Ca(2+) oscillations and signaling represent a basic mechanism for controlling many cellular events. Activation of mouse eggs entrains a temporal series of Ca(2+)-dependent events that include cortical granule exocytosis, cell cycle resumption with concomitant decreases in MPF and MAP kinase activities, and recruitment of maternal mRNAs. The outcome is a switch in cellular differentiation, i.e., the conversion of the egg into the zygote. By activating mouse eggs with experimentally controlled and precisely defined Ca(2+) transients, we demonstrate that each of these events is initiated by a different number of Ca(2+) transients, while their completion requires a greater number of Ca(2+) transients than for their initiation. This combination of differential responses to the number of Ca(2+) transients provides strong evidence that a single Ca(2+) transient-driven signaling system can initiate and drive a cell into a new developmental pathway, as well as can account for the temporal sequence of cellular changes associated with early development.
Although retroviruses can integrate their DNA into a large number of sites in the host genome, factors controlling the specificity of integration remain controversial and poorly understood. To assess the effects of transcriptional activity on integration in vivo, we created quail cell clones containing a construct with a minigene cassette, whose expression is controlled by the papilloma virus E2 protein. From these clones we derived transcriptionally active subclones expressing the wild-type E2 protein and transcriptionally silent subclones expressing a mutant E2 protein that binds its target DNA but is unable to activate transcription. By infecting both clones and subclones with avian leukosis virus and using a PCR-based assay to determine viral DNA integration patterns, we were able to assess the effects of both protein binding and transcriptional activity on retroviral DNA integration. Contrary to the hypothesis that transcriptional activity enhances integration, we found an overall decrease in integration into our gene cassette in subclones expressing the wild-type E2 protein.We also found a decrease in integration into our gene cassette in subclones expressing the mutant E2 protein, but only into the protein binding region. Based on these findings, we propose that transcriptionally active DNA is not a preferred target for retroviral integration and that transcriptional activity may in fact be correlated with a decrease in integration.
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