Inhibitor‐of‐apoptosis proteins (IAPs), including neuronal apoptosis inhibitory protein (NAIP), inhibit cell death. Other IAPs inhibit key caspase proteases which effect cell death, but the mechanism by which NAIP acts is unknown. Here we report that NAIP, through its third baculovirus inhibitory repeat domain (BIR3), binds the neuron‐restricted calcium‐binding protein, hippocalcin, in an interaction promoted by calcium. In neuronal cell lines NSC‐34 and Neuro‐2a, over‐expression of the BIR domains of NAIP (NAIP‐BIR1–3) counteracted the calcium‐induced cell death induced by ionomycin and thapsigargin. This protective capacity was significantly enhanced when NAIP‐BIR1–3 was co‐expressed with hippocalcin. Over‐expression of the BIR3 domain or hippocalcin alone did not substantially enhance cell survival, but co‐expression greatly increased their protective effects. These data suggest synergy between NAIP and hippocalcin in facilitating neuronal survival against calcium‐induced death stimuli mediated through the BIR3 domain. Analysis of caspase activity after thapsigargin treatment revealed that caspase‐3 is activated in NSC‐34, but not Neuro‐2a, cells. Thus NAIP, in conjunction with hippocalcin, can protect neurons against calcium‐induced cell death in caspase‐3‐activated and non‐activated pathways.

The ERM protein family members ezrin, radixin, and moesin are cytoskeletal effector proteins linking actin to membrane-bound proteins at the cell surface. Here we report on the cloning of myosin regulatory light chain interacting protein (MIR), a protein with an ERMhomology domain and a carboxyl-terminal RING finger, that is expressed, among other tissues, in brain. MIR is distributed in cultured COS cells, in a punctated manner as shown using enhanced green fluorescent protein (EGFP)-tagged MIR and by staining with a specific antibody for MIR. In the yeast two-hybrid system and in transfected COS cells, MIR interacts with myosin regulatory light chain B, which in turn regulates the activity of the actomyosin complex. Overexpression of MIR cDNA in PC12 cells abrogated neurite outgrowth induced by nerve growth factor (NGF) without affecting TrkA signaling. The results show that MIR, a novel ERMlike protein, affects cytoskeleton interactions regulating cell motility, such as neurite outgrowth.Dynamic changes in cell shape and movements involve interactions between proteins in the cytoskeleton and the plasma membrane. Ezrin, radixin, moesin, and the related tumor suppressor merlin link the actin cytoskeleton to membrane-bound proteins located at membrane extension sites, such as microvilli, membrane rufflings, and at cell-cell contacts (1-3). ERM 1 proteins are also involved in cell adhesion, influencing the redistribution of various cell adhesion molecules, such as ICAM-1 (4, 5). The amino-terminal domain of ERM proteins, comprising about 300 amino acids, is conserved among the different proteins and merlin and exhibits a high degree of sequence similarity (1-3). This amino terminus is related to proteins in the large band 4.1 protein superfamily (6), including some protein tyrosine phosphatases (7) and talin (8), and it is thought to bind to membrane proteins. In contrast, the interaction of ERM proteins with F-actin seems to involve both the amino-and carboxyl-terminal region (5). Merlin, however, lacks the F-actin binding site in the carboxyl terminus, but binds it in a different manner (9).ERM proteins are present in a wide variety of tissues, notably in epithelial cells in culture (10). In contrast to other tissues, less is known about the function of ERM proteins in the nervous system. During development and regeneration, neurons grow over large distances in a process controlled by extrinsic factors and cytoskeleton interactions. Some ERM proteins have been found in the growth cones of neuronal cells (11,12), suggesting an effect on neurite outgrowth. Indeed, recently it was shown by an antisense approach, that radixin and moesin are important for growth cone morphology and motility (13). Although important for dynamic interactions between cytoskeleton and membranes, the mechanism by which ERM proteins mediate their effects on cell motility is not fully understood (5). In this study, we have identified a novel ERM-like protein, called myosin regulatory light chain interacting protein (MIR), that binds to the...

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