Structural analysis of MDM2 RING separates degradation from regulation of p53 transcription activity

MDM2–MDMX complexes bind the p53 tumor-suppressor protein, inhibiting p53's transcriptional activity and targeting p53 for proteasomal degradation. Inhibitors that disrupt binding between p53 and MDM2 efficiently activate a p53 response, but their use in the treatment of cancers that retain wild-type p53 may be limited by on-target toxicities due to p53 activation in normal tissue. Guided by a novel crystal structure of the MDM2–MDMX–E2(UbcH5B)–ubiquitin complex, we designed MDM2 mutants that prevent E2–ubiquitin binding without altering the RING-domain structure. These mutants lack MDM2's E3 activity but retain the ability to limit p53′s transcriptional activity and allow cell proliferation. Cells expressing these mutants respond more quickly to cellular stress than cells expressing wild-type MDM2, but basal p53 control is maintained. Targeting the MDM2 E3-ligase activity could therefore widen the therapeutic window of p53 activation in tumors

Predicted Functions of MdmX in Fine-Tuning the Response of p53 to DNA Damage

Tumor suppressor protein p53 is regulated by two structurally homologous proteins, Mdm2 and MdmX. In contrast to Mdm2, MdmX lacks ubiquitin ligase activity. Although the essential interactions of MdmX are known, it is not clear how they function to regulate p53. The regulation of tumor suppressor p53 by Mdm2 and MdmX in response to DNA damage was investigated by mathematical modeling of a simplified network. The simplified network model was derived from a detailed molecular interaction map (MIM) that exhibited four coherent DNA damage response pathways. The results suggest that MdmX may amplify or stabilize DNA damage-induced p53 responses via non-enzymatic interactions. Transient effects of MdmX are mediated by reservoirs of p53∶MdmX and Mdm2∶MdmX heterodimers, with MdmX buffering the concentrations of p53 and/or Mdm2. A survey of kinetic parameter space disclosed regions of switch-like behavior stemming from such reservoir-based transients. During an early response to DNA damage, MdmX positively or negatively regulated p53 activity, depending on the level of Mdm2; this led to amplification of p53 activity and switch-like response. During a late response to DNA damage, MdmX could dampen oscillations of p53 activity. A possible role of MdmX may be to dampen such oscillations that otherwise could produce erratic cell behavior. Our study suggests how MdmX may participate in the response of p53 to DNA damage either by increasing dependency of p53 on Mdm2 or by dampening oscillations of p53 activity and presents a model for experimental investigation

Complement Enhances the Clearance of Large-sized Soluble Iga Aggregates in Rats

In the present study the involvement of the complement system (C) in the clearance of soluble IgA aggregates in the rat was studied. Monoclonal monomeric IgA (mIgA) antibody (which does not activate C) or aggregated polymeric IgA (aIgA; which activates C) were administered intravenously to phosphate-buffered saline-treated and complement-depleted [Cobra venom factor (CVF)-treated] rats and assessed for clearance from the circulation. In the control rats, mIgA was cleared in a biphasic fashion with a first half-life (T1/2) of 29.5 +/- 14.2 min and a second T1/2 of 230 +/- 176 min. No differences were observed in clearance of mIgA in CVF-treated rats as compared to PBS-treated rats. In PBS-treated rats, aIgA with a size between 20S and 150S disappeared very rapidly from the circulation with a first T1/2 of 1.1 +/- 0.4 min and a second T1/2 of 23.2 +/- 11.3 min. In CVF-treated rats the clearance of aIgA was significantly delayed as compared to that in control rats, namely with a first T1/2 of 7.3 +/- 2.6 min and a second T1/2 of 64.2 +/- 19.4 min. Immunohistochemical studies of the liver (which is the main site of clearance of aIgA) revealed that Kupffer cells (KC) are mainly responsible for the uptake of aIgA. Furthermore, in PBS-treated rats aIgA deposition was accompanied by C3 deposition in the KC. In CVF-treated rats, the percentage of KC containing aIgA was significantly lower during the first 16 min after aIgA administration as compared to PBS treated rats. In addition no detectable C3 was found in KC of CVF-treated rats. These results indicate that KC plays an important role in the clearance of large molecular weight IgA in rats and that C facilitates the clearance of these complexes from the circulation

Physical planning in place-making through design and image building

This paper discusses how physical planning works with place-making through urban design to support image creation in a municipality that aims to create a new image for its city. Two large-scale urban development projects in Sweden are analysed. The analysis focuses on official plans and documentation, and expert interviews. It distinguishes how place-making by physical planning happens through aestheticisation, the design of physical environments for specific groups of inhabitants and by granting visual and symbolic coherence to an area aligned with the new image for the city. The construction of this image is kick-started by a flagship development, but requires that subsequent projects support complementing urban visions, so as to grant coherence to the perceptions of the new image. In the process, heterogeneity, variation and conflict are tamed and urbanity is framed for "creative classes". The paper thus highlights potential dilemmas that place-making might create for the practice of physical planning in urban development and contributes to the discussions on the transformation of public sector urban planning in the context of urban entrepreneurialism