The NHB1 (N-terminal homology box 1) sequence in transcription factor Nrf1 is required to anchor it to the endoplasmic reticulum and also to enable its asparagine-glycosylation

Yiguo Zhang, John M. Lucocq, Masayuki Yamamoto, John D. Hayes

Research output: Contribution to journalArticlepeer-review

66 Citations (Scopus)

Abstract

Nrf1 (nuclear factor-erythroid 2 p45 subunit-related factor 1) is negatively controlled by its NTD (N-terminal domain) that lies between amino acids 1 and 124. This domain contains a leucine-rich sequence, called NHB1 (N-terminal homology box 1; residues 11-30), which tethers Nrf1 to the ER (endoplasmic reticulum). Electrophoresis resolved Nrf1 into two major bands of approx. 95 and 120 kDa. The 120-kDa Nrf1 form represents a glycosylated protein that was present exclusively in the ER and was converted into a substantially smaller polypeptide upon digestion with either peptide:N-glycosidase F or endoglycosidase H. By contrast, the 95-kDa Nrf1 form did not appear to be glycosylated and was present primarily in the nucleus. NHB1 and its adjacent residues conform to the classic tripartite signal peptide sequence, comprising n-, h- and c-regions. The h-region (residues 11-22), but neither the n-region (residues 1-10) nor the c-region (residues 23-30), is required to direct Nrf1 to the ER. Targeting Nrf1 to the ER is necessary to generate the 120-kDa glycosylated protein. The n-region and c-region are required for correct membrane orientation of Nrf1, as deletion of residues 2-10 or 23-30 greatly increased its association with the ER and the extent to which it was glycosylated. The NHB1 does not contain a signal peptidase cleavage site, indicating that it serves as an ER anchor sequence. Wild-type Nrf1 is glycosylated through its Asn/Ser/Thr-rich domain, between amino acids 296 and 403, and this modification was not observed in an Nrf1Δ299-400 mutant. Glycosylation of Nrf1 was not necessary to retain it in the ER.

Original languageEnglish
Pages (from-to)161-172
Number of pages12
JournalBiochemical Journal
Volume408
Issue number2
DOIs
Publication statusPublished - 2007 Dec 1
Externally publishedYes

Keywords

  • Endoplasmic reticulum
  • Glycosylation
  • Non-alcoholic steatohepatitis (NASH)
  • Nuclear factor-erythroid 2-p45 subunit-related factor (Nrf)
  • Oxidative stress
  • Signal anchor sequence

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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