Iron (Fe) deficiency is harmful to plant growth and development. Plants can sense Fe deficiency conditions and modulate the expression of Fe deficiency-responsive genes in order to maintain Fe homeostasis. POPEYE (PYE) was characterized as a negative regulator of the Fe homeostasis-associated genes. However, the mechanism by which PYE regulates Fe homeostasis remains unclear. 

In a study published in Journal of Experimental Botany, researchers from the Xishuangbanna Tropical Botanical Garden (XTBG) of the Chinese Academy of Sciences showed that the PYE negatively regulates bHLH Ib genes in two different ways and that the conditional nuclear localization of PYE is crucial for Fe homeostasis. 

To further investigate the functions of PYE in the Fe deficiency response, the researchers constructed PYE-overexpressing plants (PYEox). The results showed that PYE overexpression disrupted the Fe deficiency response. PYE negatively affected the expression of genes associated with Fe homeostasis. PYE directly repressed the transcription of bHLH Ib genes (bHLH38, bHLH39, bHLH100, and bHLH101) by associating with their promoters.  

Subcellular localization analysis indicated that PYE localized in both the cytoplasm and the nucleus. PYE contained a Nuclear Export Signal (NES) which is required for the cytoplasmic localization of PYE. Mutation of the NES enhanced the repressive function of PYE, resulting in the downregulation of Fe deficiency responsive genes.  

Co-expression assays revealed that three bHLH IVc members (bHLH104, bHLH105/ILR3, and bHLH115) facilitated the nuclear accumulation of PYE. Conversely, PYE indirectly repressed the transcriptional activating ability of bHLH IVc. Additionally, PYE directly negatively regulated its own transcription. 

"Our study provides insights into the complicated Fe deficiency response signaling pathway and enhances the understanding of PYE functions," said LIANG Gang of XTBG.