Artificial microRNAs against hypoxia-responsive small protein encoding genes reveal HRSP41 and 42 as regulators of development and stress tolerance
- 1Department of Development and Hormone Action, Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas – Universidad Politécnica de Valencia), Valencia, 46022 Spain
Abstract
Background: Small proteins remain poorly characterized in Arabidopsis thaliana, partly because of genetic redundancy and the difficulty of identifying loss-of-function mutants. To address this gap, we investigated 58 hypoxia-responsive small protein genes (HRSPs) encoding proteins of 80–100 amino acids using a homology-based artificial microRNA (amiRNA) strategy.
Results: Homozygous amiRNA lines were screened for developmental traits and stress-related phenotypes. Two closely related paralogs, HRSP41 and HRSP42, emerged as key regulators with both shared and distinct functions. HRSP41 promoted flowering under short-day conditions, whereas HRSP42 delayed flowering and specifically increased seed tolerance to oxidative stress. Both genes acted together to reduce the inhibitory effect of abscisic acid on seed germination. Transcriptome profiling revealed distinct downstream effects: HRSP42 silencing was associated with repression of glutathione metabolism, whereas HRSP41 silencing affected glycerolipid metabolism and cell wall-related pathways. Under hypoxia, simultaneous downregulation of HRSP41 and HRSP42 altered transcriptional, post-transcriptional, and translational responses and intensified the repression of photosynthesis-related genes.
Conclusions: Despite their high sequence similarity, HRSP41 and HRSP42 control plant development and stress adaptation through partially overlapping but clearly distinct molecular mechanisms. Our results identify these paralogs as previously uncharacterized regulators of flowering, oxidative stress responses, ABA sensitivity, and hypoxia-responsive gene expression.
Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1186/s12870-026-09666-5.
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BibTeX
@article{castillo2026artificial,
title = {{Artificial microRNAs against hypoxia-responsive small protein encoding genes reveal HRSP41 and 42 as regulators of development and stress tolerance}},
author = {Mari-Cruz Castillo and Sandra Sánchez-Palomares and José León},
journal = {BMC Plant Biology},
year = {2026},
month = aug,
doi = {10.1186/s12870-026-09666-5},
url = {https://doi.org/10.1186/s12870-026-09666-5},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Artificial microRNAs against hypoxia-responsive small protein encoding genes reveal HRSP41 and 42 as regulators of development and stress tolerance
AU - Mari-Cruz Castillo
AU - Sandra Sánchez-Palomares
AU - José León
PY - 2026
DA - 2026/08/01
JO - BMC Plant Biology
DO - 10.1186/s12870-026-09666-5
UR - https://doi.org/10.1186/s12870-026-09666-5
AB - Background: Small proteins remain poorly characterized in Arabidopsis thaliana, partly because of genetic redundancy and the difficulty of identifying loss-of-function mutants. To address this gap, we investigated 58 hypoxia-responsive small protein genes (HRSPs) encoding proteins of 80–100 amino acids using a homology-based artificial microRNA (amiRNA) strategy. Results: Homozygous amiRNA lines were screened for developmental traits and stress-related phenotypes. Two closely related paralogs, HRSP41 and HRSP42, emerged as key regulators with both shared and distinct functions. HRSP41 promoted flowering under short-day conditions, whereas HRSP42 delayed flowering and specifically increased seed tolerance to oxidative stress. Both genes acted together to reduce the inhibitory effect of abscisic acid on seed germination. Transcriptome profiling revealed distinct downstream effects: HRSP42 silencing was associated with repression of glutathione metabolism, whereas HRSP41 silencing affected glycerolipid metabolism and cell wall-related pathways. Under hypoxia, simultaneous downregulation of HRSP41 and HRSP42 altered transcriptional, post-transcriptional, and translational responses and intensified the repression of photosynthesis-related genes. Conclusions: Despite their high sequence similarity, HRSP41 and HRSP42 control plant development and stress adaptation through partially overlapping but clearly distinct molecular mechanisms. Our results identify these paralogs as previously uncharacterized regulators of flowering, oxidative stress responses, ABA sensitivity, and hypoxia-responsive gene expression. Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1186/s12870-026-09666-5.
SN - 1471-2229
ER -