IQGAP1 and IQGAP2 are Reciprocally Altered in Hepatocellular Carcinoma

Background  

IQGAP1 and IQGAP2 are homologous members of the IQGAP family of scaffold proteins. Accumulating evidence implicates IQGAPs in tumorigenesis. We recently reported that IQGAP2 deficiency leads to the development of hepatocellular carcinoma (HCC) in mice. In the current study we extend these findings, and investigate IQGAP1 and IQGAP2 expression in human HCC.     

Quaternary structures of tumor suppressor p53 and a specific p53 DNA complex

The homotetrameric tumor suppressor p53 consists of folded core and tetramerization domains, linked and flanked by intrinsically disordered segments that impede structure analysis by x-ray crystallography and NMR. Here, we solved the quaternary structure of human p53 in solution by a combination of small-angle x-ray scattering, which defined its shape, and NMR, which identified the core domain interfaces and showed that the folded domains had the same structure in the intact protein as in fragments. We combined the solution data with electron microscopy on immobilized samples that provided medium resolution 3D maps. Ab initio and rigid body modeling of scattering data revealed an elongated cross-shaped structure with a pair of loosely coupled core domain dimers at the ends, which are accessible for binding to DNA and partner proteins. The core domains in that open conformation closed around a specific DNA response element to form a compact complex whose structure was independently determined by electron microscopy. The structure of the DNA complex is consistent with that of the complex of four separate core domains and response element fragments solved by x-ray crystallography and contacts identified by NMR. Electron microscopy on the conformationally mobile, unbound p53 selected a minor compact conformation, which resembled the closed conformation, from the ensemble of predominantly open conformations. A multipronged structural approach could be generally useful for the structural characterization of the rapidly growing number of multidomain proteins with intrinsically disordered regions.     

Structural insights into eRF3 and stop codon recognition by eRF1

Eukaryotic translation termination is mediated by two interacting release factors, eRF1 and eRF3, which act cooperatively to ensure efficient stop codon recognition and fast polypeptide release. The crystal structures of human and Schizosaccharomyces pombe full-length eRF1 in complex with eRF3 lacking the GTPase domain revealed details of the interaction between these two factors and marked conformational changes in eRF1 that occur upon binding to eRF3, leading eRF1 to resemble a tRNA molecule. Small-angle X-ray scattering analysis of the eRF1/eRF3/GTP complex suggested that eRF1''s M domain contacts eRF3''s GTPase domain. Consistently, mutation of Arg192, which is predicted to come in close contact with the switch regions of eRF3, revealed its important role for eRF1''s stimulatory effect on eRF3''s GTPase activity. An ATP molecule used as a crystallization additive was bound in eRF1''s putative decoding area. Mutational analysis of the ATP-binding site shed light on the mechanism of stop codon recognition by eRF1.     

Investigation of intravenous delivery of nanoliposomal topotecan for activity against orthotopic glioblastoma xenografts

Achieving effective treatment outcomes for patients with glioblastoma (GBM) has been impeded by many obstacles, including the pharmacokinetic limitations of antitumor agents, such as topotecan (TPT). Here, we demonstrate that intravenous administration of a novel nanoliposomal formulation of TPT (nLS-TPT) extends the survival of mice with intracranial GBM xenografts, relative to administration of free TPT, because of improved biodistribution and pharmacokinetics of the liposome-formulated drug. In 3 distinct orthotopic GBM models, 3 weeks of biweekly intravenous therapy with nLS-TPT was sufficient to delay tumor growth and significantly extend animal survival, compared with treatment with free TPT (P ≤ .03 for each tumor tested). Analysis of intracranial tumors showed increased activation of cleaved caspase-3 and increased DNA fragmentation, both indicators of apoptotic response to treatment with nLS-TPT. These results demonstrate that intravenous delivery of nLS-TPT is a promising strategy in the treatment of GBM and support clinical investigation of this therapeutic approach.     

Vascular and inflammatory stresses mediate atherosclerosis via RAGE and its ligands in apoE-/- mice

Endothelial dysfunction is a key triggering event in atherosclerosis. Following the entry of lipoproteins into the vessel wall, their rapid modification results in the generation of advanced glycation endproduct epitopes and subsequent infiltration of inflammatory cells. These inflammatory cells release receptor for advanced glycation endproduct (RAGE) ligands, specifically S100/calgranulins and high-mobility group box 1, which sustain vascular injury. Here, we demonstrate critical roles for RAGE and its ligands in vascular inflammation, endothelial dysfunction, and atherosclerotic plaque development in a mouse model of atherosclerosis, apoE-/- mice. Experiments in primary aortic endothelial cells isolated from mice and in cultured human aortic endothelial cells revealed the central role of JNK signaling in transducing the impact of RAGE ligands on inflammation. These data highlight unifying mechanisms whereby endothelial RAGE and its ligands mediate vascular and inflammatory stresses that culminate in atherosclerosis in the vulnerable vessel wall.