Regulation of epigenetics and development by the xenobiotic/oxidative stress response factor Keap1 in Drosophila

Abstract

Environmental toxins can cause diseases or developmental defects when not removed from cells, but the molecular mechanisms are not fully understood. The Keap1-Nrf2 complex is an important regulator of detoxification. Nrf2 is a transcription factor that activates expression of detoxification genes. Keap1 usually binds to Nrf2 in the cytoplasm and targets it for degradation. Toxins disrupt the Keap1-Nrf2 interaction, allowing Nrf2 to enter the nucleus and activate transcription. Interestingly, Keap1 and Nrf2 also bind to chromatin and regulate developmental genes. Thus, perhaps the developmental roles of Keap1-Nrf2 mediate the effects of toxins on development. To address this overall goal, this research used Drosophila to elucidate the developmental functions and molecular mechanisms of dKeap1 (Drosophila Keap1).Studying the developmental functions of dKeap1 is complicated because disrupting dKeap1 can cause mis-regulation of toxin response and lead to developmental defects. We found that a dKeap1 truncation with the C-terminal domain removed (dKeap1-ΔCTD) could still regulate expression of detoxifying genes but could no longer enter the nucleus to bind to chromatin. This allowed us to distinguish the developmental defects directly related to dKeap1 from the indirect effects of mis-regulated toxin response. We found that dKeap1-ΔCTD flies had defects in viability, fertility, and adipose tissue. These phenotypes likely resulted from mis-regulated developmental gene expression as we found that ecdysone biosynthetic/response genes and adipogenesis genes were down-regulated in specific tissues. We hypothesized that dKeap1 regulates transcription through controlling high-order chromatin structure. Here, we found that dKeap1 interacts with Lamin Dm0 proteins, which are intermediate filament proteins that form the nuclear lamina and organize the overall chromatin architecture. We also found that dKeap1 is required for the maintenance of a normal nuclear lamina, that dKeap1 overexpression redistributes the heterochromatin marker H3K9me2 along the chromosome arms and that dKeap1 and Lamin Dm0 function in the same genetic pathway. These results support a model where dKeap1 regulates chromatin structure and gene expression through interaction with Lamin proteins. Understanding the roles of dKeap1 in chromatin regulation and development can lead to a better understanding of the roles of Keap1-Nrf2 in disease and how environmental toxins influence epigenetics and development at the molecular level.