The miR319/bHLH094 Module Regulates Creeping Bentgrass Thermotolerance by Modulating Auxin Biosynthesis and...

Area of Science:

• Plant Science
• Molecular Biology
• Stress Physiology

Background:

• MicroRNA319 (miR319) is known to regulate plant development and stress responses like drought and salt.
• The specific role of miR319 in thermotolerance, especially in cool-season grasses, is not well understood.

Purpose of the Study:

• To investigate the function of miR319 in heat tolerance of creeping bentgrass (Agrostis stolonifera).
• To identify the target gene of miR319 and elucidate the underlying molecular mechanisms of heat response.

Main Methods:

• Identification of miR319 target gene (AsbHLH094) using transgenic creeping bentgrass lines overexpressing miR319 (OE319).
• Functional characterization of AsbHLH094 in transgenic tobacco plants.
• Protein-protein interaction assays to study interactions with Aux/IAA proteins.
• Transcriptomic analysis to assess changes in auxin biosynthesis and signaling pathways.
• Measurement of endogenous indole-3-acetic acid (IAA) levels and assessment of exogenous IAA application effects.

Main Results:

• miR319 negatively regulates heat tolerance in creeping bentgrass.
• AsbHLH094, a bHLH transcription factor, was identified as a direct target of miR319 and its expression was downregulated in OE319 lines.
• Overexpression of AsbHLH094 enhanced heat tolerance in transgenic tobacco.
• AsbHLH094 interacts with the auxin signaling component AsIAA1.
• OE319 plants showed downregulated auxin biosynthesis and response genes, leading to reduced auxin accumulation.
• AsbHLH094 overexpression in tobacco induced auxin-related gene expression and increased auxin levels.
• High temperatures increased endogenous IAA levels in creeping bentgrass, and exogenous IAA improved heat tolerance.

Conclusions:

• A novel miR319-AsbHLH094 regulatory module is identified in creeping bentgrass.
• This module modulates auxin biosynthesis and signaling pathways to contribute to heat stress responses.
• Understanding this mechanism provides insights into improving thermotolerance in cool-season grasses.