• Journal of Life Science
• /
• v.15 no.6 s.73
• /
• pp.904-908
• /
• 2005

Recently the pathological actions of CagA of Helicobacter pylori (H. pylori) on gastric epithelial cells have been reported. CagA is directly injected into the host cytoplasm and undergoes tyrosine phosphorylation in the cells. In addition, translocated CagA forms a physical complex with SHP-2. There are two major CagA subtypes according to the amino acid sequence in the 3'region of CagA; i) the East Asian type (A-B-D of EPIYA motifs) and ii) the Western type (A-B-C of EPIYA motifs). Repeated EPIYA motifs in the 3'region of CagA are involved in the interaction with SHP-2. The East Asian type conferred stronger SHP-2 binding activity than the Westrrn type of CagA. Here we analyzed the amino acid sequences of the SHP-2 binding site of cagA gene in H. pyzori, and investigated whether there is my relationship between the diversities of cagA and the disease out-come in Korea. Most of Korean H. pylori strains showed A-B-D motifs(the East Asian type), and only one strain showed A-B-B-D motifs. In Korea, the incidence of atrophic gastritis and gastric cancer is significantly high compared with Western countries. The high frequency of the East Asian type CagA among Korean H. pylori strains would be involved in increasing the risk of gastric cancer in Korean populations.

• IMMUNE NETWORK
• /
• v.2 no.2
• /
• pp.72-78
• /
• 2002

Background: The precise mechanism by which CTLA-4 regulates T cell immune responses is still not fully understood. Previously we proposed that CTLA-4 could downregulate T cell function by modulating a signaling cascade initiated from the T cell receptor complex. The evidence for this notion comes from our findings that CTLA-4 associated with the T cell receptor zeta (TCR zeta) chain, and hence regulated TCR zeta phosphorylation by co-associated SHP-2 tyrosine phosphatase (1). In this report, we investigated whether any other signaling molecules could be involved in the CTLA-4 signaling pathway. Methods: We have taken biochemical approaches, such as immunoprecipitation followed by autoradiography or immunoblotting, to identify the molecules associated with CTLA-4. To perform these assays, we used activated primary T cells and ectopically transfected 293 cells. Various truncation mutants of CTLA-4 were used to map the interaction site on CTLA-4. Results: We found that in addition to TCR zeta and SHP-2, a recently cloned small adaptor molecule, SAP (SLAM-associated protein), was also able to associate with CTLA-4. We identified the domain of SAP association in CTLA-4 being a motif involving GVYVKM. This motif has been previously found to bind SHP-2 through its phosphorylated tyrosine interaction with SH-2 domain of SHP-2. Indeed, co-expression of SAP and SHP-2 reduced their binding to CTLA-4 significantly, suggesting that SAP and SHP-2 compete for the common binding site, GVYVKM. Thus, by blocking SHP-2 recruitment SAP could function as a negative regulator of CTLA-4. Conclusion: Taken together, our data suggest the existence of complicate signaling cascade in regulating CTLA-4 function, and further provide evidence that SAP can act either as a positive or negative regulator depending on the nature of the associating receptors.

• The Korean Journal of Physiology and Pharmacology
• /
• v.27 no.4
• /
• pp.417-426
• /
• 2023

The TRPM4 gene encodes a Ca²⁺-activated monovalent cation channel called transient receptor potential melastatin 4 (TRPM4) that is expressed in various tissues. Dysregulation or abnormal expression of TRPM4 has been linked to a range of diseases. We introduced the hemagglutinin (HA) tag into the extracellular S6 loop of TRPM4, resulting in an HA-tagged version called TRPM4-HA. This TRPM4-HA was developed to investigate the purification, localization, and function of TRPM4 in different physiological and pathological conditions. TRPM4-HA was successfully expressed in the intact cell membrane and exhibited similar electrophysiological properties, such as the current-voltage relationship, rapid desensitization, and current size, compared to the wild-type TRPM4. The presence of the TRPM4 inhibitor 9-phenanthrol did not affect these properties. Furthermore, a wound-healing assay showed that TRPM4-HA induced cell proliferation and migration, similar to the native TRPM4. Co-expression of protein tyrosine phosphatase, non-receptor type 6 (PTPN6 or SHP1) with TRPM4-HA led to the translocation of TRPM4-HA to the cytosol. To investigate the interaction between PTPN6 and tyrosine residues of TRPM4 in enhancing channel activity, we generated four mutants in which tyrosine (Y) residues were substituted with phenylalanine (F) at the N-terminus of TRPM4. The YF mutants displayed properties and functions similar to TRPM4-HA, except for the Y256F mutant, which showed resistance to 9-phenanthrol, suggesting that Y256 may be involved in the binding site for 9-phenanthrol. Overall, the creation of HA-tagged TRPM4 provides researchers with a valuable tool to study the role of TRPM4 in different conditions and its potential interactions with other proteins, such as PTPN6.