Heme acts through the Bach1b/Nrf2a-MafK pathway to regulate exocrine peptidase precursor genes in porphyric zebrafish

Abstract

Using a zebrafish model of hepatoerythropoietic porphyria (HEP), we identify a previously unknown mechanism underlying heme-mediated regulation of exocrine zymogens. Zebrafish bach1b, nrf2a and mafK are all expressed in the zebrafish exocrine pancreas. Overexpression of bach1b or knockdown of nrf2a result in the downregulation of the expression of the exocrine zymogens, whereas overexpression of nrf2a or knockdown of bach1b cause their upregulation. In vitro luciferase assays demonstrate that heme activates the zymogens in a dosage-dependent manner and that the zymogen promoter activities require the integral Maf recognition element (MARE) motif. The Bach1b-MafK heterodimer represses the zymogen promoters, whereas the Nrf2a-MafK heterodimer activates them. Furthermore, chromatin immunoprecipitation (ChIP) assays show that MafK binds to the MARE sites in the 5′ regulatory regions of the zymogens. Taken together, these data indicate that heme stimulates the exchange of Bach1b for Nrf2a at MafK-occupied MARE sites and that, particularly in heme-deficient porphyria, the repressive Bach1b-MafK heterodimer dominates, which can be exchanged for the activating Nrf2a-MafK heterodimer upon treatment with hemin. These results provide novel insights into the regulation of exocrine function, as well as the pathogenesis of porphyria, and should be useful for designing new therapies for both types of disease.

The Transcription Factor Bach1 Suppresses the Developmental Angiogenesis of Zebrafish

“…Bach1b contains both BTB and BZIP domains and mediates the regulatory role of heme in transcription of the zymogens in zebrafish [17, 18]. Recent study has identified that Bach1b is expressed ubiquitously from one cell to 48 hpf [18], and this observation is consistent with our observation.…”

“…Bach1b plays an important role in Nrf2-MafK pathway in zebrafish [17]. Thus, we overexpressed Bach1b in zebrafish embryos to investigate the role of Bach1 during vascular development.…”

“…In the zebrafish genome, there are two Bach1 homolog genes, bach1a and Bach1b, which are located on chromosomes 15 and 10, respectively [17]. Bach1b contains both BTB and BZIP domains and mediates the regulatory role of heme in transcription of the zymogens in zebrafish [17, 18].…”

Nrf2 and Nrf2-related proteins in development and developmental toxicity: Insights from studies in zebrafish (Danio rerio)

“…Similar to the findings with nrf1 and nrf2 , zebrafish also possess duplicated keap1 genes ( keap1a and keap1b [227, 229]) and duplicated bach1 genes ( bach1a, bach1b [250, 251]). Together, these results demonstrate that duplicated zebrafish genes are common in the Nrf pathways, as they are in some other pathways involved in regulation of response to environmental stressors [252, 253].…”

“…The expression of nrf2a mRNA at 24 hpf is restricted to the olfactory system, but by 5-dpf this gene is more widely expressed, appearing also in gill, liver, and intestine [266]. Expression of nrf2a mRNA is also seen in the exocrine pancreas at 84 hpf [250]. The expression of nrf3 mRNA is localized to fore, mid- and hindbrain and pectoral fin buds between 24- and 36-hpf [240].…”