Sumoylation-deficient Prdx6 repairs aberrant Sumoylation-mediated Sp1 dysregulation-dependent Prdx6 repression and cell injury in aging and oxidative stress.

Bhavana Chhunchha,Eri Kubo,Prerna Singh,Dhirendra P. Singh

doi:10.18632/aging.101547

Abstract

Progressive deterioration of antioxidant response in aging is a major culprit in the initiation of age-related pathobiology induced by oxidative stress. We previously reported that oxidative stress leads to a marked reduction in transcription factor Sp1 and its mediated Prdx6 expression in lens epithelial cells (LECs) leading to cell death. Herein, we examined how Sp1 activity goes awry during oxidative stress/aging, and whether it is remediable. We found that Sp1 is hyper-Sumoylated at lysine (K) 16 residue in aging LECs. DNA binding and promoter assays revealed, in aging and oxidative stress, a significant reduction in Sp1 overall binding, and specifically to Prdx6 promoter. Expression/overexpression assay revealed that the observed reduction in Sp1-DNA binding activity was connected to its hyper-Sumoylation due to increased reactive oxygen species (ROS) and Sumo1 levels, and reduced levels of Senp1, Prdx6 and Sp1. Mutagenesis of Sp1 at K16R (arginine) residue restored steady-state, and improved Sp1-DNA binding activity and transactivation potential. Extrinsic expression of Sp1K16R increased cell survival and reduced ROS levels by upregulating Prdx6 expression in LECs under aging/oxidative stress, demonstrating that Sp1K16R escapes the aberrant Sumoylation processes. Intriguingly, the deleterious processes are reversible by the delivery of Sumoylation-deficient Prdx6, an antioxidant, which would be a candidate molecule to restrict aging pathobiology.

Highlights

All cells or organisms encounter many types of environmental stresses and apoptotic stimuli
Age-related increases of reactive oxygen species (ROS) levels in lens epithelial cells (LECs) were connected to progressive decline in Sp1 and Prdx6 expression and Sp1-DNA binding activity to its GC rich elements
We observed that the levels of both Sp1 and Prdx6 mRNA in human lens epithelial cells (hLECs) declined with aging, and this loss was more significant in aged cells (Fig. 1B, 56y onward)

Summary

Introduction

All cells or organisms encounter many types of environmental stresses and apoptotic stimuli. Many biologically relevant factors in the cellular and external environments, such as chemical, ultraviolet B (UVB) radiation, H2O2 and growth factors, have been shown to initiate reactive oxygen species (ROS)-evoked adverse signaling in cells due to malfunction of antioxidant defense system. We and other investigators have reported that Prdx maintains cellular homeostasis by maintaining calcium homeostasis and cell membrane and DNA integrity by optimizing ROS levels against the stressors such as H2O2, UVB, paraquat, endoplasmic reticulum (ER) stress and overstimulated growth factors [7,8,9]. Our published report has shown that expression of the antioxidant defense gene Prdx and its regulator Sp1 decline significantly in an age-dependent www.aging‐us.com manner, which is directly associated with increased production of ROS and cell death in human lens epithelial cells (hLECs) [10, 11]

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