A cotton LIM domain-containing protein (GhWLIM5) is involved in bundling actin filaments

LIM-domain proteins play important roles in cellular processes in eukaryotes. In this study, a LIM protein gene, GhWLIM5, was identified in cotton. Quantitative RT-PCR analysis showed that GhWLIM5 was expressed widely in different cotton tissues and had a peak in expression during fiber elongation. GFP fluorescence assay revealed that cotton cells expressing GhWLIM5:eGFP fusion gene displayed a network distribution of eGFP fluorescence, suggesting that GhWLIM5 protein is mainly localized to the cell cytoskeleton. When GhWLIM5:eGFP transformed cells were stained with rhodamine-phalloidin there was consistent overlap in eGFP and rhodamine-palloidin signals, demonstrating that GhWLIM5 protein is colocalized with the F-actin cytoskeleton. In addition, high-speed cosedimentation assay verified that GhWLIM5 directly bound actin filaments, while low cosedimentation assay and microscopic observation indicated that GhWLIM5 bundled F-actin in vitro. Increasing amounts of GhWLIM5 protein were able to protect F-actin from depolymerization in vitro in the presence of Lat B (an F-actin depolymerizer). Our results contribute to a better understanding of the biochemical role of GhWLIM5 in modulating the dynamic F-actin network in cotton.