The selective post-translational processing of transcription factor Nrf1 yields distinct isoforms that dictate its ability to differentially regulate gene expression.
Upon translation, the N-terminal homology box 1 (NHB1) signal anchor sequence of Nrf1 integrates it within the endoplasmic reticulum (ER) whilst its transactivation domains [TADs, including acidic domain 1 (AD1), the flanking Asn/Ser/Thr-rich (NST) domain and AD2] are transiently translocated into the ER lumen, whereupon the NST domain is glycosylated to yield an inactive 120-kDa glycoprotein. Subsequently, these TADs are retrotranslocated into extra-luminal subcellular compartments, where Nrf1 is deglycosylated to yield an active 95-kDa isoform. Herein, we report that AD1 and AD2 are required for the stability of the 120-kDa Nrf1 glycoprotein, but not that of the non-glycosylated/de-glycosylated 95-kDa isoform. Degrons within AD1 do not promote proteolytic degradation of the 120-kDa Nrf1 glycoprotein. However, repositioning of AD2-adjoining degrons (i.e. DSGLS-containing SDS1 and PEST2 sequences) into the cyto/nucleoplasm enables selective topovectorial processing of Nrf1 by the proteasome and/or calpains to generate a cleaved active 85-kDa Nrf1 or a dominant-negative 36-kDa Nrf1γ. Production of Nrf1γ is abolished by removal of SDS1 or PEST2 degrons, whereas production of the cleaved 85-kDa Nrf1 is blocked by deletion of the ER luminal-anchoring NHB2 sequence (aa 81-106). Importantly, Nrf1 activity is positively and/or negatively regulated by distinct doses of proteasome and calpain inhibitors.
1] The NSFC-funded Laboratory of Cell Biochemistry and Gene Regulation, College of Medical Bioengineering and Faculty of Life Sciences, Chongqing University, No. 174 Shazheng Street, Shapingba District, Chongqing 400044, China [2] Jacqui Wood Cancer Centre, James Arrott Drive, Division of Cancer Research, Medical Research Institute, Ninewells Hospital &Medical School, University of Dundee, DD1 9SY, Scotland, UK.
Yiguo
Y
Zhang
The NSFC-funded Laboratory of Cell Biochemistry and Gene Regulation, College of Medical Bioengineering and Faculty of Life Sciences, Chongqing University, No. 174 Shazheng Street, Shapingba District, Chongqing 400044, China.
Shaojun
S
Li
The NSFC-funded Laboratory of Cell Biochemistry and Gene Regulation, College of Medical Bioengineering and Faculty of Life Sciences, Chongqing University, No. 174 Shazheng Street, Shapingba District, Chongqing 400044, China.
Yuancai
Y
Xiang
The NSFC-funded Laboratory of Cell Biochemistry and Gene Regulation, College of Medical Bioengineering and Faculty of Life Sciences, Chongqing University, No. 174 Shazheng Street, Shapingba District, Chongqing 400044, China.
Lu
L
Qiu
The NSFC-funded Laboratory of Cell Biochemistry and Gene Regulation, College of Medical Bioengineering and Faculty of Life Sciences, Chongqing University, No. 174 Shazheng Street, Shapingba District, Chongqing 400044, China.
Huakan
H
Zhao
Jacqui Wood Cancer Centre, James Arrott Drive, Division of Cancer Research, Medical Research Institute, Ninewells Hospital &Medical School, University of Dundee, DD1 9SY, Scotland, UK.
John D
JD
Hayes
D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D019556:COS Cells; D002154:Calpain / Q000235:genetics* / Q000378:metabolism; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000235:genetics; D005786:Gene Expression Regulation; D006801:Humans; D051266:Nuclear Respiratory Factor 1 / Q000096:biosynthesis / Q000235:genetics*; D046988:Proteasome Endopeptidase Complex / Q000235:genetics; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics*; D011499:Protein Processing, Post-Translational / Q000235:genetics*; D059748:Proteolysis; D017386:Sequence Homology, Amino Acid; D015533:Transcriptional Activation / Q000235:genetics*
C495633:NRF1 protein, human; D051266:Nuclear Respiratory Factor 1; D020033:Protein Isoforms; D002154:Calpain; D046988:Proteasome Endopeptidase Complex