Abstract
The Long Evans Cinnamon (LEC) rat is highly susceptible to X-irradiation due to defective DNA double-strand break repair and is a model for hepatocellular carcinogenesis. We constructed a bacterial artificial chromosome (BAC) contig of rat completely covering the region associated with radiation susceptibility. We demonstrated that defective DNA repair in LEC is fully complemented by a 200-kb BAC, 65K18 in transient and stable transfected LEC cells. Further genetic analysis determined that the radiation susceptibility region is located in a 129-kb.
We compared genome sequences of the region between Fischer 344 (F344) and LEC, and found that an intronless Rpl36a gene is inserted into the LEC genome, but not into the F344 genome. The
gene expression of Rpl36a of LEC cells was lower than that of F344 cells, suggesting that the intronless Rpl36a in the region is a pseudogene like other intronless genes. We mapped four expressed sequence tags (ESTs) in the region. The expression of three ESTs was not different between F344 and LEC cells, but the remaining one (EST#4) was not expressed in LEC cells. To determine whether the EST#4 is associated with radiation susceptibility, we measured cell survival of EST#4 knockdown cells after irradiation. Survival of cells transfected with EST#4 siRNAs was not different from that of cells transfected with the negative control siRNA. From the present study, it is not clear whether EST#4 is associated with radiation susceptibility in the LEC rat. Further study will focus on cloning full-length of EST#4 and exploring more effective siRNAs.
