Structural insight into SUMO chain recognition and manipulation by the ubiquitin ligase RNF4

The small ubiquitin-like modifier (SUMO) can form polymeric chains that are important signals in cellular processes such as meiosis, genome maintenance and stress response. The SUMO-targeted ubiquitin ligase RNF4 engages with SUMO chains on linked substrates and catalyses their ubiquitination, which targets substrates for proteasomal degradation. Here we use a segmental labelling approach combined with solution nuclear magnetic resonance (NMR) spectroscopy and biochemical characterization to reveal how RNF4 manipulates the conformation of the SUMO chain, thereby facilitating optimal delivery of the distal SUMO domain for ubiquitin transfer

Degradation of Cdc25A by \u3b2-TrCP during S phase and in response to DNA damage

The Cdc25A phosphatase is essential for cell-cycle progression because of its function in dephosphorylating cyclin-dependent kinases. In response to DNA damage or stalled replication, the ATM and ATR protein kinases activate the checkpoint kinases Chk1 and Chk2, which leads to hyperphosphorylation of Cdc25A1\u20133. These events stimulate the ubiquitin-mediated pro- teolysis of Cdc25A1,4,5 and contribute to delaying cell-cycle progression, thereby preventing genomic instability1\u20137. Here we report that b-TrCP is the F-box protein that targets phosphory- lated Cdc25A for degradation by the Skp1/Cul1/F-box protein complex. Downregulation of b-TrCP1 and b-TrCP2 expression by short interfering RNAs causes an accumulation of Cdc25A in cells progressing through S phase and prevents the degradation of Cdc25A induced by ionizing radiation, indicating that b-TrCP may function in the intra-S-phase checkpoint. Consistent with this hypothesis, suppression of b-TrCP expression results in radioresistant DNA synthesis in response to DNA damage\u2014a phenotype indicative of a defect in the intra-S-phase checkpoint that is associated with an inability to regulate Cdc25A properly. Our results show that b-TrCP has a crucial role in mediating the response to DNA damage through Cdc25A degradation

Evaluation and Treatment of Iron Deficiency Anemia: A Gastroenterological Perspective

A substantial volume of the consultations requested of gastroenterologists are directed towards the evaluation of anemia. Since iron deficiency anemia often arises from bleeding gastrointestinal lesions, many of which are malignant, establishment of a firm diagnosis usually obligates an endoscopic evaluation. Although the laboratory tests used to make the diagnosis have not changed in many decades, their interpretation has, and this is possibly due to the availability of extensive testing in key populations. We provide data supporting the use of the serum ferritin as the sole useful measure of iron stores, setting the lower limit at 100 μg/l for some populations in order to increase the sensitivity of the test. Trends of the commonly obtained red cell indices, mean corpuscular volume, and the red cell distribution width can provide valuable diagnostic information. Once the diagnosis is established, upper and lower gastrointestinal endoscopy is usually indicated. Nevertheless, in many cases a gastrointestinal source is not found after routine evaluation. Additional studies, including repeat upper and lower endoscopy and often investigation of the small intestine may thus be required. Although oral iron is inexpensive and usually effective, there are many gastrointestinal conditions that warrant treatment of iron deficiency with intravenous iron

Key role for ubiquitin protein modification in TGFβ signal transduction

The transforming growth factor β (TGFβ) superfamily of signal transduction molecules plays crucial roles in the regulation of cell behavior. TGFβ regulates gene transcription through Smad proteins and signals via non-Smad pathways. The TGFβ pathway is strictly regulated, and perturbations lead to tumorigenesis. Several pathway components are known to be targeted for proteasomal degradation via ubiquitination by E3 ligases. Smurfs are well known negative regulators of TGFβ, which function as E3 ligases recruited by adaptors such as I-Smads. TGFβ signaling can also be enhanced by E3 ligases, such as Arkadia, that target repressors for degradation. It is becoming clear that E3 ligases often target multiple pathways, thereby acting as mediators of signaling cross-talk. Regulation via ubiquitination involves a complex network of E3 ligases, adaptor proteins, and deubiquitinating enzymes (DUBs), the last-mentioned acting by removing ubiquitin from its targets. Interestingly, also non-degradative ubiquitin modifications are known to play important roles in TGFβ signaling. Ubiquitin modifications thus play a key role in TGFβ signal transduction, and in this review we provide an overview of known players, focusing on recent advances

Cancer Genomics Identifies Regulatory Gene Networks Associated with the Transition from Dysplasia to Advanced Lung Adenocarcinomas Induced by c-Raf-1

Background: Lung cancer is a leading cause of cancer morbidity. To improve an understanding of molecular causes of disease a transgenic mouse model was investigated where targeted expression of the serine threonine kinase c-Raf to respiratory epithelium induced initialy dysplasia and subsequently adenocarcinomas. This enables dissection of genetic events associated with precancerous and cancerous lesions. Methodology/Principal Findings: By laser microdissection cancer cell populations were harvested and subjected to whole genome expression analyses. Overall 473 and 541 genes were significantly regulated, when cancer versus transgenic and non-transgenic cells were compared, giving rise to three distinct and one common regulatory gene network. At advanced stages of tumor growth predominately repression of gene expression was observed, but genes previously shown to be upregulated in dysplasia were also up-regulated in solid tumors. Regulation of developmental programs as well as epithelial mesenchymal and mesenchymal endothelial transition was a hall mark of adenocarcinomas. Additionaly, genes coding for cell adhesion, i.e. the integrins and the tight and gap junction proteins were repressed, whereas ligands for receptor tyrosine kinase such as epi- and amphiregulin were up-regulated. Notably, Vegfr- 2 and its ligand Vegfd, as well as Notch and Wnt signalling cascades were regulated as were glycosylases that influence cellular recognition. Other regulated signalling molecules included guanine exchange factors that play a role in an activation of the MAP kinases while several tumor suppressors i.e. Mcc, Hey1, Fat3, Armcx1 and Reck were significantly repressed. Finally, probable molecular switches forcing dysplastic cells into malignantly transformed cells could be identified. Conclusions/Significance: This study provides insight into molecular pertubations allowing dysplasia to progress further to adenocarcinoma induced by exaggerted c-Raf kinase activity

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference