The Nrf3 transcription factor is a membrane-bound glycoprotein targeted to the endoplasmic reticulum through its N-terminal homology box 1 sequence

Transcription factor Nrf3 (NF-E2 p45-related factor 3) is targeted to the endoplasmic reticulum (ER). Mouse Nrf3 is subject to proteolysis, Asn glycosylation, and deglycosylation reactions. It is synthesized as a ∼96-kDa protein that is subsequently converted into isoforms of ∼90, 80, and 70 kDa. In the ER, the ∼90-kDa glycoprotein is predominant and gives rise to ∼80- and ∼70-kDa isoforms. The ∼90- and ∼80-kDa polypeptides were observed in the nuclear envelope, whereas the ∼70-kDa isoform was detected primarily in the nucleoplasm. Our experiments showed the N-terminal homology box 1 (NHB1, residues 12-31) is part of a tripartite signal peptide sequence, comprising n, h, and c regions. The h region (residues 12-23) was demonstrated to target Nrf3 to the ER and is necessary for its Asn glycosylation. The n region (residues 1-11) controlled the abundance of the ∼90-kDa glycoprotein. The c region (residues 24-39) was found to contain a signal peptidase cleavage site that is responsible for production of the ∼90-kDa mature Nrf3 glycoprotein from a ∼96-kDa precursor. We have found that Nrf3 is activated by the ER stressors tunicamycin and brefeldin A, and that NHB1 is required for this response. Amino acids between the c region and NHB2 (residues 76-100) controlled the proteolytic processing of mouse Nrf3 into cleavage products of ∼80-kDa (glycated) and ∼70-kDa (non-glycated); by contrast, human Nrf3 lacked a signal peptidase cleavage site between its c region and NHB2. Lastly, data are presented suggesting that the NHB2 sequence in mouse Nrf3 may regulate the topology of the transcription factor within the ER membrane.