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Homeobox genes encode transcription factors which function in body axis patterning in the developing embryo. Recent evidence suggests that the maintenance of specific HOX expression patterns is necessary for regulating the homeostasis of adult tissues as well. In this study, HOXB7 transformed human mammary epithelial cells, MCF10A, to grow in minimally supplemented medium, to form colonies in Matrigel, and display resistance to ionizing radiation. Searching for protein partners of HOXB7 that might contribute to resistance to ionizing radiation, we identified four HOXB7-binding proteins by GST pull-down/affinity chromatography and confirmed their interactions by coimmunoprecipitation in vivo. Interestingly, all four HOXB7-binding proteins shared functions as genomic caretakers and included members of the DNA-dependent protein kinase holoenzyme (Ku70, Ku80, DNA-PK(cs)) responsible for DNA double-strand break repair by nonhomologous end joining pathway and poly(ADP) ribose polymerase. Exogenous and endogenous expression of HOXB7 enhanced nonhomologous end joining and DNA repair functions in vitro and in vivo, which were reversed by silencing HOXB7. This is the first mechanistic study providing definitive evidence for the involvement of any HOX protein in DNA double-strand break repair.

Reactive oxygen species have recently been demonstrated to play a role in numerous cellular signal transduction pathways. Here we investigate the involvement of H2O2 in Raf-1-mediated differentiation in the human medullary thyroid carcinoma (MTC) cell line TT:deltaRaf-1:ER. Catalase, but not Cu/Zn superoxide dismutase, completely inhibited Raf-1-induced differentiation of beta-estradiol-treated TT: deltaRaf-1:ER. In addition, catalase treatment down-regulated RET expression at both the mRNA and protein levels and induced apoptosis in the parental TT cell line and uninduced TT:deltaRaf-1:ER human MTC cells. These results implicate H2O2 as a downstream mediator of c-Raf-1-induced differentiation and as a survival factor in MTC cells.

The polyamine analogue, N1-ethyl-N11-[(cycloheptyl)methyl]-4,8-diazaundecane (CHENSpm)-induced programmed cell death in NCI H157 cells is accompanied by cytochrome c release, the loss of mitochondrial membrane potential, activation of caspase-3, caspase-mediated poly(ADP-ribose) polymerase cleavage, G2-M arrest, and DNA and nuclear fragmentation. Overexpression of Bcl-2 completely inhibits CHENSpm-induced cytochrome c release, caspase-3 activation, and poly(ADP-ribose) polymerase cleavage. However, Bcl-2 does not abrogate CHENSpm-induced programmed cell death. These results suggest that although cytochrome c release and activation of the caspase-3 protease cascade contribute to the rapid and efficient execution of apoptosis, a caspase cascade-independent pathway also exists and can be activated by CHENSpm treatment.