ABSTRACT
Understanding the mechanisms producing low dose ionizing radiation specific biological effects represents one of the major challenges of radiation biology. Although experimental evidence does suggest that various molecular stress response pathways may be involved in the production of low dose effects, much of the detail of those mechanisms remains elusive. We hypothesized that the regulation of various stress response pathways upon irradiation may differ from one another in complex dose-response manners, causing the specific and subtle low dose radiation effects. In the present study, the transcription level of 22 genes involved in stress responses were analyzed using RT-qPCR in normal human fibroblasts exposed to a range of gamma-doses from 1 to 200 cGy. Using the alkali comet assay, we also measured the level of DNA damages in dose-response and time-course experiments. We found non-linear dose responses for the repair of DNA damage after exposure to gamma-radiation. Alterations in gene expression were also not linear with dose for several of the genes examined and did not follow a single pattern. Rather, several patterns could be seen. Our results suggest a complex interplay of various stress response pathways triggered by low radiation doses, with various low dose thresholds for different genes.
ABSTRACT
The murine immunoglobulin heavy-chain (Igh) locus provides an important model for understanding the replication of tissue-specific gene loci in mammalian cells. We have observed two DNA replication programs with dramatically different temporal replication patterns for the Igh locus in B-lineage cells. In pro- and pre-B-cell lines and in ex vivo-expanded pro-B cells, the entire locus is replicated early in S phase. In three cell lines that exhibit the early-replication pattern, we found that replication forks progress in both directions through the constant-region genes, which is consistent with the activation of multiple initiation sites. In contrast, in plasma cell lines, replication of the Igh locus occurs through a triphasic pattern similar to that previously detected in MEL cells. Sequences downstream of the Igh-C alpha gene replicate early in S, while heavy-chain variable (Vh) gene sequences replicate late in S. An approximately 500-kb transition region connecting sequences that replicate early and late is replicated progressively later in S. The formation of the transition region in different cell lines is independent of the sequences encompassed. In B-cell lines that exhibit a triphasic-replication pattern, replication forks progress in one direction through the examined constant-region genes. Timing data and the direction of replication fork movement indicate that replication of the transition region occurs by a single replication fork, as previously described for MEL cells. Associated with the contrasting replication programs are differences in the subnuclear locations of Igh loci. When the entire locus is replicated early in S, the Igh locus is located away from the nuclear periphery, but when Vh gene sequences replicate late and there is a temporal-transition region, the entire Igh locus is located near the nuclear periphery.
