Although therapeutic interventions of signal-transduction cascades with targeted kinase inhibitors are a well-established strategy, drug-discovery efforts to identify targeted phosphatase inhibitors have proven challenging. Herein we report a series of allosteric, small-molecule inhibitors of wild-type p53-induced phosphatase (Wip1), an oncogenic phosphatase common to multiple cancers. Compound binding to Wip1 is dependent on a 'flap' subdomain located near the Wip1 catalytic site that renders Wip1 structurally divergent from other members of the protein phosphatase 2C (PP2C) family and that thereby confers selectivity for Wip1 over other phosphatases. Treatment of tumor cells with the inhibitor GSK2830371 increases phosphorylation of Wip1 substrates and causes growth inhibition in both hematopoietic tumor cell lines and Wip1-amplified breast tumor cells harboring wild-type TP53. Oral administration of Wip1 inhibitors in mice results in expected pharmacodynamic effects and causes inhibition of lymphoma xenograft growth. To our knowledge, GSK2830371 is the first orally active, allosteric inhibitor of Wip1 phosphatase.

SAMHD1 is a dGTP-activated deoxynucleoside triphosphate triphosphohydrolase (dNTPase) whose dNTPase activity has been linked to HIV/SIV restriction. The mechanism of its dGTPactivated dNTPase function remains unclear. Recent data also indicate that SAMHD1 regulates retrotransposition of LINE-1 elements. Here we report the 1.8-Å crystal structure of homotetrameric SAMHD1 in complex with the allosteric activator and substrate dGTP/dATP. The structure indicates the mechanism of dGTP-dependent tetramer formation, which requires the cooperation of three subunits and two dGTP/dATP molecules at each allosteric site. Allosteric dGTP binding induces conformational changes at the active site, allowing a more stable interaction with the substrate and explaining the dGTP-induced SAMHD1 dNTPase activity. Mutations of dGTP binding residues in the allosteric site affect tetramer formation, dNTPase activity and HIV-1 restriction. dGTP-triggered tetramer formation is also important for SAMHD1-mediated LINE-1 regulation. The structural and functional information provided here should facilitate future investigation of SAMHD1 function, including dNTPase activity, LINE-1 modulation and HIV-1 restriction.

Achieving rapid and effective hemostasis on irregularly shaped, non‐compressible visceral, and high‐pressure arterial bleeding wounds remains a critical clinical challenge. Herein, an ultrafast self‐gelling and wet adhesive polyethyleneimine/polyacrylic acid/quaternized chitosan (PEI/PAA/QCS) powder is reported as the hemostatic material and wound dressing. PEI/PAA/QCS powder deposited on bleeding wounds can rapidly absorb a large amount of blood to concentrate coagulation factors. Meanwhile, the powder can form an adhesive hydrogel in situ within 4 s upon hydration to form a pressure‐resistant physical barrier. Furthermore, PEI/PAA/QCS hydrogels can aggregate blood cells and platelets to enhance hemostasis. Depositing PEI/PAA/QCS powder on various bleeding wounds, including at the liver and heart, high‐pressure femoral artery and tail vein of rats, arrests the bleeding around 10 s with no rebleeding after ten minutes. Excellent hemostasis of PEI/PAA/QCS powder is further demonstrated against massive hemorrhage in porcine spleen and liver in vivo, which are non‐compressible organs with abundant blood supply. In addition, the powder can be used as a wound dressing to promote the healing of the full‐thickness skin wounds. The advantages of PEI/PAA/QCS powder including rapid and effective hemostasis, effective wound healing, easy usage, low cost, and adaptability to fit complex target sites make it a promising biomaterial for surgical applications.

Achieving strong adhesion of bioadhesives on wet tissues remains a challenge and an acute clinical demand because of the interfering interfacial water and limited adhesive-tissue interactions. Here we report a self-gelling and adhesive polyethyleneimine and polyacrylic acid (PEI/PAA) powder, which can absorb interfacial water to form a physically cross-linked hydrogel in situ within 2 seconds due to strong physical interactions between the polymers. Furthermore, the physically cross-linked polymers can diffuse into the substrate polymeric network to enhance wet adhesion. Superficial deposition of PEI/PAA powder can effectively seal damaged porcine stomach and intestine despite excessive mechanical challenges and tissue surface irregularities. We further demonstrate PEI/PAA powder as an effective sealant to enhance the treatment outcomes of gastric perforation in a rat model. The strong wet adhesion, excellent cytocompatibility, adaptability to fit complex sites, and easy synthesis of PEI/PAA powder make it a promising bioadhesive for numerous biomedical applications.

The ability of Saccharomyces cerevisiae to form morphologically complex colony-like structures called mats requires expression of the cell surface glycoprotein Flo11p and growth on a semisolid surface. As the mat grows, it forms two visually distinct populations called the rim (edge of the mat) and the hub (interior of the mat), which can be physically separated from one another based on their agar adherence properties. Here, we show that growth of the mat on a semisolid agar surface creates concentric glucose and pH gradients in the medium that are required for the differentiation of the hub and rim. Disruption of the pathways that respond to changing levels of glucose block mat formation by decreasing FLO11 expression. However, in wild-type cells, Flo11p is expressed in both portions of the structure. The difference in adherence between the rim and hub appears to be a consequence of the reduced adherence of Flo11p at the elevated pH of the rim.Microbes exhibit "multicellular" behaviors such as swarming and the formation of colonies, fruiting bodies, and biofilms (1,12,26,27,28). All of these behaviors depend on cells that interact with one another and the local environment. For example, fruiting body formation in Myxococcus xanthus occurs in response to starvation conditions (12). Biofilm formation is regulated by more diverse stimuli, depending on the microbe, but can be divided into two basic categories, surface conditions and nutrient conditions (15,16,20). Many of these multicellular behaviors depend upon a solid support and are not manifest in liquid cultures.Mat formation in the baker's yeast Saccharomyces cerevisiae has many of the features of other microbial multicellular behaviors. The formation of mats is dependent upon the nature of the surface, the concentration of glucose, and the genetic background of the strain (24). Mats are formed on semisolid agar surfaces (0.3%) and not in liquid, and experiments have shown that the surface is required for the initial formation of this structure (19).As S. cerevisiae grows on the wet surface of a semisolid agar petri plate, it forms a mat that spreads over the agar. The mat is a structure with an interior region called the hub, which is distinguished by channels and wrinkles, and a smooth periphery called the rim (Fig. 1C). Mat formation has been shown to be dependent on the FLO11 gene: a flo11⌬ null mutant fails to form a mat but, instead, grows as a poorly spreading mass of cells (Fig. 1A) (24). FLO11 is a member of a superfamily of genes encoding cell-surface adhesin proteins found in S. cerevisiae and other yeasts including Candida albicans and Candida glabrata (31).In this report, we designed experiments to identify aspects of the semisolid surface that create differences between the rim and hub. We found that the mat forms glucose and pH gradients in the agar, and these gradients lead to alterations in mat formation. These results suggest that a higher pH in the rim may alter Flo11p function so that the mat is less adherent, thereby permitting the vang...

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SAMHD1 is the only known eukaryotic deoxynucleoside triphosphate triphosphohydrolase (dNTPase) and is a major regulator of intracellular dNTP pools. It has been reported to be a potent inhibitor of retroviruses such as HIV-1 and endogenous retrotransposons. Previous crystal structures have revealed that SAMHD1 is activated by dGTP-dependent tetramer formation. However, recent data have indicated that the primary activator of SAMHD1 is GTP, not dGTP. Therefore, how its dNTPase activity is regulated needs to be further clarified. Here, five crystal structures of the catalytic core of SAMHD1 in complex with different combinations of GTP and dNTPs are reported, including a GTP-bound dimer and four GTP/dNTP-bound tetramers. The data show that human SAMHD1 contains two unique activator-binding sites in the allosteric pocket. The primary activator GTP binds to one site and the substrate dNTP (dATP, dCTP, dUTP or dTTP) occupies the other. Consequently, both GTP and dNTP are required for tetramer activation of the enzyme. In the absence of substrate binding, SAMHD1 adopts an inactive dimer conformation even when complexed with GTP. Furthermore, SAMHD1 activation is regulated by the concentration of dNTP. Thus, the level of dNTP pools is elegantly regulated by the self-sensing ability of SAMHD1 through a novel activation mechanism.

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