Protein kinase Cη-MARCKS axis associations with epithelial barrier disturbance and inflammatory readouts in...

Intestinal epithelial barrier disruption is a hallmark of ulcerative colitis (UC). Protein kinase Cη (PKCη) and its substrate myristoylated alanine-rich C-kinase substrate (MARCKS) have been implicated in cytoskeletal organization and barrier regulation, but their roles in UC remain incompletely defined. PKCη expression and its association with disease severity were assessed using Gene Expression Omnibus (GEO) datasets, dextran sodium sulfate (DSS)-induced colitis mouse model, and UC patient colon tissues. Adenoviral shRNA-mediated PKCη knockdown was performed in mice, with outcomes including body weight, colon length, disease activity index (DAI), histological changes, and permeability. PKCη, MARCKS, and tight junction proteins were assessed by RT-qPCR, Western blot, and immunohistochemistry. LPS-stimulated Caco-2 cells with PKCη knockdown/overexpression, with or without MARCKS knockdown, were used for validation. Protein interaction was tested by Co-immunoprecipitation (Co-IP). Cytokines (TNF-α, IL-1β, IL-6) were measured by ELISA. PKCη expression was elevated in UC mucosa and in DSS-induced colitis mice. In mice, PKCη knockdown was associated with reduced disease activity, lower intestinal permeability, decreased pro-inflammatory cytokines, and a more favorable tight-junction profile. Similar directional changes were observed in LPS-stimulated Caco-2 cells, whereas PKCη overexpression showed opposite effects. Co-immunoprecipitation suggested that PKCη and MARCKS occur in the same protein complex, and PKCη modulation was accompanied by changes in MARCKS abundance and phosphorylation. MARCKS knockdown partially attenuated the effects of PKCη overexpression. PKCη may contribute to epithelial barrier dysfunction and inflammation in settings relevant to ulcerative colitis, possibly through modulation of MARCKS expression and phosphorylation. Further studies are required to establish causality and to assess whether the PKCη-MARCKS axis has therapeutic relevance.