Storage proteins are essential for seed germination and seedling growth, as they provide an indispensable nitrogen source and energy. Our previous report highlighted the defective endosperm development in the serine hydroxymethyltransferase 4 (OsSHMT4) gene mutant, floury endosperm20-1 (flo20-1). However, the alterations in storage protein content and distribution within the flo20-1 endosperm remained unclear. Here, the immunocytochemistry analyses revealed a deficiency in storage protein accumulation in flo20-1. Electron microscopic observation uncovered abnormal morphological structures in protein bodies (PBI and PBII) in flo20-1. Immunofluorescence labeling demonstrated that aberrant prolamin composition could lead to the subsequent formation and deposition of atypical structures in protein body I (PBI), and decreased levels of glutelins and globulin resulted in protein body II (PBII) malformation. Further RNA-seq data combined with qRT-PCR results indicated that altered transcription levels of storage protein structural genes were responsible for the abnormal synthesis and accumulation of storage protein, which further led to non-concentric ring structural PBIs and amorphous PBIIs. Collectively, our findings further underscored that OsSHMT4 is required for the synthesis and accumulation of storage proteins and storage organelle formation in endosperm cells.

Cotton is recognized as the primary source of essential natural fibers for the global textile industry, supporting its sustainability and development. However, adverse environmental conditions such as drought severely constrain cotton production; thus, developing stress-tolerant cultivars via molecular breeding is essential for maintaining yield stability. Here, a comprehensive functional dissection was conducted on GhGA2ox15, a gibberellin 2-oxidase gene derived from Gossypium hirsutum L. This gene encodes a key catabolic enzyme implicated in the deactivation of endogenous bioactive GAs and the modulation of stress adaptation. We characterized GhGA2ox15, a GA2ox gene from upland cotton that modulates endogenous bioactive GA levels and abiotic stress tolerance. Bioinformatics and sequence analyses confirmed that GhGA2ox15 is a canonical C₂₀-GA2ox subfamily member, with conserved DIOX_N and 2OG-FeII_Oxy domains and marked similarity to orthologs in Arabidopsis and rice. Tobacco subcellular localization assays indicated that GhGA2ox15 resides in both the nucleus and the cytoplasm. In transgenic Arabidopsis and Oryza sativa lines, GhGA2ox15 overexpression was shown to increase drought tolerance, while virus-induced gene silencing (VIGS) of GhGA2ox15 yielded significantly compromised drought resistance. Physiological assays linked GhGA2ox15 silencing to impaired reactive oxygen species (ROS) detoxification. The suppressed lines displayed markedly lower antioxidant enzyme activities, concomitant ROS accumulation in leaves, and attenuated transcription of drought-responsive marker genes. Our findings delineate the mechanistic role of GhGA2ox15 in drought adaptation and highlight its potential utility in breeding drought-tolerant cotton.