Helicobacter pylori-induced gastric cancer is orchestrated by MRCKβ-mediated Siah2 phosphorylation
Open Access
- 3 February 2021
- journal article
- research article
- Published by Springer Science and Business Media LLC in Journal of Biomedical Science
- Vol. 28 (1), 1-17
- https://doi.org/10.1186/s12929-021-00710-0
Abstract
Background: Helicobacter pylori-mediated gastric carcinogenesis is initiated by a plethora of signaling events in the infected gastric epithelial cells (GECs). The E3 ubiquitin ligase seven in absentia homolog 2 (Siah2) is induced in GECs in response to H. pylori infection. Posttranslational modifications of Siah2 orchestrate its function as well as stability. The aim of this study was to evaluate Siah2 phosphorylation status under the influence of H. pylori infection and its impact in gastric cancer progression. Methods: H. pylori-infected various GECs, gastric tissues from H. pylori-infected GC patients and H. felis-infected C57BL/6 mice were evaluated for Siah2 phosphorylation by western blotting or immunofluorescence microscopy. Coimmunoprecipitation assay followed by mass spectrometry were performed to identify the kinases interacting with Siah2. Phosphorylation sites of Siah2 were identified by using various plasmid constructs generated by site-directed mutagenesis. Proteasome inhibitor MG132 was used to investigate proteasome degradation events. The importance of Siah2 phosphorylation on tumorigenicity of infected cells were detected by using phosphorylation-null mutant and wild type Siah2 stably-transfected cells followed by clonogenicity assay, cell proliferation assay, anchorage-independent growth and transwell invasion assay. Results: Siah2 was phosphorylated in H. pylori-infected GECs as well as in metastatic GC tissues at residues serine6 (Ser6) and threonine279 (Thr279). Phosphorylation of Siah2 was mediated by MRCKβ, a Ser/Thr protein kinase. MRCKβ was consistently expressed in uninfected GECs and noncancer gastric tissues but its level decreased in infected GECs as well as in metastatic tissues which had enhanced Siah2 expression. Infected murine gastric tissues showed similar results. MRCKβ could phosphorylate Siah2 but itself got ubiquitinated from this interaction leading to the proteasomal degradation of MRCKβ and use of proteasomal inhibitor MG132 could rescue MRCKβ from Siah2-mediated degradation. Ser6 and Thr279 phosphorylated-Siah2 was more stable and tumorigenic than its non-phosphorylated counterpart as revealed by the proliferation, invasion, migration abilities and anchorage-independent growth of stable-transfected cells. Conclusions: Increased level of Ser6 and Thr279-phosphorylated-Siah2 and downregulated MRCKβ were prominent histological characteristics of Helicobacter-infected gastric epithelium and metastatic human GC. MRCKβ-dependent Siah2 phosphorylation stabilized Siah2 which promoted anchorage-independent survival and proliferative potential of GECs. Phospho-null mutants of Siah2 (S6A and T279A) showed abated tumorigenicity.Keywords
Funding Information
- Science and Engineering Research Board (SB/SO/BB-0015/2014)
This publication has 45 references indexed in Scilit:
- Helicobacter pylori-induced gastric inflammation and gastric cancerCancer Letters, 2014
- Identification of a Src Tyrosine Kinase/SIAH2 E3 Ubiquitin Ligase Pathway That Regulates C/EBPδ Expression and Contributes to Transformation of Breast Tumor CellsMolecular and Cellular Biology, 2012
- Cdc42-dependent formation of the ZO-1/MRCKβ complex at the leading edge controls cell migrationThe EMBO Journal, 2011
- Large-Scale Prediction of Human Protein−Protein Interactions from Amino Acid Sequence Based on Latent Topic FeaturesJournal of Proteome Research, 2010
- Dissecting Activation of the PAK1 Kinase at Protrusions in Living CellsOnline Journal of Public Health Informatics, 2009
- An inducible autoregulatory loop between HIPK2 and Siah2 at the apex of the hypoxic responseNature, 2008
- A Tripartite Complex Containing MRCK Modulates Lamellar Actomyosin Retrograde FlowCell, 2008
- Fibroblast-led collective invasion of carcinoma cells with differing roles for RhoGTPases in leading and following cellsNature, 2007
- Regulation of the Ring Finger E3 Ligase Siah2 by p38 MAPKOnline Journal of Public Health Informatics, 2006
- Sequence and structure-based prediction of eukaryotic protein phosphorylation sitesJournal of Molecular Biology, 1999