The role of Nrf1 and Nrf2 in the regulation of glutathione and redox dynamics in the developing zebrafish embryo

Abstract

Redox signaling is important for embryogenesis, guiding pathways that govern processes crucial for embryo patterning, including cell polarization, proliferation, and apoptosis. Exposure to pro-oxidants during this period can be deleterious, resulting in altered physiology, teratogenesis, later-life diseases, or lethality. We previously reported that the glutathione antioxidant defense system becomes increasingly robust, including a doubling of total glutathione and dynamic shifts in the glutathione redox potential at specific stages during embryonic development in the zebrafish, Danio rerio. However, the mechanisms underlying these changes are unclear, as is the effectiveness of the glutathione system in ameliorating oxidative insults to the embryo at different stages. Here, we examine how the glutathione system responds to the model pro-oxidants tert-butylhydroperoxide and tert-butylhydroquinone at different developmental stages, and the role of Nuclear factor erythroid 2-related factor (Nrf) proteins in regulating developmental glutathione redox status. Embryos became increasingly sensitive to pro-oxidants after 72 h post-fertilization (hpf), after which the duration of the recovery period for the glutathione redox potential was increased. To determine whether the doubling of glutathione or the dynamic changes in glutathione redox potential are mediated by zebrafish paralogs of Nrf transcription factors, morpholino oligonucleotides were used to knock down translation of Nrf1 and Nrf2 (nrf1a, nrf1b, nrf2a, nrf2b). Knockdown of Nrf1a or Nrf1b perturbed glutathione redox state until 72 hpf. Knockdown of Nrf2 paralogs also perturbed glutathione redox state but did not significantly affect the response of glutathione to pro-oxidants. Nrf1b morphants had decreased gene expression of glutathione synthesis enzymes, while hsp70 increased in Nrf2b morphants. This work demonstrates that despite having a more robust glutathione system, embryos become more sensitive to oxidative stress later in development, and that neither Nrf1 nor Nrf2 alone appear to be essential for the response and recovery of glutathione to oxidative insults.

MGST1, a GSH transferase/peroxidase essential for development and hematopoietic stem cell differentiation

“…The peak of MGST1 expression is supported by other studies in zebrafish [43] . In addition, an enhanced sensitivity to pro-oxidants has been described at 72 hpf [44] , in zebrafish, an observation that fits well with the noted MGST1 decline. With few exceptions, other GSTs do not show a developmental peak in zebrafish development [45] .…”

Mapping glutathione utilization in the developing zebrafish (Danio rerio) embryo

“…The ability of the GSH system to respond and recover from oxidizing conditions changes with developmental stage. Zebrafish embryos are increasingly resistant to oxidizing exposures from 18 h post fertilization (hpf) (when the majority of the endoderm derived organs start developing) −72 hpf (most major endoderm-derived organs have developed and the embryo hatches); after hatching, embryos become much more sensitive to pro-oxidant exposures [35]. This is in keeping with changes in the concentration of GSH in zebrafish embryos during development, which nearly doubles between 24 to 36 hpf [33].…”

Unravelling paralogous gene expression dynamics during three-spined stickleback embryogenesis

“…48 X scf. 692 jamb (−/*) I 61 jamc (*/*) VII 61 XVI I nanos1 (*/*) IV 62 nrf1 (*/−) V 63 XV XI pax2 (−/*) VI 64 ppar alpha (*/*) IV 65 V XIX tle3 (groucho3) (*/−) XIX 66 snai1 (*/−) XVII 67 II scf. 180 wnt4 (*/*) XVII 68 pou2f2 (*/−) XX 69 XX X pou3f2 (*/*) XX 69 stat1 (*/*) I 70 XVIII XVI tbx5 (*/*) XIV 71 lbx1 (*/*) IX 72 XIII VI wt1 (*/*) XIX 73 adam17 (*/−) scf.…”