Search bioRxiv⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.64898/2026.09.27.754851

The conserved function of bHLH121 and overexpression of N-terminal bHLH121 fragment suppresses iron deficiency response in Arabidopsis

Zhou, P.·Liu, C.·Pan, Y.·Fei, Y.·Shen, R.·Wang, R.·Lan, P.
View original source ↗Export metadata ↓

Abstract

Iron (Fe) homeostasis is fundamental to plant growth and development and is centrally regulated by a well-conserved network of basic helix-loop-helix (bHLH) transcription factors. In Arabidopsis thaliana, bHLH121 regulates the expression of FER-like iron deficiency-induced transcription factor (FIT) and clade Ib bHLH38/39/100/101 genes, among others. However, its functional conservation remains unexplored, and some contradictory results have been reported regarding the effects of bHLH121 overexpression. Here, we investigated the evolutionary conservation of bHLH121 through complementary tests in the Arabidopsis bhlh121 mutant created by CRISPR/Cas9 gene editing. We also investigated the effects of ectopically expressing full-length, N-terminal, and C-terminal fragments of bHLH121 on plant responses to Fe deficiency in Arabidopsis. Transgenic plants overexpressing the full-length bHLH121 driven by the cauliflower mosaic virus 35S promoter (bHLH121 ox) resulted in shorter roots and more severe leaf chlorosis compared to wild-type plants under Fe limiting conditions. Moreover, overexpression of the N-terminal fragment (N121 ox) rendered plants highly sensitive to Fe deficiency, whereas overexpression of the C terminal fragment (C121 ox) did not. Consistent with the phenotypic observations, N121 ox plants accumulated lower Fe content, and the ferric chelate reductase activity was significantly reduced. In N121 ox plants, the induction of key Fe regulatory genes, especially FIT, was significantly inhibited under Fe deficiency. Confocal imaging showed that both N121-GFP and C121-GFP fusion proteins could localize to the nucleus. Electrophoretic mobility shift assay showed that both full-length bHLH121 and N121 could effectively bind the probes derived from the promoter of FIT. The present work reveals a conserved function of bHLH121 in Fe homeostasis across several dicot species and demonstrates that the N121 may possess a dominant nature in Fe deficiency responses.

Explore related subjects

plant biologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Zhou, P., Liu, C., Pan, Y., Fei, Y., Shen, R., Wang, R., Lan, P.. 2026-09-28. The conserved function of bHLH121 and overexpression of N-terminal bHLH121 fragment suppresses iron deficiency response in Arabidopsis. https://doi.org/10.64898/2026.09.27.754851

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Specialisation of photosynthetic biochemistry in C4 chloroplasts potentiates distinct cell-specific functionality of different chloroplast-to-nucleus retrograde signalling pathways.

Chloroplasts are key environmental sensors for plant cells. Upon exposure to abiotic stresses such as excess sunlight and heat, reactive oxygen species (ROS) in chloroplasts are sensed to trigger chloroplast-to-nucleus signalling pathways for acclimatory nuclear gene expression. There is extensive biochemical- and cell type-specialisation of chloroplasts, such as those in heat-resilient plants performing C4 photosynthesis, but how these specialisations intersect with chloroplast-to-nucleus signalling remains unclear. Here we show that specific chloroplast-to-nucleus signalling networks are functional in specialised leaf cell types of the C4 model species, Setaria viridis. RNA-sequencing, electron paramagnetic resonance (EPR) spectroscopy and metabolomics on cell type-enriched fractions, complemented with mutant analyses provided evidence for cell-specific enrichment of chloroplast-to-nucleus retrograde signalling pathways correlating with the photosynthetic specialisation in these cell types. Singlet oxygen (1O2)-responsive beta-cyclocitral ({beta}-CC) /dihydroactinidiolide (DHA) signalling and hydrogen peroxide (H2O2)-responsive 3'-phosphoadenosine 5'-phosphate (PAP) signalling pathways were specific to mesophyll cells, in line with enrichment of photosystem II and linear electron flow to photosystem I, respectively in this cell type. In contrast, bundle sheath cells were enriched for the rapid, ROS-independent retrograde signalling pathway mediated by the Calvin cycle product, dihydroxyacetone phosphate (DHAP), consistent with the exclusive localisation of the Calvin cycle to C4 bundle sheath cells. Taken together, our results provide insights into how chloroplast- and cell type-specialisation intersect with stress signalling in plant cells and identify photosynthetic biochemistry as a key driver of cell-specific cellular signalling.

plant biology↗
bioRxiv2026

The periplasmic substrate-binding protein ThuE is required for sucrose uptake and multiplication of Pseudomonas syringae pv. tomato DC3000 in tomato leaves

Phytopathogenic bacteria depend on host-derived carbon for successful colonization of plant tissue. While sucrose is among the most abundant sugars in the leaf apoplast, the steps that limit its acquisition by Pseudomonas syringae are not known. In this study, we used base editing to generate mutants of six candidate sucrose utilization genes in P. syringae pv. tomato DC3000 (Pto). Loss of either the sucrose hydrolase ScrB or both levansucrases had no detectable effect, whereas the growth defect in the loss of the outer membrane porin mutant scry was overcome by raising external sucrose. Loss of the periplasmic substrate-binding protein ThuE impaired growth both in vitro and in planta, reducing bacterial populations to approximately two orders of magnitude lower in tomato leaves. Sucrose import through a single non-redundant inner membrane ABC transporter is therefore required for Pto multiplication in planta and serves as a potential target for controlling bacterial speck disease.

plant biology↗
bioRxiv2026

Spatial expression evolution of duplicated genes post-whole genome duplication in cultivated tomato

The mechanisms by which whole genome duplication contributes to expression divergence of duplicate genes, which can impact the evolution of form and function, has been a long-standing question in evolutionary biology. We compared expression patterns of paralogs derived from the most recent Solanaceae-specific genome triplication event across 23 tissues and inferred ancestral expression states to investigate patterns of expression evolution. While the partitioning of ancestral expression states is traditionally considered the second most common (after the gene loss) trajectory of duplicate expression evolution, we found that the ancestral expression states tended to be retained by most (~66%) duplicates, with ~6% of gene lineages experiencing expression gain events and ~20% experiencing expression losses. The genes that gained expression in fruit tissues are significantly enriched with functions associated with fruit quality. Notably, asymmetrical expression divergence is prevalent among duplicates, especially when ancestral genes were either not expressed or were highly expressed. Our study demonstrates the utility of integrating paralog expression comparisons with phylogenetic analyses to document the potential evolutionary history of spatial expression patterns and gain insight into how expression divergence contributes to the evolution of function across tissues.

plant biology↗