细胞功能测定

The eight SOCS (Suppressor of Cytokine Signaling) proteins encoded in the human genome all contain a C-terminal domain, the SOCS box, that allows them to function as E3 ubiquitin ligases and thereby catalyze the ubiquitination of components of the JAK/STAT signaling pathway. This activity is ...

The suppressors of cytokine signalling (SOCS) protein family consist of eight members (SOCS 1–7, and CIS). SOCS1 and SOCS3 are the best-studied family members and have been shown to act as negative feedback inhibitors of the JAK/STAT signalling pathway. To study the physiological roles of the SO ...

The Suppressor of Cytokine Signalling (SOCS) proteins are a family of negative regulators characterized by a central SH2 domain and C-terminal SOCS box motif. Cytokine Inducible SH2-containing protein (CIS), SOCS1, 2 and 3 are rapidly upregulated in response to cytokine stimulation and ...

STAT proteins are activated by diverse cellular stimuli including cytokine and growth factor receptor signaling, proto-oncogene and oncogene expression, and cellular stress mediators. In most cases, canonical STAT activation by a particular treatment or cellular condition r ...

The transcription factor Signal Tranducer and Activator of Transcription 5 (STAT5) plays an important role in many biological processes. To study STAT5 biology, several different constructs have been designed that render STAT5 constitutively active. These constructs have now be ...

Multiple experimental tools have demonstrated that cytokine-induced STAT activation entails the transition of dimer conformations rather than de novo dimerization. In this chapter, we describe the utilization of analytical ultracentrifugation (AUC) as a powerful techniq ...

The ability to observe the dynamic localization of a protein in living cells can provide critical insight to its mode of action and functional molecular interactions. To this purpose, green fluorescent protein (GFP) has served as a powerful tool to tag STAT proteins for microscopic visualiz ...

Phosphorylation of signal transducers and activators of transcription (STATs) indicates their involvement in active signaling. Here we describe immunohistochemical staining procedures for detection and identification of the cellular localization of phospho-STAT2 ...

Flow cytometry can be used to study STAT phosphorylation on a per cell basis. Cells are fixed, permeablized, and stained with antibodies that specifically bind to the phosphorylated form of the STAT protein. This allows the tyrosine phosphorylation of a single STAT to be studied within a heterog ...

Acetylation of signal transducer and activator of transcription (STAT) proteins has been recognized as a significant mechanism for the regulation of their cellular functions. Site-specific antibodies are available only for a minority of STATs. The detection of acetylated STATs by i ...

Here we describe the preparation of nuclear extracts and the electrophoretic mobility shift assay (EMSA) for the detection of STAT species. We use the method for the investigation of STAT1 and STAT3 homo- and heterodimers and show how the preparation of the extracts can influence the distribu ...

Mutations in the JAK2 gene are prevalent in the human myeloid malignancies, being present in virtually all cases of polycythemia vera, and a significant proportion of patients with other myeloproliferative disorders. Various methods for the detection of acquired mutations in this ge ...

The JAK family of protein tyrosine kinases are now recognized as important participants in a wide range of pathologies, from cancer to inflammatory diseases. In the last decade, the drive to develop drugs targeting members of this family has begun to deliver a panel of small molecule inhibitors of ...

The JAK/STAT signaling pathway has essential roles in multiple developmental processes, including stem cell maintenance, immune responses, and cellular proliferation. As a result, it has been extensively studied in both vertebrate systems and lower complexity models, such as Dro ...

The turnover rate of different protein species in a signal transduction network strongly affects the impact of the given species on the outcome of a stimulus. Whereas stable, long-lived proteins mainly account for the transmission of a signal, unstable short-lived species often comprise ...

Janus Kinases (JAKs) are the key effector kinases that initiate intracellular signalling cascades in response to cytokines and growth factors. As such, a large number of cytoplasmic proteins interact with JAKs both as substrates and as components of regulatory machinery designed to en ...

The discovery that a range of myeloproliferative diseases and leukemias are associated with Janus Kinase (JAK) mutations has highlighted the importance of JAK/STAT signalling in disease and sparked a renewed interest in developing JAK inhibitors. In vitro kinase assays are the most di ...

Jak binding to cytokine receptors has been shown to be a complex and tight interaction. When studying loss-of-function or gain-of-function mutants of the Jaks or cytokine receptors it is often necessary to know if a certain mutant still associates correctly in the context of the signaling comp ...

Activation of Janus kinases (Jaks) occurs through autophosphorylation of key tyrosine residues located primarily within their catalytic domain. Phosphorylation of these tyrosine residues facilitates access of substrates to the active site and serves as an intrinsic indicat ...

Identifying molecules that serve as markers for cell aging is a goal that has been pursued by several groups. Senescence-associated β galactosidase (SA-βgal) staining is broadly used and very easily detected. β-gal is a lysosomal enzyme strongly correlated to the progression of cell senes ...