SummaryIn cancer treatment, apoptosis is a well-recognized cell death mechanism through which cytotoxic agents kill tumor cells. Here we report that dying tumor cells use the apoptotic process to generate potent growth-stimulating signals to stimulate the repopulation of tumors undergoing radiotherapy. Surprisingly, activated caspase 3, a key executioner of apoptosis, plays key roles in the growth stimulation. One downstream effector that caspase 3 regulates is prostaglandin E2, which can potently stimulates growth of surviving tumor cells. Deficiency of caspase 3 either in tumor cells or in tumor stroma caused significant tumor sensitivity to radiotherapy in xenograft or mouse tumors. In human cancer patients, higher levels of activated caspase 3 in tumor tissues are correlated with significantly increased rate of recurrence and deaths. We propose the existence of a “Phoenix Rising” pathway of cell death-induced tumor repopulation in which caspase 3 plays key roles.

The essential splicing factor human UAP56 (hUAP56) is a DExD/H-box protein known to promote prespliceosome assembly. Here, using a series of hUAP56 mutants that are defective for ATP-binding, ATP hydrolysis, or dsRNA unwindase/helicase activity, we assess the relative contributions of these biochemical functions to pre-mRNA splicing. We show that prespliceosome assembly requires hUAP56's ATP-binding and ATPase activities, which, unexpectedly, are required for hUAP56 to interact with U2AF 65 and be recruited into splicing complexes. Surprisingly, we find that hUAP56 is also required for mature spliceosome assembly, which requires, in addition to the ATP-binding and ATPase activities, hUAP56's dsRNA unwindase/helicase activity. We demonstrate that hUAP56 directly contacts U4 and U6 snRNAs and can promote unwinding of the U4/U6 duplex, and that both these activities are dependent on U2AF 65. Our results indicate that hUAP56 first interacts with U2AF 65 in an ATP-dependent manner, and subsequently with U4/U6 snRNAs to facilitate stepwise assembly of the spliceosome. Pre-mRNA splicing occurs in a ribonucleoprotein (RNP) complex called the spliceosome, which comprises a large number of proteins and several U small nuclear RNP particles (snRNPs) termed U1, U2, U4, U5, and U6. Assembly of the spliceosome on the pre-mRNA substrate proceeds in a stepwise manner and involves the recognition of intron-defining splice signals (for review, see Hertel and Graveley 2005). Spliceosome assembly is initiated by the formation of complex E, in which U1 snRNP is stably associated with the 5Ј splice site, SF1 is associated with the branchpoint, and U2 snRNP auxiliary factor (U2AF) subunits U2AF 65 and U2AF 35 are associated with the polypyrimidine (Py)-tract and 3Ј splice site, respectively. Subsequently, SF1 is replaced by U2 snRNP at the branchpoint, leading to the formation of complex A (the prespliceosome). Mature spliceosome assembly occurs upon entry of the U4/U6 и U5 tri-snRNP to form complex B, followed by structural rearrangements to form the catalytically active complex C, in which U2 and U6 snRNAs interact, and U6 replaces U1 at the 5Ј splice site.Many steps in spliceosome assembly require ATP hydrolysis and are mediated by a series of splicing factors that are members of the DExD/H-box protein family, the founding member of which is eIF-4A, a known RNA helicase (for review, see Staley and Guthrie 1998;Silverman et al. 2003;Rocak and Linder 2004). Several of these DExD/H-box splicing factors have been shown to possess an ATP-dependent RNA unwinding/helicase activity (Laggerbauer et al. 1998;Raghunathan and Guthrie 1998;Wagner et al. 1998;Wang et al. 1998) and are thought to use ATP hydrolysis as a driving force to modulate specific RNA structural rearrangements during spliceosome assembly. DExD/H-box proteins typically have a series of conserved sequence motifs; structural, mutational, and biochemical analyses have suggested roles for these motifs in ATP-binding, ATP hydrolysis (ATPase), RNA-binding, and dsRNA unwinding/hel...

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The molecular mechanisms involved in the derivation of induced pluripotent stem cells (iPSCs) from differentiated cells are poorly understood. Here we report that caspases 3 and 8, two proteases associated with apoptotic cell death, play critical roles in induction of iPSCs from human fibroblasts. Activation of caspases 3 and 8 occurs soon after transduction of iPSC-inducing transcription factors. Oct-4, a key iPSC transcription factor, is responsible for the activation. Inhibition of caspase 3 or 8 in human fibroblast cells partially or completely prevents the induction of iPSCs, respectively. Furthermore, retinoblastoma susceptibility (Rb) protein appears to be one of the factors that act downstream of the caspases. We propose that caspases are key facilitators of nuclear reprogramming in iPSC induction.