doiSerbia

Contribution of O-GlcNAc modification of NF-κB p65 in the attenuation of diabetes-induced haptoglobin expression in rat liver

Martinović Vesna I. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Arambašić-Jovanović Jelena D. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Bogojević Desanka B. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Ivanović Anđelija S. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Otašević Vesna M. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Stančić Ana T. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia)
Grigorov Ilijana I. (Department of Molecular Biology, Institute for Biological Research “Siniša Stanković”, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia), iligri@ibiss.bg.ac.rs

Haptoglobin (Hp) is a hemoglobin-binding protein that prevents free hemoglobin-induced tissue oxidative damage. In streptozotocin-induced diabetic rats, the initial elevation of Hp expression in the serum and liver tends to decrease with diabetes progression, contributing to increased oxidative stress. Glucose toxicity and diabetic complications are closely related to increased modification of nucleocytoplasmic proteins by O-linked-N-acetylglucosamine (O-GlcNAc). We examined the contribution of O-GlcNAcylation of NF-κB p65 to changes in liver Hp expression in diabetic rats. WGA-affinity chromatography revealed a progressive increase in O-GlcNAcylation in nuclear NF-κB p65 during eight weeks of diabetes. DNA-affinity chromatography followed by immunoblot analysis revealed that decreased Hp expression at 4 and 8 weeks of diabetes was accompanied by the absence of Hp gene hormone-responsive element (HRE) occupancy with NF-κB p65, low occupancy with glucocorticoid receptor (GR), and almost no changes in STAT3 occupancy compared to 2 weeks, when Hp expression was highest. Coimmunoprecipitation experiments indicate that these events were the result of impaired NF-κB p65/STAT3 and GR/STAT3 interactions. Results suggest that the attenuation of Hp expression associated with diabetes was at least in part the result of O-GlcNAcylation of NF-κB p65, which prevents the formation of an effective transcription initiation complex on the Hp gene promoter.

Keywords: haptoglobin, gene expression, diabetes, NF-kB p65, O-linked-N-acetylglucosamine modification

Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 451-03-68/2020-14/200007