The GET pathway modulates azole susceptibility through sterol metabolic remodeling and protein trafficking in filamentous fungi
4 affiliations
- 1State Key Laboratory of Microbial Diversity and Innovative Utilization, Institute of Microbiology, Chinese Academy of Sciences
- 2College of Life Sciences, University of Chinese Academy of Sciences
- 3National Institute for Radiological Protection, China CDC
- 4Shandong Jinniu Group Co., Ltd.
Abstract
Azole antifungals are widely used to control fungal infection, yet resistance mechanisms beyond drug efflux and target modification remain insufficiently understood. Here, we identified the Guided Entry of Tail-anchored proteins (GET) pathway as a conserved regulator of azole susceptibility in Neurospora crassa and Aspergillus fumigatus . Deletion or dysfunction of the core GET components Get-3 or Get-4 confers resistance to multiple azoles without impairing hyphal growth or sporulation. Mechanistically, GET deficiency neither reduced intracellular accumulation nor altered drug efflux and target expression or total ergosterol levels. Instead, GET-deficient strains displayed significantly reduced accumulation of the toxic sterol intermediate 14α-methyl-3,6-diol under azole stress, a change that may limit azole-induced sterol toxicity and contribute to reduced azole susceptibility. Proximity labeling and functional validation indicated that GET deficiency perturbs the trafficking of multiple azole-resistance-associated proteins, including the key tail-anchored protein transporter Emp-47 and mitochondrial Cox subunits Cox-4 and Cox-15, which deletion enhanced azole resistance. Transcriptomic and functional analyses further revealed that GET disruption altered the expression of some azole resistance-associated genes, which involve in membrane transport, metabolism, and cell wall organization, independent of canonical stress pathways. In this study, 19 previously uncharacterized genes modulating azole susceptibility are uncovered. Collectively, our findings identify the GET pathway as a non-classical regulator of azole susceptibility associated with protein-trafficking defects and metabolic adaptation in filamentous fungi.
IMPORTANCE: Understanding non-canonical mechanisms of azole resistance is crucial given the rising failure of antifungal therapies. Here, we identify the GET pathway as an evolutionarily conserved regulator of azole susceptibility in both a model fungus and the major pathogen Aspergillus fumigatus . We uncover 19 previously unknown resistance-modulating genes and reveal a conceptually distinct mechanism whereby GET disruption confers resistance independently of classical efflux pumps or target alterations. Instead, GET dysfunction impairs trafficking of Emp-47 and Cox subunits, reduces toxic sterol intermediate accumulation, and alters resistance-associated gene expression. These findings establish protein trafficking as a new layer of stress adaptation and provide a mechanistic framework for further investigation of fungal azole susceptibility.
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BibTeX
@article{ye2026pathway,
title = {{The GET pathway modulates azole susceptibility through sterol metabolic remodeling and protein trafficking in filamentous fungi}},
author = {Shuting Ye and Ziyu Cui and Mi Zhou and Chengcheng Hu and Pengju Yu and Long Zhang and Shaojie Li and Xianyun Sun},
journal = {mBio},
year = {2026},
month = oct,
doi = {10.1128/mbio.02021-26},
url = {https://journals.asm.org/doi/10.1128/mbio.02021-26},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - The GET pathway modulates azole susceptibility through sterol metabolic remodeling and protein trafficking in filamentous fungi
AU - Shuting Ye
AU - Ziyu Cui
AU - Mi Zhou
AU - Chengcheng Hu
AU - Pengju Yu
AU - Long Zhang
AU - Shaojie Li
AU - Xianyun Sun
PY - 2026
DA - 2026/10/07
JO - mBio
DO - 10.1128/mbio.02021-26
UR - https://journals.asm.org/doi/10.1128/mbio.02021-26
AB - Azole antifungals are widely used to control fungal infection, yet resistance mechanisms beyond drug efflux and target modification remain insufficiently understood. Here, we identified the Guided Entry of Tail-anchored proteins (GET) pathway as a conserved regulator of azole susceptibility in Neurospora crassa and Aspergillus fumigatus . Deletion or dysfunction of the core GET components Get-3 or Get-4 confers resistance to multiple azoles without impairing hyphal growth or sporulation. Mechanistically, GET deficiency neither reduced intracellular accumulation nor altered drug efflux and target expression or total ergosterol levels. Instead, GET-deficient strains displayed significantly reduced accumulation of the toxic sterol intermediate 14α-methyl-3,6-diol under azole stress, a change that may limit azole-induced sterol toxicity and contribute to reduced azole susceptibility. Proximity labeling and functional validation indicated that GET deficiency perturbs the trafficking of multiple azole-resistance-associated proteins, including the key tail-anchored protein transporter Emp-47 and mitochondrial Cox subunits Cox-4 and Cox-15, which deletion enhanced azole resistance. Transcriptomic and functional analyses further revealed that GET disruption altered the expression of some azole resistance-associated genes, which involve in membrane transport, metabolism, and cell wall organization, independent of canonical stress pathways. In this study, 19 previously uncharacterized genes modulating azole susceptibility are uncovered. Collectively, our findings identify the GET pathway as a non-classical regulator of azole susceptibility associated with protein-trafficking defects and metabolic adaptation in filamentous fungi. IMPORTANCE: Understanding non-canonical mechanisms of azole resistance is crucial given the rising failure of antifungal therapies. Here, we identify the GET pathway as an evolutionarily conserved regulator of azole susceptibility in both a model fungus and the major pathogen Aspergillus fumigatus . We uncover 19 previously unknown resistance-modulating genes and reveal a conceptually distinct mechanism whereby GET disruption confers resistance independently of classical efflux pumps or target alterations. Instead, GET dysfunction impairs trafficking of Emp-47 and Cox subunits, reduces toxic sterol intermediate accumulation, and alters resistance-associated gene expression. These findings establish protein trafficking as a new layer of stress adaptation and provide a mechanistic framework for further investigation of fungal azole susceptibility.
SN - 2150-7511
ER -