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<p>Sucrose non-fermenting 1-related protein kinase (SnRK1) is a conserved central regulator of energy signaling and stress responses in plants. However, the functional roles of the <i>SnRK1β</i> regulatory subunit in turfgrass species remain largely unexplored. In this study, we identified and characterized <i>PpOSK4</i>, an <i>SnRK1β</i> homologue in Kentucky bluegrass (<i>Poa pratensis</i> L.), through integrative transcriptomic, molecular, and transgenic approaches. Mining of drought-responsive transcriptome data (PRJNA517968; three biological replicates per treatment; DESeq2, <i>q</i> &lt; 0.05, |log₂FC| &gt; 1) identified 38 differentially expressed SnRK family genes, among which <i>PpOSK4</i> was selected for functional validation based on its significant drought responsiveness and high sequence conservation with rice <i>OSK4</i> (95.4% nucleotide identity). <i>PpOSK4</i> was preferentially expressed in roots and the cultivar ‘Midnight II’, induced by salt stress, high nitrogen, and exogenous hormones (GABA, IAA, SA, GA), but suppressed by drought and low nitrogen. Expression was inversely correlated with exogenous sucrose concentration, consistent with the canonical SnRK1 energy-sensing mechanism. Heterologous overexpression of <i>PpOSK4</i> in three independent homozygous T2 <i>Arabidopsis</i> lines (OE2, OE4, OE7; 57–72-fold overexpression) caused pronounced dwarfism (21–56% height reduction), reduced stomatal density, and enhanced drought and salt tolerance. In contrast, homologous overexpression in two independent Kentucky bluegrass lines (OE4, OE5; 19–31-fold overexpression) did not affect plant height but markedly enhanced leaf epicuticular wax biosynthesis and significantly improved drought and salt tolerance, as evidenced by higher chlorophyll retention, relative water content, and antioxidant enzyme (SOD, POD) activities, along with reduced electrolyte leakage and reactive oxygen species accumulation. This species-specific functional divergence between monocots and dicots represents a central finding, underscoring the necessity of validating gene functions in target species. This study provides the first functional characterization of an SnRK1β subfamily member in turfgrass, offering new molecular targets for stress-resistant breeding.</p>

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Functional analysis of PpOSK4 in Kentucky bluegrass: molecular basis for inhibiting Arabidopsis growth and enhancing drought resistance in Kentucky bluegrass

  • 期刊论文

  • 发表日期：

    2026年3月17日

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  • Yifeng Jin,
  • Xue You,
  • Haoran Yu,
  • Yansong Gao,
  • Qi Zhen,
  • Yushu Wang,
  • Yang Chen,
  • Miao He

摘要

Sucrose non-fermenting 1-related protein kinase (SnRK1) is a conserved central regulator of energy signaling and stress responses in plants. However, the functional roles of the SnRK1β regulatory subunit in turfgrass species remain largely unexplored. In this study, we identified and characterized PpOSK4, an SnRK1β homologue in Kentucky bluegrass (Poa pratensis L.), through integrative transcriptomic, molecular, and transgenic approaches. Mining of drought-responsive transcriptome data (PRJNA517968; three biological replicates per treatment; DESeq2, q < 0.05, |log₂FC| > 1) identified 38 differentially expressed SnRK family genes, among which PpOSK4 was selected for functional validation based on its significant drought responsiveness and high sequence conservation with rice OSK4 (95.4% nucleotide identity). PpOSK4 was preferentially expressed in roots and the cultivar ‘Midnight II’, induced by salt stress, high nitrogen, and exogenous hormones (GABA, IAA, SA, GA), but suppressed by drought and low nitrogen. Expression was inversely correlated with exogenous sucrose concentration, consistent with the canonical SnRK1 energy-sensing mechanism. Heterologous overexpression of PpOSK4 in three independent homozygous T2 Arabidopsis lines (OE2, OE4, OE7; 57–72-fold overexpression) caused pronounced dwarfism (21–56% height reduction), reduced stomatal density, and enhanced drought and salt tolerance. In contrast, homologous overexpression in two independent Kentucky bluegrass lines (OE4, OE5; 19–31-fold overexpression) did not affect plant height but markedly enhanced leaf epicuticular wax biosynthesis and significantly improved drought and salt tolerance, as evidenced by higher chlorophyll retention, relative water content, and antioxidant enzyme (SOD, POD) activities, along with reduced electrolyte leakage and reactive oxygen species accumulation. This species-specific functional divergence between monocots and dicots represents a central finding, underscoring the necessity of validating gene functions in target species. This study provides the first functional characterization of an SnRK1β subfamily member in turfgrass, offering new molecular targets for stress-resistant breeding.