Chemical inhibition of DNA-PKcs impairs the activation and cytotoxicity of CD4+ helper and CD8+ effector T cells.

Modulation of T cell activity is an effective strategy for the treatment of autoimmune diseases, immune-related disorders and cancer. This highlights a critical need for the identification of proteins that regulate T cell function. The kinase DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is emerging as a potent regulator of the immune system, spurring interest in its use as a therapeutic target. In murine models of immune-related diseases including asthma and rheumatoid arthritis, treatment with small-molecule DNA-PKcs inhibitors decreased the disease severity. Additionally, DNA-PKcs inhibitors reduced T cell-mediated graft rejection in a murine allogenic skin graft model. These in vivo studies suggest the use of DNA-PKcs inhibitors as immunotherapy for autoimmune and T cell-mediated disorders. In this study, we sought to characterize further the effects of DNA-PKcs inhibitors on T cells to better understand their clinical potential. We determined that inhibition of DNA-PKcs using inhibitor NU7441 and the inhibitors currently in clinical trials for cancer therapy, M3184 and AZD7648, abrogated the activation of murine and human CD4+ and CD8+ T cells as evidenced by the reduced expression of the activation markers CD69 and CD25. Furthermore, inhibition of DNA-PKcs impeded metabolic pathways and the proliferation of activated T cells. This reduced the ability of OTI-CD8+ T cells to kill cancer cells and the expression of IFNγ and cytotoxic genes. These results highlight a critical role for DNA-PKcs in T cells and validate future studies using DNA-PKcs inhibitors as immune modulation therapy for the treatment of immune-related diseases.

Gender differences in driving under the influence (DUI) program client characteristics: Implications for treatment delivery.

The purpose of this descriptive study was to examine gender differences in the characteristics of clients in a large Driving Under the Influence (DUI) program in Southern California. We analyzed secondary de-identified data from a large DUI program for the years 2009-2014 (n = 19,619). Sociodemographic characteristics, measures of physical and mental comorbidity, and alcohol use severity measures were compared for male and female clients. Women averaged 32.85 years of age (SD = 10.70), while men were slightly older at 34.2 years (SD = 11.19). Females comprised an increasingly greater percentage of the client population over the time period studied (27.6%-30.7%). In a multivariable model, compared to male clients, females were more likely to be White non-Hispanic, not currently married, and younger. Women were more likely than men to report anxiety, depression, and a history of domestic violence. Blood alcohol content at arrest and measures of hazardous drinking did not differ significantly by gender. Results suggested that gender-specific DUI programs might be useful.

Inuit Elderly: A Systematic Review of Peer Reviewed Journal Articles.

Over the last century, Inuit have experienced rapid social changes that have greatly impacted their way of life, health, and intergenerational traditions. Although there is a growing body of research concerning Inuit youth, relatively little is known about elderly Inuit. In an effort to bridge this knowledge gap, a systematic review of peer-reviewed journal articles was conducted. This review identified a dearth of research on older Inuit, and highlighted limitations in service provision to this primarily rural and isolated population. Implications for policy and practice and recommendations for future research are also discussed.

Multicopy plasmids affect replisome positioning in Bacillus subtilis.

The DNA replication machinery, various regions of the chromosome, and some plasmids occupy characteristic subcellular positions in bacterial cells. We visualized the location of a multicopy plasmid, pHP13, in living cells of Bacillus subtilis using an array of lac operators and LacI-green fluorescent protein (GFP). In the majority of cells, plasmids appeared to be highly mobile and randomly distributed. In a small fraction of cells, there appeared to be clusters of plasmids located predominantly at or near a cell pole. We also monitored the effects of the presence of multicopy plasmids on the position of DNA polymerase using a fusion of a subunit of DNA polymerase to GFP. Many of the plasmid-containing cells had extra foci of the replisome, and these were often found at uncharacteristic locations in the cell. Some of the replisome foci were dynamic and highly mobile, similar to what was observed for the plasmid. In contrast, replisome foci in plasmid-free cells were relatively stationary. Our results indicate that in B. subtilis, plasmid-associated replisomes are recruited to the subcellular position of the plasmid. Extending this notion to the chromosome, we postulated that the subcellular position of the chromosomally associated replisome is established by the subcellular location of oriC at the time of initiation of replication.

DksA: a critical component of the transcription initiation machinery that potentiates the regulation of rRNA promoters by ppGpp and the initiating NTP.

Ribosomal RNA (rRNA) transcription is regulated primarily at the level of initiation from rRNA promoters. The unusual kinetic properties of these promoters result in their specific regulation by two small molecule signals, ppGpp and the initiating NTP, that bind to RNA polymerase (RNAP) at all promoters. We show here that DksA, a protein previously unsuspected as a transcription factor, is absolutely required for rRNA regulation. In deltadksA mutants, rRNA promoters are unresponsive to changes in amino acid availability, growth rate, or growth phase. In vitro, DksA binds to RNAP, reduces open complex lifetime, inhibits rRNA promoter activity, and amplifies effects of ppGpp and the initiating NTP on rRNA transcription, explaining the dksA requirement in vivo. These results expand our molecular understanding of rRNA transcription regulation, may explain previously described pleiotropic effects of dksA, and illustrate how transcription factors that do not bind DNA can nevertheless potentiate RNAP for regulation.