Wnt/β-catenin signaling plays a central role in development and is also involved in a diverse array of diseases. Binding of Wnts to the coreceptors Frizzled and LRP6/5 leads to phosphorylation of PPPSPxS motifs in the LRP6/5 intracellular region and the inhibition of GSK3β bound to the scaffold protein Axin. However, it remains unknown how GSK3β is specifically inhibited upon Wnt stimulation. Here, we show that overexpression of the intracellular region of LRP6 containing a Ser/Thr rich cluster and a PPPSPxS motif impairs the activity of GSK3β in cells. Synthetic peptides containing the PPPSPxS motif strongly inhibit GSK3β in vitro only when they are phosphorylated. Microinjection of these peptides into Xenopus embryos confirms that the phosphorylated PPPSPxS motif potentiates Wnt-induced second body axis formation. In addition, we show that the Ser/Thr rich cluster of LRP6 plays an important role in LRP6 binding to GSK3β. These observations demonstrate that phosphorylated LRP6/5 both recruits and directly inhibits GSK3β using two distinct portions of its cytoplasmic sequence, and suggest a novel mechanism of activation in this signaling pathway.

AIMP2/p38 is a scaffolding protein required for the assembly of the macromolecular tRNA synthetase complex. Here, we describe a previously unknown function for AIMP2 as a positive regulator of p53 in response to genotoxic stresses. Depletion of AIMP2 increased resistance to DNA damage-induced apoptosis, and introduction of AIMP2 into AIMP2-deficient cells restored the susceptibility to apoptosis. Upon DNA damage, AIMP2 was phosphorylated, dissociated from the multi-tRNA synthetase complex, and translocated into the nuclei of cells. AIMP2 directly interacts with p53, thereby preventing MDM2-mediated ubiquitination and degradation of p53. Mutations in AIMP2, affecting its interaction with p53, hampered its ability to activate p53. Nutlin-3 recovered the level of p53 and the susceptibility to UV-induced cell death in AIMP2-deficient cells. This work demonstrates that AIMP2, a component of the translational machinery, functions as proapoptotic factor via p53 in response to DNA damage.A minoacyl-tRNA synthetases (ARSs) are the enzymes that ligate specific amino acids to tRNAs before protein synthesis. In higher eukaryotic systems, nine different ARSs form an intriguing macromolecular complex with three nonenzymatic factors called ARS-interacting, multifunctional proteins (AIMPs) (1, 2). Many of these complex-forming ARSs, as well as AIMPs, play diverse regulatory roles that are not directly related to protein synthesis (2). Among the three AIMPs, AIMP1 is secreted as a cytokine working in immune, angiogenesis, and wound-healing processes (3-7) and also functions as a hormone controlling glucose homeostasis (8). AIMP3 is a tumor suppressor required for chromosome integrity (9, 10). Although AIMP2 is critical for the assembly of the multi-ARS complex (11), it also suppresses cell proliferation via down-regulation of c-Myc (12). In addition, AIMP2 was shown to be involved in Parkinson's disease, inducing neural cell death (13). However, it is yet to be determined how AIMP2 is involved in the control of cell death. In this work, we investigated the functional significance and molecular behavior of AIMP2 during the control of cell death and the relationship of AIMP2 associated with the multi-ARS complex and its proapoptotic activity. Results AIMP2-Deficient Cells AreResistant to Cell Death. To see the importance of AIMP2 during the control of cell death, we subjected 12.5-d AIMP2 ϩ/ϩ and AIMP2 Ϫ/Ϫ mouse embryonic fibroblasts (MEFs) to UV irradiation and compared their apoptotic sensitivity. The apoptotic cells, indicated by the subG1 portion, were increased Ϸ3-fold by UV irradiation in AIMP2 ϩ/ϩ but not in AIMP2 Ϫ/Ϫ cells (Fig. 1A). Transfection of AIMP2 into AIMP2 Ϫ/Ϫ MEFs restored the apoptotic sensitivity to UV irradiation (Fig. 1B). We also compared the apoptotic response of AIMP2 ϩ/ϩ and AIMP2 Ϫ/Ϫ MEFs to UV irradiation by monitoring caspase-3 activation. Procaspase-3 cleavage resulting in caspase-3 generation was observed in AIMP2 ϩ/ϩ but not in AIMP2 Ϫ/Ϫ cells (Fig. 1C). Suppression of AIMP2 via gene-specific siRNA...

Human fragile histidine triad (FHIT) is known to be a putative tumor suppressor gene because of frequent deletion and mutation in various kinds of human cancers, in particular, human lung cancer. However, its biological function as tumor suppressor gene, has not been demonstrated. In fact, there are conflict results on the role of FHIT. Oncogenic property of FHIT has been proposed such as overeactivation of ATR/CHK1 pathway in FHIT deficient cells, and low tumor incidence in FHIT-deficient mice. In contrast, tumor suppressive functions such as cell cycle inhibition and induction of apoptosis also proposed. Here we provide the evidences about unexpected working mechanism of FHIT as a tumor alarm gene. FHIT can awake the intracellular tumor suppressive system through reduction or deletion itself. FHIT reduction or deleted cells significantly increase the p21/WAF1 expression as well as p14/ARF-p53. It is achieved at posttranslation level and p53-dependent manner. So loss of FHIT can sequester the cell cycle in G1 phase. FHIT reduction can be achieved by oncogene activation and DNA damage. Thus, FHIT-deficient cells did not response to additional tumor risk and accumulation of additional genetic mutation is easily fixed in genome, which may contribute to tumor progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A56.