Discovery of an exosite on the SOCS2-SH2 domain that enhances SH2 binding to phosphorylated ligands

Suppressor of cytokine signaling (SOCS)2 protein is a key negative regulator of the growth hormone (GH) and Janus kinase (JAK)-Signal Transducers and Activators of Transcription (STAT) signaling cascade. The central SOCS2-Src homology 2 (SH2) domain is characteristic of the SOCS family proteins and is an important module that facilitates recognition of targets bearing phosphorylated tyrosine (pTyr) residues. Here we identify an exosite on the SOCS2-SH2 domain which, when bound to a non-phosphorylated peptide (F3), enhances SH2 affinity for canonical phosphorylated ligands. Solution of the SOCS2/F3 crystal structure reveals F3 as an α-helix which binds on the opposite side of the SH2 domain to the phosphopeptide binding site. F3:exosite binding appears to stabilise the SOCS2-SH2 domain, resulting in slower dissociation of phosphorylated ligands and consequently, enhances binding affinity. This biophysical enhancement of SH2:pTyr binding affinity translates to increase SOCS2 inhibition of GH signaling

Mediators of lifestyle behaviour changes in obese pregnant women. Secondary analyses from the DALI lifestyle randomised controlled trial

A better understanding of what drives behaviour change in obese pregnant overweight women is needed to improve the effectiveness of lifestyle interventions in this group at risk for gestational diabetes (GDM). Therefore, we assessed which factors mediated behaviour change in the Vitamin D and Lifestyle Intervention for GDM Prevention (DALI) Lifestyle Study. A total of 436 women, with pre-pregnancy body mass index ≥29 kg/m 2 , ≤19 + 6 weeks of gestation and without GDM, were randomised for counselling based on motivational interviewing (MI) on healthy eating and physical activity, healthy eating alone, physical activity alone, or to a usual care group. Lifestyle was measured at baseline, and at 24–28 and 35–37 weeks of gestation. Outcome expectancy, risk perception, task self-efficacy and social support were measured at those same time points and considered as possible mediators of intervention effects on lifestyle. All three interventions resulted in increased positive outcome expectancy for GDM reduction, perceived risk to the baby and increased task self-efficacy. The latter mediated intervention effects on physical activity and reduced sugared drink consumption. In conclusion, our MI intervention was successful in increasing task self-efficacy, which was related to improved health behaviours

Investigations of the suppressor of cytokine signalling 5

© 2016 Dr. Edmond Michael LinossiThe SOCS family of proteins provide crucial negative regulation of cytokine and growth factor signalling. They achieve this through multiple mechanisms, including ubiquitination and degradation of target substrates by the proteasome, direct inhibition of enzyme activity and steric competition for key binding sites in signalling proteins. Whilst the biology and mechanisms of action have been well defined for a number of SOCS family members, a clear physiological function has not been described for SOCS5. This is due in part to discrepancies in the literature, where exogenous expression of SOCS5 in mice suggests a role in CD4+ T cells that is not supported by analysis of SOCS5- deficient mice. It has also been implicated as a negative regulator of EGFR signalling and its expression is inversely correlated with receptor levels in aggressive hepatocellular carcinoma patients. SOCS5 shares a high degree of sequence homology with Socs36E of Drosophila, which is known to regulate EGFR and JAK signalling in vivo. In this work, I aimed to identify SOCS5 interacting proteins to better understand its role. I have characterised two specific targets in JAK1 and ShcA and extended the search for interacting proteins using mass spectrometry to broadly define the SOCS5 interactome in HEK293T cells. These studies revealed an extensive network of interactions, likely mediated by the poorly defined N-terminal region of SOCS5, which was heavily modified by phosphorylation. Most interestingly, in the PyMT mouse model of breast cancer, which is driven by many of the identified SOCS5 interacting proteins, SOCS5- deficient mice exhibited accelerated disease onset and tumour growth. These data show for the first time that SOCS5 plays an important protective role in the mammary gland. This was supported by analyses of SOCS5 in human breast cancer that shows patients with low SOCS5 expression exhibit a worse clinical outcome and more aggressive disease

State of the structure address on MET receptor activation by HGF

The MET receptor tyrosine kinase (RTK) and its cognate ligand hepatocyte growth factor (HGF) comprise a signaling axis essential for development, wound healing and tissue homeostasis. Aberrant HGF/MET signaling is a driver of many cancers and contributes to drug resistance to several approved therapeutics targeting other RTKs, making MET itself an important drug target. In RTKs, homeostatic receptor signaling is dependent on autoinhibition in the absence of ligand binding and orchestrated set of conformational changes induced by ligand-mediated receptor dimerization that result in activation of the intracellular kinase domains. A fundamental understanding of these mechanisms in the MET receptor remains incomplete, despite decades of research. This is due in part to the complex structure of the HGF ligand, which remains unknown in its full-length form, and a lack of high-resolution structures of the complete MET extracellular portion in an apo or ligand-bound state. A current view of HGF-dependent MET activation has evolved from biochemical and structural studies of HGF and MET fragments and here we review what these findings have thus far revealed

A survey of the kinome pharmacopeia reveals multiple scaffolds and targets for the development of novel anthelmintics

International audienceAbstract Over one billion people are currently infected with a parasitic nematode. Symptoms can include anemia, malnutrition, developmental delay, and in severe cases, death. Resistance is emerging to the anthelmintics currently used to treat nematode infection, prompting the need to develop new anthelmintics. Towards this end, we identified a set of kinases that may be targeted in a nematode-selective manner. We first screened 2040 inhibitors of vertebrate kinases for those that impair the model nematode Caenorhabditis elegans . By determining whether the terminal phenotype induced by each kinase inhibitor matched that of the predicted target mutant in C. elegans , we identified 17 druggable nematode kinase targets. Of these, we found that nematode EGFR, MEK1, and PLK1 kinases have diverged from vertebrates within their drug-binding pocket. For each of these targets, we identified small molecule scaffolds that may be further modified to develop nematode-selective inhibitors. Nematode EGFR, MEK1, and PLK1 therefore represent key targets for the development of new anthelmintic medicines

Suppressor of cytokine signaling (SOCS) 5 utilises distinct domains for regulation of JAK1 and interaction with the adaptor protein Shc-1

Suppressor of Cytokine Signaling (SOCS)5 is thought to act as a tumour suppressor through negative regulation of JAK/STAT and epidermal growth factor (EGF) signaling. However, the mechanism/s by which SOCS5 acts on these two distinct pathways is unclear. We show for the first time that SOCS5 can interact directly with JAK via a unique, conserved region in its N-terminus, which we have termed the JAK interaction region (JIR). Co-expression of SOCS5 was able to specifically reduce JAK1 and JAK2 (but not JAK3 or TYK2) autophosphorylation and this function required both the conserved JIR and additional sequences within the long SOCS5 N-terminal region. We further demonstrate that SOCS5 can directly inhibit JAK1 kinase activity, although its mechanism of action appears distinct from that of SOCS1 and SOCS3. In addition, we identify phosphoTyr317 in Shc-1 as a high-affinity substrate for the SOCS5-SH2 domain and suggest that SOCS5 may negatively regulate EGF and growth factor-driven Shc-1 signaling by binding to this site. These findings suggest that different domains in SOCS5 contribute to two distinct mechanisms for regulation of cytokine and growth factor signaling

Structure and Functional Characterization of the Conserved JAK Interaction Region in the Intrinsically Disordered N‑Terminus of SOCS5

SOCS5 can negatively regulate both JAK/STAT and EGF-receptor pathways and has therefore been implicated in regulating both the immune response and tumorigenesis. Understanding the molecular basis for SOCS5 activity may reveal novel ways to target key components of these signaling pathways. The N-terminal region of SOCS5 coordinates critical protein interactions involved in inhibition of JAK/STAT signaling, and a conserved region within the N-terminus of SOCS5 mediates direct binding to the JAK kinase domain. Here we have characterized the solution conformation of this conserved JAK interaction region (JIR) within the largely disordered N-terminus of SOCS5. Using nuclear magnetic resonance (NMR) chemical shift analysis, relaxation measurements, and NOE analysis, we demonstrate the presence of preformed structural elements in the JIR of mouse SOCS5 (mSOCS5_175–244), consisting of an α-helix encompassing residues 224–233, preceded by a turn and an extended structure. We have identified a phosphorylation site (Ser211) within the JIR of mSOCS5 and have investigated the role of phosphorylation in modulating JAK binding using site-directed mutagenesis

Elevated cytokine and chemokine production in lungs of <i>Socs4^R108X/R108X</i> is associated with an increased influx of T cells.

<p>(A) Cytokine and chemokine levels were analysed by ELISA and Bioplex in lung homogenates at day 3 post-infection with X31 virus. Mean data ± S.E.M. are shown for biological replicates (n = 4 for Balb/c, n = 5 for <i>Socs4^R108X/R108X</i>), * indicates p<0.05, **<0.005. (B) Phenotypic analysis of lung hematopoietic subsets in <i>Socs4^R108X/R108X</i> and Balb/c mice at day 3 post-infection. Flow cytometry analysis was performed on homogenized lungs and extracted BAL. Data plotted include combined cell numbers from lungs and BAL, * indicates p<0.05, **<0.005. (C) Expression of <i>Socs4</i> mRNA in immune cells recovered from BAL and in the lungs of Balb/c mice infected with X31 virus. Mean data ± S.E.M. are shown for n = 3 biological replicates, u/inf = uninfected.</p

Reduced downregulation of CD62L expression on <i>Socs4^R108X/R108X</i> CD8 cells in following X31 influenza infection.

<p>(A) Flow cytometric analysis showing the gating strategy and percentages of CD62L^hi and CD62L^l°CD8 T cells on day 5 post-infection with X31 influenza virus. Representative dot plots are shown from control Balb/c and <i>Socs4^R108X/R108X</i> mice. (B) Total CD8 T cell numbers in the lymph node (MLN), lungs (BAL) and spleen (SPL) of <i>Socs4^R108X/R108X</i> and Balb/C mice on day d5, d6 and d7 following infection with X31 influenza virus. (C) Total number and percentages of CD62L^hi and CD62L^l°CD8 cells in the MLN of <i>Socs4^R108X/R108X</i> and Balb/C mice following X31 influenza virus infection.</p