The relationship of natural killer cell counts, perforin mRNA and CD2 expression to post-exercise natural killer cell activity in humans

The aim of this study was to further investigate the mechanism of suppression of natural killer (NK) cell cytotoxic activity In peripheral blood following strenuous exercise. Blood was collected for analysis of NK cell concentration, cytotoxic activity, CD2 surface expression and perforin gene expression from runners (RUN, n = 6) and resting controls (CONTROL, n = 4) pre-exercise, 0, 1.5, 5, and 24 h following a 60-min treadmill run at 80% of VO2 peak. Natural killer cytotoxic activity, measured using a whole blood chromium release assay, fluctuated minimally in the CONTROL group and increased by 63% and decreased by 43% 0 and 1.5 h post-exercise, respectively, in the RUN group (group x time, P < 0.001). Lytic index (cytotoxic activity per cell) did not change. Perforin mRNA, measured using quantitative real-time polymerase chain reaction (ORT-PCR) decreased from pre- to post-exercise and remained decreased through 24 h, The decrease from pre- to 0 In post-exercise was seen predominately in the RUN group and was inversely correlated r = - 0.95) to pre-exercise perform mRNA. The NK cell surface expression of CD2 (lymphocyte function-associated antigen-2) was determined using fluorescent antibodies and flow cytometry, There was no change in the proportion of NK cells expressing CD2 or CD2 density, We conclude that (1) numerical redistribution accounted for most of the change in NK cytotoxic activity following a strenuous run, (2) decrease in perforin gene expression during the run was inversely related to pre-exercise levels but did not parallel changes in cytotoxic activity, and (3) CD2 surface expression was not affected by exercise.

A role for the GCC-box in jasmonate-mediated activation of the PDF1.2 gene of arabidopsis

The PDF1.2 gene of Arabidopsis encoding a plant defensin is commonly used as a marker for characterization of the jasmonate-dependent defense responses. Here, using PDF1.2 promoter-deletion lines linked to the beta-glucoronidase-reporter gene, we examined putative promoter elements associated with jasmonate-responsive expression of this gene. Using stably transformed plants, we first characterized the extended promoter region that positively regulates basal expression from the PDF1.2 promoter. Second, using promoter deletion constructs including one from which the GCC-box region was deleted, we observed a substantially lower response to jasmonate than lines carrying this motif. In addition, point mutations introduced into the core GCC-box sequence substantially reduced jasmonate responsiveness, whereas addition of a 20-nucleotide-long promoter element carrying the core GCC-box and flanking nucleotides provided jasmonate responsiveness to a 35S minimal promoter. Taken together, these results indicated that the GCC-box plays a key role in conferring jasmonate responsiveness to the PDF1.2 promoter. However, deletion or specific mutations introduced into the core GCC-box did not completely abolish the jasmonate responsiveness of the promoter, suggesting that the other promoter elements lying downstream from the GCC-box region may also contribute to jasmonate responsiveness. In other experiments, we identified a jasmonate- and pathogen-responsive ethylene response factor transcription factor, AtERF2, which when overexpressed in transgenic Arabidopsis plants activated transcription from the PDF1.2, Thi2.1, and PR4 (basic chitinase) genes, all of which contain a GCC-box sequence in their promoters. Our results suggest that in addition to their roles in regulating ethylene-mediated gene expression, ethylene response factors also appear to play important roles in regulating jasmonate-responsive gene expression, possibly via interaction with the GCC-box.