Expression of inducible nitric oxide synthase (iNOS) is central to many of the systemic effects associated with sepsis. iNOS expression and nitric oxide (NO) production alter multiple functions, including cardiac contractility, vasomotor tone, intestinal epithelial permeability, and leukocyte recruitment. Utilizing both in vivo and in vitro murine models of endotoxemia (LPS), we have previously demonstrated that NO feedback inhibits its own synthesis by increasing transcription of osteopontin (OPN), a potent trans-repressor of iNOS expression. In this competitive renewal, we propose to characterize the pathway by which OPN feeds back to downregulate iNOS transcription. In in vivo, ex vivo, and in vitro murine models of LPS- and cecal ligation and puncture (CLP) mediated sepsis, our studies show that: 1) OPN acts with STAT-interacting LIM (SLIM) protein to ubiquitinate (Ub) an essential iNOS transcription factor, STAT1, for 26s proteasome mediated degradation and inhibit STAT1 dependent iNOS expression, 2) STAT1 is not ubiquitinated in the absence of SLIM, and 3) survival of OPN null or SLIM null mice is significantly decreased indicating the functional relevance of this pathway in the pathophysiology of sepsis. We hypothesize that OPN acts through SLIM as an E3 ubiquitin ligase to degrade STAT1 protein and inhibit iNOS transcription in sepsis. We will focus on the following specific aims which are critical to defining the mechanisms underlying OPN mediated STAT1 degradation in murine models of LPS and CLP mediated sepsis. 1) We will identify the OPN-regulated E3 ligase which transfers ubiquitin to STAT1, focusing initially on SLIM protein. 2) We will define the role of OPN in regulating expression and/or activation of SLIM. 3) We will confirm in vivo relevance of the OPN-STAT1-Ub pathway in murine models of LPS stimulation and/or CLP that incorporate OPN null and SLIM null animals. The role of OPN in the regulation of STAT1 dependent protein expression in sepsis has not been previously explored. Our proposed studies will utilize iNOS as a specific example of a STAT1 dependent protein to define OPN as a unique and as yet, poorly characterized, trans-activator of STAT1 degradation. Characterization of this regulatory pathway may identify potential regulatory targets for therapy in septic shock.
Death as the result of severe infection and injury remains a critical problem in the U.S. This research examines the regulation of a major player in the breakdown of normal bodily functions following major infection and injury. If successful, our studies may result in prevention of this breakdown.
|Weber, C E; Kothari, A N; Wai, P Y et al. (2015) Osteopontin mediates an MZF1-TGF-Î²1-dependent transformation of mesenchymal stem cells into cancer-associated fibroblasts in breast cancer. Oncogene 34:4821-33|
|Seth, Devanshi; Duly, Alastair; Kuo, Paul C et al. (2014) Osteopontin is an important mediator of alcoholic liver disease via hepatic stellate cell activation. World J Gastroenterol 20:13088-104|
|Hunter, Cedric; Bond, Jennifer; Kuo, Paul C et al. (2012) The role of osteopontin and osteopontin aptamer (OPN-R3) in fibroblast activity. J Surg Res 176:348-58|
|Bhattacharya, Syamal D; Mi, Zhiyong; Talbot, Lindsay J et al. (2012) Human mesenchymal stem cell and epithelial hepatic carcinoma cell lines in admixture: concurrent stimulation of cancer-associated fibroblasts and epithelial-to-mesenchymal transition markers. Surgery 152:449-54|
|Bhattacharya, Syamal D; Mi, Zhiyong; Kim, Victoria M et al. (2012) Osteopontin regulates epithelial mesenchymal transition-associated growth of hepatocellular cancer in a mouse xenograft model. Ann Surg 255:319-25|
|Talbot, Lindsay Jones; Mi, Zhiyong; Bhattacharya, Syamal Dave et al. (2011) Pharmacokinetic characterization of an RNA aptamer against osteopontin and demonstration of in vivo efficacy in reversing growth of human breast cancer cells. Surgery 150:224-30|
|Diesen, Diana L; Kuo, Paul C (2011) Nitric oxide and redox regulation in the liver: part II. Redox biology in pathologic hepatocytes and implications for intervention. J Surg Res 167:96-112|
|Mi, Zhiyong; Bhattacharya, Syamal D; Kim, Victoria M et al. (2011) Osteopontin promotes CCL5-mesenchymal stromal cell-mediated breast cancer metastasis. Carcinogenesis 32:477-87|
|Syn, Wing-Kin; Choi, Steve S; Liaskou, Evaggelia et al. (2011) Osteopontin is induced by hedgehog pathway activation and promotes fibrosis progression in nonalcoholic steatohepatitis. Hepatology 53:106-15|
|Zhao, Wei; Wang, Lijuan; Zhang, Meng et al. (2011) NF-*B- and AP-1-mediated DNA looping regulates osteopontin transcription in endotoxin-stimulated murine macrophages. J Immunol 186:3173-9|
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