Functional characterization of wheat GSK kinase TaSK41 and screening for interacting proteins

SK41, a plant glycogen synthase kinase 3 (GSK3) /SHAGGY-like kinases gene, plays an important role in grain development and grain weight formation. To further elucidate the biological function of TaSK41 in regulating wheat grain development and its underlying molecular mechanisms, this study examined the expression patterns of TaSK41 across different tissues, its subcellular localization, the grain phenotypes of transgenic rice overexpressing TaSK41, and its interacting proteins. qRT-PCR analysis revealed that TaSK41 was ubiquitously expressed, with particularly high expression in spikes, early developing grains, ovaries, and seed coats. Subcellular localization assays showed that the TaSK41-GFP fusion protein was predominantly localized in both the cytoplasm and the nucleus. Overexpression of TaSK41 in transgenic rice led to a reduction in thousand-grain weight, accompanied by significant decreases in both grain length and width. Through yeast two-hybrid screening, 17 candidate proteins were identified as potential interactors with TaSK41. Among these, full-length interaction validation was conducted for TaARF4and TaBSK3, both of which are associated with grain development. Point-to-point assays confirmed that TaSK41 interacts with the full-length TaARF4 protein. This interaction was further validated in vivo using a luciferase complementation assay, confirming that TaSK41 physically interacts with TaARF4 in plant cells. These findings provide a theoretical basis for future investigations into the molecular mechanisms by which TaSK41 regulates grain weight formation in wheat.