Little is known about the cellular mechanisms modulating the shift in balance from a state of survival to cell death by caspasemediated apoptosis in response to a lethal stress. Here we show that the RNA-binding protein HuR has an important function in mediating this switch. During caspase-mediated apoptosis, HuR is cleaved to generate two cleavage products (CPs). Our data demonstrate that the cleavage of HuR switches its function from being a prosurvival factor under normal conditions to becoming a promoter of apoptosis in response to a lethal stress. In the absence of an apoptotic stimuli, HuR associates with and promotes the expression of caspase-9 and prothymosin a (ProT) mRNAs, and pro-and antiapoptotic factors, respectively, both of which have been characterized as important players in determining cell fate. During the early steps of caspase-mediated apoptosis, however, the level of caspase-9 protein increases, while ProT remains unchanged. Under these conditions, the two HuR-CPs selectively bind to and stabilize caspase-9 mRNA, but do not bind to ProT. Hence, taken together, our data show that by maintaining a threshold of expression of proapoptotic factors such as caspase-9 in response to a lethal stress, the HuR-CPs help a cell to switch from resisting death to undergoing apoptosis.

In response to severe stress, apoptotic cell death is engaged. Apoptosis is a well orchestrated process that involves the activation and implication of many factors. In this study, we identified a role for the nuclear trafficking factor TRN2 (transportin 2) in cell death. TRN2 is normally responsible for the nuclear import of the RNA-binding protein HuR. During apoptosis, however, HuR accumulates in the cytoplasm. This is due to the caspase-mediated cleavage of the cytoplasmic fraction of HuR. One of the cleavage fragments generated by this processing of HuR interacts with TRN2 and thus blocks the re-import of HuR into the nucleus. This concentrates HuR in the cytoplasm, advancing apoptosis. Therefore, increasing or decreasing the levels of TRN2 has an inverse consequential effect on cell death, demonstrating for the first time the role of a nucleocytoplasmic transport factor in apoptosis.When confronted with stress, eukaryotic cells undergo different responses depending on the severity and duration of the stress (1). Although adaptation and survival are at one end of these responses, cell death can also be the outcome. Apoptotic cell death is a complex pathway involving the activation and cleavage of several proteins, some of the most important of which are cysteine-aspartic proteases (caspases). This family of proteases then cleaves specific substrates, allowing the progression of cell death (2-5). In addition to causing the organized destruction of a cell, caspase-mediated cleavage can also serve to activate target proteins that have important roles in advancing apoptosis themselves. Recently, the RNA-binding protein HuR was identified as one such factor (6). Under normal conditions, HuR is an important regulator of gene expression, controlling the localization, stabilization, and translation of many different mRNAs (7,8). The targets of HuR are involved in many cellular processes, including cell growth, differentiation, and survival and apoptotic cell death. Likewise, the importance of HuR in regulating these processes has been shown, given that its absence or overexpression in different systems can have potent effects on cell physiology (6, 9 -12).The role of HuR as a promoter of cell death was identified when it was found that caspase-3 and caspase-7 cleave HuR in response to lethal stress (6), such as with the use of the apoptotic stimulator drug staurosporine (STS). 4 The cleavage of HuR, occurring at Asp-226 of this protein, yields two cleavage products: HuR-CP1 (26 kDa) and HuR-CP2 (8 kDa). Mutation of this cleavage site prevents apoptosis, whereas providing these two cleavage products can stimulate cell death under nonlethal conditions (13). It was shown that HuR-CP2 helps promote apoptosis by binding to putative HLA-associated protein-I (PHAPI) (6), an activator of apoptosis and a known ligand of full-length HuR. However, the implication of other protein partners has not yet been explored.Intriguingly, prior to its cleavage, HuR, which typically shuttles between the nucleus and the cytoplasm, must ha...