulatory T lymphocytes are essential to maintain homeostasis of the immune system, limiting the magnitude of effector responses and allowing the establishment of immunological tolerance. Two main types of regulatory T cells have been identified--natural and induced (or adaptive)-and both play significant roles in tuning down effector immune responses. Adaptive CD4(+)Foxp3(+) regulatory T (iTreg) cells develop outside the thymus under a variety of conditions. These include not only antigen presentation under subimmunogenic or noninflammatory conditions, but also chronic inflammation and infections. We speculate that the different origin of iTreg cells (noninflammatory versus inflammatory) results in distinct properties, including their stability. iTreg cells are also generated during homeostasis of the gut and in cancer, although some cancers also favor expansion of natural regulatory T (nTreg) cells. Here we review how iTreg cells develop and how they participate in immunological tolerance, contrasting, when possible, iTreg cells with nTreg cells.

SignificanceTumors are often surrounded and invaded by bone marrow-derived cells. Imaging the infiltration of such immune cells into tumors may therefore be an attractive means of detecting tumors or of tracking the response to anticancer therapy. We show that it is possible to detect these cells noninvasively by positron emission tomography (PET) via the surface markers displayed by them. The ability to monitor the immune response in the course of therapy will enable early determination of the efficacy of treatment and will inform decisions as to whether treatment should be stopped or continued. Noninvasive monitoring could therefore change how therapies are applied and assessed, to the benefit of many patients.