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Colitis Is Associated with Loss of the Histidine Phosphatase LHPP and Upregulation of Histidine Phosphorylation in Intestinal Epithelial Cells.

ABSTRACT: Protein histidine phosphorylation (pHis) is a posttranslational modification involved in cell cycle regulation, ion channel activity and phagocytosis. Using novel monoclonal antibodies to detect pHis, we previously reported that the loss of the histidine phosphatase LHPP (phospholysine phosphohistidine inorganic pyrophosphate phosphatase) results in elevated pHis levels in hepatocellular carcinoma. Here, we show that intestinal inflammation correlates with the loss of LHPP in dextran sulfate sodium (DSS)-treated mice and in inflammatory bowel disease (IBD) patients. Increased histidine phosphorylation was observed in intestinal epithelial cells (IECs), as determined by pHis immunofluorescence staining of colon samples from a colitis mouse model. However, the ablation of Lhpp did not cause increased pHis or promote intestinal inflammation under physiological conditions or after DSS treatment. Our observations suggest that increased histidine phosphorylation plays a role in colitis, but the loss of LHPP is not sufficient to increase pHis or to cause inflammation in the intestine.

SUBMITTER: Linder M

PROVIDER: S-EPMC10452693 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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Colitis Is Associated with Loss of the Histidine Phosphatase LHPP and Upregulation of Histidine Phosphorylation in Intestinal Epithelial Cells.

Linder Markus M Liko Dritan D Kancherla Venkatesh V Piscuoglio Salvatore S Hall Michael N MN

Biomedicines 20230801 8

Protein histidine phosphorylation (pHis) is a posttranslational modification involved in cell cycle regulation, ion channel activity and phagocytosis. Using novel monoclonal antibodies to detect pHis, we previously reported that the loss of the histidine phosphatase LHPP (phospholysine phosphohistidine inorganic pyrophosphate phosphatase) results in elevated pHis levels in hepatocellular carcinoma. Here, we show that intestinal inflammation correlates with the loss of LHPP in dextran sulfate sod ...[more]

PMID: 37626656

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Project description:Intestinal epithelial barrier protects intestine from infection and injury, while chronic inflammation is a trigger for tumorigenesis. As a member of tight junctions (TJs) family, Claudin-7 (Cldn-7) is dedicated to maintaining cell polarity and TJs barrier integrity, and closely related to the development of inflammation and tumors. However, potential roles of Cldn-7 in intestinal inflammation and colitis-associated colorectal cancer (CAC) have not been well characterized in vivo. Here, we analyzed the expression profile of Cldn-7 in inflammatory bowel disease (IBD) and CAC. Colitis and colitis-cancer transformation models were established based on inducible intestinal conditional Cldn-7 gene knockout mice (Cldn7fl/fl;villin-CreERT2), by intraperitoneal injection of azomethane (AOM) and dextran sodium sulfate (DSS) feeding. Cldn-7 knockout promoted susceptibility to colitis and CAC, aggravated clinical symptoms, severely damaged intestinal epithelium, increased mucosal inflammation accompanied dysregulated cell proliferation-apoptosis. Epithelial barrier integrity was destroyed, and intercellular permeability was increased. After AOM/DSS induction, tumor burden and volume were increased, characterized by enhanced proliferation and activation of Wnt/β-catenin signaling pathway. Mechanistically, Cldn-7 deficiency promoted colitis and subsequently malignant transformation by destroying TJs integrity and increasing inflammatory cascade. Overall, based on Cldn-7 knockout mouse model, we have first demonstrated the key roles of Cldn-7 in maintaining intestinal homeostasis and preventing IBD and consequent CAC. Abbreviations: AJs: adherens junctions; AOM: azomethane; Cldn-7: Claudin-7; CRC: colorectal cancer; CAC: colitis-associated colorectal cancer; CD: Crohn's disease; DSS: dextran sodium sulfate; DAI: disease activity index; EMT: epithelial-mesenchymal transition; FITC: fluorescence isothiocyanate; HB: hemoglobin; IBD: inflammatory bowel disease; IECs: intestinal epithelial cells; ISCs: intestinal stem cells; PLT: platelet; RBC: red blood cell; ROS: reactive oxygen species; TAM: tamoxifen; TJs: tight junctions; TCF/LEF: T-cell factor/lymphoid enhancer factor; UC: ulcerative colitis; WBC: white blood cell.

Project description:Self-renewal of embryonic stem cells (ESCs) is orchestrated by a vast number of genes at the transcriptional and translational levels. However, the molecular mechanisms of post-translational regulatory factors in ESC self-renewal remain unclear. Histidine phosphorylation, also known as hidden phosphorylation, cannot be detected by conventional experimental methods. A recent study defined phospholysine phosphohistidine inorganic pyrophosphate phosphatase (LHPP) as a histidine phosphatase, which regulates various biological behaviors in cells via histidine dephosphorylation. In this study, the doxycycline (DOX)-induced hLHPP-overexpressing mouse ESCs and mouse LHPP silenced mESCs were constructed. Quantitative polymerase chain reaction (qPCR), western blotting analysis, immunofluorescence, Flow cytometry, colony formation assays, alkaline phosphatase (AP) and bromodeoxyuridine (Brdu) staining were performed. We found that the histidine phosphorylation level was strikingly reduced following LHPP overexpression. Besides, the expression of Oct4 and Lefty1, indispensable genes in the process of ESCs self-renewal, was significantly down-regulated, while markers related to the differentiation were markedly elevated. Moreover, LHPP-mediated histidine dephosphorylation induced G0/G1 phase arrest in mESCs, suggesting LHPP was implicated in cell proliferation and cell cycle. Conversely, silencing of Lhpp promoted the self-renewal of mESCs and reversed the RA induced increased expression of genes associated with differentiation. Mechanistically, our findings suggested that the enzymatic active site of LHPP was the cysteine residue at position 226, not 53. LHPP-mediated histidine dephosphorylation lowered the expression levels of β-catenin and the cell cycle-related genes CDK4 and CyclinD1, while it up-regulated the cell cycle suppressor genes P21 and P27. Taken together, our findings reveal that LHPP-mediated histidine dephosphorylation plays a role in the self-renewal of ESCs. LHPP-mediated histidine dephosphorylation inhibited the self-renewal of ESCs by negatively regulating the Wnt/β-catenin pathway and downstream cell cycle-related genes, providing a new perspective and regulatory target for ESCs self-renewal.

Dataset Information

Colitis Is Associated with Loss of the Histidine Phosphatase LHPP and Upregulation of Histidine Phosphorylation in Intestinal Epithelial Cells.

Publications

Colitis Is Associated with Loss of the Histidine Phosphatase LHPP and Upregulation of Histidine Phosphorylation in Intestinal Epithelial Cells.

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