Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair.

The partitioning-defective 3 (Par3), a key component in the conserved Par3/Par6/aPKC complex, plays fundamental roles in cell polarity. Herein we report the identification of Ku70 and Ku80 as novel Par3-interacting proteins through an in vitro binding assay followed by liquid chromatography-tandem mass spectrometry. Ku70/Ku80 proteins are two key regulatory subunits of the DNA-dependent protein kinase (DNA-PK), which plays an essential role in repairing double-strand DNA breaks (DSBs). We determined that the nuclear association of Par3 with Ku70/Ku80 was enhanced by gamma-irradiation (IR), a potent DSB inducer. Furthermore, DNA-PKcs, the catalytic subunit of DNA-PK, interacted with the Par3/Ku70/Ku80 complex in response to IR. Par3 over-expression or knockdown was capable of up- or downregulating DNA-PK activity, respectively. Moreover, the Par3 knockdown cells were found to be defective in random plasmid integration, defective in DSB repair following IR, and radiosensitive, phenotypes similar to that of Ku70 knockdown cells. These findings identify Par3 as a novel component of the DNA-PK complex and implicate an unexpected link of cell polarity to DSB repair.

Immunomodulatory activities of HERBSnSENSES Cordyceps -- in vitro and in vivo studies.

The commercially available HERBSnSENSEStrade mark Cordyceps (HSCS) belongs to a cultivated strain of Cordyceps sinensis whose immunomodulatory activities has been renowned in traditional Chinese medicine (TCM) for centuries. The present report is the first that describes its immunomodulatory features through a series of in vitro and in vivo experiments. We measured, in peripheral blood mononuclear cells the in vitro effects of HSCS on the gene expression of cytokines and cytokine receptors, cytokine release, and surface expression of cytokine receptors using cDNA expression array, cytometric bead array (CBA), and immunoflorescence staining, respectively, as well as macrophage phagocytosis and monocyte production of H2O2 using flow cytometry. Sixty female BALB/c mice were fed with either HSCS (40 mg/kg/day) or water consecutively for 14 days. Proliferation, cytokine liberation, and CD3/4/8 expression of splenic cells were measured using 5-bromo-2'-deoxyuridine proliferation ELISA, CBA, and cytometry immunoflorescence staining, respectively. In vitro results demonstrated that HSCS induced the production of interleukin(IL)-1beta, IL-6, IL-10 and tumor necrosis factor alphaalpha from PBMC, augmented surface expression of CD25 on lymphocytes, and elevated macrophage phagocytosis and monocyte production of H2O2. In vivo results showed that HSCS did not induce splenomegaly and cytokine overliberation. Our results possibly provide the biochemical basis for future clinical trials.