ANP32E, a Protein Involved in Steroid-Refractoriness in Ulcerative Colitis, Identified by a Systems Biology Approach
JOURNAL OF CROHNS & COLITIS
Authors: Loren, V.; Garcia-Jaraquemada, A.; Naves, J. E.; Carmona, X.; Manosa, M.; Aransay, A. M.; Lavin, J. L.; Sanchez, I.; Cabre, E.; Manye, J.; Domenech, E.
Abstract
Background and Aims Steroid-refractoriness is a common and unpredictable phenomenon in ulcerative colitis [UC], but there are no conclusive studies on the molecular functions involved. We aimed to assess the mechanism of action related to steroid failure by integrating transcriptomic data from UC patients, and updated molecular data on UC and glucocorticoids. Methods MicroRNA [miRNA] and mRNA expression were evaluated by sequencing and microarrays, respectively, from rectal biopsies of patients with moderately-to-severe active UC, obtained before and on the third day of steroid treatment. The differential results were integrated into the mathematical models generated by a systems biology approach. Results This computational approach identified 18 proteins that stand out either by being associated with the mechanism of action or by providing a means to classify the patients according to steroid response. Their biological functions have been linked to inflammation, glucocorticoid-induced transcription and angiogenesis. All the selected proteins except ANP32E [a chaperone which has been linked to the exchange of H2A.z histone and promotes glucocorticoid receptor-induced transcription] had previously been related to UC and/or glucocorticoid-induced biological actions. Western blot and immunofluorescence assays confirmed the implication of this chaperone in steroid failure in patients with active UC. Conclusions A systems biology approach allowed us to identify a comprehensive mechanism of action of steroid-refractoriness, highlighting the key role of steroid-induced transcription and the potential implication of ANP32E in this phenomenon.
Livin Regulates H2A.X-Y142 Phosphorylation and Promotes Autophagy in Colon Cancer Cells via a Novel Kinase Activity
FRONTIERS IN ONCOLOGY
Authors: Ge, Yang; Liu, Bao-lin; Cui, Jun-peng; Li, Shu-qiang
Abstract
Objective: To investigate Livin-mediated regulation of H2A.X-Y142 phosphorylation via a novel kinase activity and its effect on autophagy in colon cancer cells. Methods: The interaction between Livin and H2A.X was tested by immunoprecipitation. H2A.X-/- HCT116 cells were transfected with human influenza hemagglutinin (HA)-tagged WT or Y142F phospho-dead mutantH2A.X plasmids. GST-tagged recombinant Livin protein was used to perform in vitro pull-down experiment and kinase assay. H2A.X-/-Livin+/+ SW480 cells were co-transfected with H2A.X-WT/H2A.X-Y142F plasmid and LC3 EGFP-tagged plasmid to explore whether H2A.X-Y142F was involved in Livin-mediated autophagy induced by starvation in colon cancer cells. Results: Co-immunoprecipitation studies confirmed that Livin interacted with H2A.X and that it was phosphorylation dependent. In vitro kinase assay confirmed that Livin could phosphorylate H2A.X. Knockdown of Livin (Livin-/-) in SW480 cells or HCT116 cells canceled the starvation-induced autophagy in colon cancer cells; H2A.X-/-Livin+/+ SW480 cells transfected with H2A.X-WT activated autophagy induced by starvation while cells transfected with H2A.X-Y142F had no significant difference; Livin-H2A.X-Y142F axis activated autophagy in colon cancer cells through transcriptionally regulating ATG5 and ATG7. Conclusion: Livin promotes autophagy in colon cancer cells via regulating the phosphorylation of H2A.X-Y142.
COA
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