Fang Xiaolan, Lu Qiuheng, Emoto Kazou, Adler Paul N
Biology Department, Cell Biology Department, Cancer Center, Morphogenesis and Regenerative Medicine Institute, University of Virginia, Charlottesville, VA 22903, USA.
BMC Dev Biol. 2010 Apr 20;10:40. doi: 10.1186/1471-213X-10-40.
Cell polarity is a common feature of eukaryotic cells. The NDR kinases have been found to regulate polarized growth in both animal cells and fungi. Drosophila Tricornered is an NDR kinase that is essential for the normal polarized growth of extensions of epidermal cells and for the tiling and branching of dendrites of da sensory neurons. Tricornered function requires interacting with the large Furry protein (3479 amino acid).
We constructed a furry (fry) transgene and established that it rescued the lethality of fry null mutations. The encoded protein was tagged at both its amino and carboxy termini and this allowed us to demonstrate that the protein existed as an uncleaved protein in vivo. We used the C terminal GFP tag to follow the protein in vivo and found it to be highly mobile. Interestingly Fry accumulated at the distal tip of growing bristles. We established that Fry and Trc could be co-immunoprecipitated from wing discs.
The mobility of Fry in both bristles and dendrites suggests that it could function in directing/mediating the intracellular transport needed for polarized growth. Our observations that full length Fry and Trc show only partial co-localization in growing bristles while an amino terminal fragment of Fry shows close to complete co-localization with Trc suggests that the interaction between these proteins is transient and regulated.
细胞极性是真核细胞的一个共同特征。已发现NDR激酶可调节动物细胞和真菌中的极性生长。果蝇三角蛋白是一种NDR激酶，对于表皮细胞延伸的正常极性生长以及da感觉神经元树突的平铺和分支至关重要。三角蛋白的功能需要与大的Furry蛋白（3479个氨基酸）相互作用。
我们构建了一个furry（fry）转基因，并确定它挽救了fry无效突变的致死性。编码的蛋白质在其氨基和羧基末端都有标签，这使我们能够证明该蛋白质在体内以未切割的形式存在。我们使用C末端GFP标签在体内追踪该蛋白质，发现它具有高度的流动性。有趣的是，Fry聚集在生长刚毛的远端。我们确定Fry和Trc可以从翅芽中共免疫沉淀。
Fry在刚毛和树突中的流动性表明它可能在指导/介导极性生长所需的细胞内运输中发挥作用。我们观察到全长Fry和Trc在生长的刚毛中仅部分共定位，而Fry的氨基末端片段与Trc几乎完全共定位，这表明这些蛋白质之间的相互作用是短暂的且受调控的。
