Characterization of a non-specific lipid transfer protein from Betula platyphylla  and identification of Bp ns LTPs  response to drought stress

The non-specific lipid transfer protein (nsLTP) is a plant gene family that significantly influences plant growth, development, and adaptation to environmental stresses, particularly in response to biotic and abiotic stresses. This study characterized the nsLTP family and identified 23 nsLTP genes in birch ( Betula platyphylla ). Based on the sequence similarity and the presence of eight cysteine amino acid residues, BpnsLTP proteins were clustered into five types: type I, III, IV, V and VI. Seven nsLTPs were screened using by drought stress transcriptome analysis and qRT-PCR results proved that three nsLTP genes were positively responded to drought stress and four nsLTP genes were negatively responded to drought stress. Co-expression and go enrichment analysis revealed that genes co expressed with LTP genes were enriched in which futher domastrated these LTP were invleved plant abiotic stress. Subcellular localization experiment showed that BpnsLTPI.3 proteins were localized on cell membrane. Obtaining BpnsLTPI.3 overexpressing birch transgenic lines through leaf disk method, after treating transgenic and wild-type lines with 10% PEG6000 stress, various physiological and biochemical parameters were assessed. It was observed that the drought tolerance of the transgenic lines was significantly lower compared to the wild-type lines. Specifically, NBT and Evans blue staining indicated more pronounced stress-induced damage in the transgenic lines than in the wild-type. Moreover, the transgenic lines exhibited lower levels of protective enzymes such as POD and SOD, while the content of MDA and H ₂ O ₂ was higher in comparison to the wild-type lines. These results suggest that BpnsLTPI.3 , under the PEG6000 stress treatment, modulates protective enzyme activities, scavenges reactive oxygen species, and maintains cellular integrity in a manner that ultimately diminishes the drought tolerance of the transgenic birch plants. These results provide new insights into the biological functions of the nsLTP family of B. platyphylla in response to abiotic stress.