Mammalian cells have a remarkable diverse repertoire of response to genotoxic stress that damage DNA. Cellular responses to DNA damaging agents will initially exhibit gene induction, which is regulated by complex mechanism(s) and probably involves multiple signaling pathways. In this paper, we demonstrate that induction of ATF3 protein, a member of the ATF/CREB family of transcription factors, by ionizing radiation (IR) requires normal cellular p53 function. In contrast, induction of ATF3 after UV radiation (UV) or Methyl methanesulphonate (MMS) is independent of p53 status. Induction of ATF3 by DNA damage is rapid, transient, and through a transcriptional mechanism. The ATF3 promoter is induced by UV and MMS, but not by IR. In addition, ATF3 promoter can be activated by MEKK1, an upstream activator of the ERK and JNK kinase pathway, but not induced following p53 expression. Those results indicate that regulation of ATF3 induction after DNA damage utilizes both the p53-dependent andindependent pathways, and may also involve MAP kinase signaling pathways. Using the tetracyclineinducible system (tet-off), we have found that overexpression of ATF3 protein moderately suppresses cell growth. Interestingly, over-expression of ATF3 protein is able to slow down progression of cells from G1 to S phase, indicating that ATF3 protein might play a negative role in the control of cell cycle progression.

In response to DNA damage, the cell cycle checkpoint is an important biological event in maintaining genomic fidelity. Gadd45, a p53-regulated and DNA damage inducible protein, has recently been demonstrated to play a role in the G2-M checkpoint in response to DNA damage. In the current study, we further investigated the biochemical mechanism(s) involved in the GADD45-activated cell cycle G2-M arrest. Using the tetracycline-controlled system (tet-off), we established GADD45-inducible lines in HCT116 (wild-type p53) and Hela (inactivated p53 status) cells. Following inducible expression of the Gadd45 protein, cell growth was strongly suppressed in both HCT116 and Hela cells. Interestingly, HCT116 cells revealed a significant G2-M arrest but Hela cells failed to arrest at the G2-M phases, indicating that the GADD45-activated G2-M arrest requires normal p53 function. The GADD45-induced G2-M arrest was observed independent of p38 kinase activity. Importantly, induction of Gadd45 protein resulted in a reduction of nuclear cyclin B1 protein, whose nuclear localization is critical for the completion of G2-M transition. The reduced nuclear cyclin B1 levels correlated with inhibition of Cdc2/cyclin B1 kinase activity. Additionally, overexpression of cyclin B1 substantially abrogated the GADD45-induced cell growth suppression. Therefore, GADD45 inhibition of Cdc2 kinase activity through alteration of cyclin B1 subcellular localization may be an essential step in the GADD45-induced cell cycle G2-M arrest and growth suppression.

More than half of the healthy humans tested had cytotoxic antibodies to GT-KO PBMC, but allosensitized patients will be at no greater risk of rejecting a pig xenograft by a humoral mechanism.

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Two-thirds of baboon sera demonstrated cytotoxicity to GT-KO PBMC. After GT-KO organ transplantation, if an elicited antibody response develops, it is likely to cause rapid graft rejection.