Estimating exotic gene flow into native pine stands: zygotic vs. gametic components.

Monitoring contemporary gene flow from widespread exotic plantations is becoming an important problem in forest conservation genetics. In plants, where both seed and pollen disperse, three components of exotic gene flow with potentially unequal consequences should be, but have not been, explicitly distinguished: zygotic, male gametic and female gametic. Building on a previous model for estimating contemporary rates of zygotic and male gametic gene flow among plant populations, we present here an approach that additionally estimates the third (female gametic) gene flow component, based on a combination of uni- and biparentally inherited markers. Using this method and a combined set of chloroplast and nuclear microsatellites, we estimate gene flow rates from exotic plantations into two Iberian relict stands of maritime pine (Pinus pinaster) and Scots pine (Pinus sylvestris). Results show neither zygotic nor female gametic gene flow but moderate (6-8%) male gametic introgression for both species, implying significant dispersal of pollen, but not of seeds, from exotic plantations into native stands shortly after introduced trees reached reproductive maturity. Numerical simulation results suggest that the model yields reasonably accurate estimates for our empirical data sets, especially for larger samples. We discuss conservation management implications of observed levels of exposure to nonlocal genes and identify research needs to determine potentially associated hazards. Our approach should be useful for plant ecologists and ecosystem managers interested in the vectors of contemporary genetic connectivity among discrete plant populations.

Protein kinase CK2 as regulator of cell survival: implications for cancer therapy.

Recent studies have generated sufficient information to warrant a consideration of protein kinase CK2 as a potential target for cancer therapy. CK2 is a ubiquitous and highly conserved protein serine/threonine kinase that has long been considered to play a role in cell growth and proliferation. It is essential for cell survival, and considerable evidence suggests that it can also exert potent suppression of apoptosis in cells. This is important since the cancer phenotype is characterized by deregulation of not only proliferation but also of apoptosis. In normal cells, the level of CK2 appears to be tightly regulated, and cells resist a change in their intrinsic level of CK2. However, in all the cancers that have been examined an elevation of CK2 has been observed. Further, it appears that modest deregulation in the CK2 expression imparts a potent oncogenic potential to the cells. Disruption of CK2 by treatment of cells with antisense CK2 results in induction of apoptosis in a time and dose-dependent manner. Thus, we propose that down-regulation of CK2 by employing specific strategies to deliver antisense CK2 in vivo could have a potential role in cancer therapy.

A wound-induced [Ca2+]i increase and its transcriptional activation of immediate early genes is important in the regulation of motility.

Upon mechanical wounding of a confluent quiescent monolayer, cells move into the denuded zone. However, it is not well known what signaling cascade connects release from contact inhibition to cell movement at the wound edge. Mechanical wounding induced an increase in the concentration of intracellular free Ca2+ ([Ca2+]i) in endothelial cells at the wound edge. The [Ca2+]i signal was required for the transcriptional activation of two immediate early genes (IEGs), c-fos and c-jun, since blocking Ca2+ influx with Gd3+ or EGTA reduced IEG transcription, while augmenting Ca2+ influx increased IEG transcription. The transcriptional activation of the IEGs depended on protein kinase C and calmodulin-dependent protein kinase since treatment with the inhibitors Calphostin C and KN-62 significantly reduced IEG expression. Briefly blocking Ca2+ influx also produced a long-term reduction of cell motility, while augmenting Ca2+ influx increased cell motility. To evaluate whether expression of IEGs might control cell movement, we microinjected sense or antisense cDNA to c-fos into cells after wounding. Antisense c-fos cDNA inhibited motility, while sense cDNA increased motility rates. These results suggested that the [Ca2+]i rise, induced by wounding, regulated the initiation of subsequent motility through the transcriptional activation of IEGs during wounding.

Quantitative assessment of leading edge adhesion: reattachment kinetics modulated by injury-derived intracellular calcium predict wound closure rates in endothelial monolayers.

Migrating cells continually develop new substrate attachments at the leading edge (LE) in order to maintain traction for movement. This study evaluates the relationship between LE adhesion and wound closure by modulating injury-derived intracellular free Ca2+ ([Ca2+]i) signaling in endothelial cell (EC) monolayers following scrape-wounding. These data show that brief treatment with increased extracellular Ca2+ ([Ca2+]e) during wounding accelerated wound area closure rates by 50-65%, while brief treatments with calcium influx inhibitors reduced rates by 30-50%. Fura-2 studies in wounded monolayers indicated supranormal [Ca2+]e during wounding increased (by 52%), while influx-inhibitors decreased (by 36%) the percentage of cells exhibiting elevated plateau [Ca2+]i levels. Quantitative time-lapse interference reflection microscopy (IRM) together with indirect alphavbeta3 integrin immunofluorescence was used to measure the effects of 100 microM Gd3+ and 5 mM [Ca2+]e treatment on fractional LE adhesion after wounding. Influx inhibition blocked development of increased injury-derived LE adhesion. Measurements indicated a linear relationship (r2 = 0.99, 0.98) between LE adhesion, development rates (quantified as an association rate constant) and steady state wound closure rates. Changes in filopodial activity, as indicated by phase contrast microscopy, did not correlate with changes in wound closure rates, but an association existed between the percentile peak [Ca2+]i response and the initiation of filopodial activity, suggesting a role for filopodia in mediating Ca2+-sensitive acceleration. Taken together, our data suggest that injury-derived [Ca2+]i signaling may regulate wound closure rates by an adhesion-mediated mechanism.