Molecular And Cellular Mechanisms Whereby The Nrf1 Transcription Factor Determines Redox Homeostasis And Protein Quality Control

Intracellular reactive oxygen species(ROS)are a group of small molecules with high oxidative activity,arising from the process of cell metabolism.ROS are not only the by-products of metabolism but also serve as indispensable signal molecules in preserving the healthy life processes,but the excessive ROS are viewed as an important culprit,contributing to cancer and other chronic diseases.In all living organisms,the redox homeostasis must be steadily maintained by controlling the production and elimination of intracellular ROS.Conversely,certain extents of the redox imbalances are considered to be closely associated with a variety of chronic diseases,including neurodegenerative diseases,cardiovascular diseases,inflammatory and autoimmune diseases,diabetes,aging,and cancer.Nrf1 and Nrf2,which belong to the CNC-b ZIP family,are two principal regulators of intracellular redox homeostasis by coordinately governing transcriptional expression of distinct subsets of cognate genes through their functional heterodimers with a partner of small Maf(s Maf)or other b ZIP proteins(e.g.,AP-1 and ATF4),that are recruited for directly binding to those consensus sequences,called antioxidant and/or electrophile response elements(AREs/Ep REs),within their target promoter regions.In addition,Nrf1 and Nrf2 are also involved in many other cellular processes,including antioxidant,inflammation,cell proliferation,protein homeostasis,senescence,DNA damage repair and cell metabolism.Nrf2 is known to play a crucial role in the maintenance of cellular redox homeostasis by regulating the biosynthesis,utilization,and regeneration of glutathione,thioredoxin,and NADPH.Nrf1 is also indispensable for the above processes,however,the function and mechanism of Nrf1 are still poorly understood.Meanwhile,the coordination between Nrf1 and Nrf2 in the maintenance of redox homeostasis remains elusive,to date.In this study,the following results have been obtained by exploring the biochemistry and biological function of Nrf1 in maintaining redox and protein homeostasis:(1)Knockout of Nrf1 or Nrf2 in Hep G2 cells can change the cellular redox homeostasis and increase the intracellular ROS levels.Knockout of Nrf2 will down-regulate the antioxidant genes,and thus elevating intracellular ROS.In Nrf1-deficient cells accumulated Nrf2 leads to an evident enhancement of antioxidant,detoxifying and cytoprotective systems,but increased Nrf2 failed to lower intracellular ROS.Compared with Nrf2–∕– cells,knockout of Nrf1 resulted in higher ROS level and oxidative damage.(2)Loss of Nrf1 causes an impairment of mitochondrial functioning,down-regulation of mitochondrial complex subunits,reduction of the number of mitochondria and crista in mitochondria,and down-regulation of related genes controlling mitochondrial division.(3)The expressions of PGC1α,αpalNRF1,GABPα,TFAM that regulate cell respiration and mitochondrial biogenesis were down-regulated in Nrf1α–∕– cells.Further studies found that Nrf1 alone or in combination with Nrf2 directly regulates αPalNRF1,GABPα and PGC1α for determining the mitochondrial homeostasis.(4)Lack of Nrf1 increased cellular antioxidant ability,but the raised glycolysis level by increased Nrf2 can increase ATP production capacity and pressure on the damaged mitochondria,thus increasing the generation of ROS.Glycolytic inhibition led to distinct extents of decreased ROS yield in Nrf1α–∕– cells.(5)The increased Nrf2 protein in Nrf1α–∕– cells inhibited the expression of UCP2 through mi R-195 and mi R-497,thereby increasing the production of ROS.Restoring the expression of UCP2 could reduce the production of ROS.(6)Deficiency of Nrf1 inhibited the repair of mitochondrial function by mitochondrial stress.And the drastically increased ROS level in Nrf1α–∕– cells could activate cancer-related pathways,such as PI3K/AKT,MAPK,HIF1α,NF-κB,AP-1,and inhibit the tumor suppressor genes P53 and PTEN.(7)The deubiquitinating enzyme USP19 can rescue Nrf1 from ER-associated degradation by removing ubiquitin molecules within Nrf1,and thus prolong its half-life to increase the transcriptional activity of this CNC-b ZIP factor.(8)USP19 can regulate proteasome subunit expression by deubiquitinating Nrf1,thereby affecting protein homeostasis in cells.In turn,Nrf1 and Nrf2 can transcriptionally activate the expression of USP19.In summary,Nrf1 acts as a potent integrator of redox regulation by multi-hierarchical network mechanisms,and Nrf1 plays a important role in linking ER with mitochondria.