Lee J K, Coyne R S, Dubreuil R R, Goldstein L S, Branton D
Department of Cellular and Developmental Biology, Harvard University, Cambridge, Massachusetts 02138.
J Cell Biol. 1993 Dec;123(6 Pt 2):1797-809. doi: 10.1083/jcb.123.6.1797.
We show that the alpha-spectrin gene is essential for larval survival and development by characterizing several alpha-spectrin mutations in Drosophila. P-element minigene rescue and sequence analysis were used to identify the alpha-spectrin gene as the l(3)dre3 complementation group of the Dras-Roughened-ecdysoneless region of chromosome 3 (Sliter et al., 1988). Germ line transformants carrying an alpha-spectrin cDNA, whose expression is driven by the ubiquitin promoter, fully rescued the first to second instar lethality characteristic of the l(3)dre3 alleles. The molecular defects in two gamma-ray-induced alleles were identified. One of these mutations, which resulted in second instar lethality, contained a 73-bp deletion in alpha-spectrin segment 22 (starting at amino acid residue 2312), producing a premature stop codon between the two EF hands found in this segment. The second mutation, which resulted in first instar lethality, contained a 20 base pair deletion in the middle of segment 1 (at amino acid residue 92), resulting in a premature stop codon. Examination of the spectrin-deficient larvae revealed a loss of contact between epithelial cells of the gut and disruption of cell-substratum interactions. The most pronounced morphological change was seen in tissues of complex cellular architecture such as the middle midgut where a loss of cell contact between cup-shaped cuprophilic cells and neighboring interstitial cells was accompanied by disorganization of the cuprophilic cell brush borders. Our examination of spectrin deficient larvae suggests that an important role of non-erythroid spectrin is to stabilize cell to cell interactions that are critical for the maintenance of cell shape and subcellular organization within tissues.
通过对果蝇中几种α-血影蛋白突变进行特征分析,我们发现α-血影蛋白基因对幼虫的存活和发育至关重要。利用P因子微型基因拯救和序列分析,将α-血影蛋白基因鉴定为3号染色体Dras-粗糙-无蜕皮激素区域的l(3)dre3互补群(Sliter等人,1988年)。携带由泛素启动子驱动表达的α-血影蛋白cDNA的生殖系转化体,完全拯救了l(3)dre3等位基因特有的一龄到二龄致死性。鉴定了两个γ射线诱导等位基因中的分子缺陷。其中一个导致二龄致死的突变,在α-血影蛋白第22段(从氨基酸残基2312开始)中有一个73bp的缺失,在该段发现的两个EF手之间产生了一个提前终止密码子。第二个突变导致一龄致死,在第1段中间(氨基酸残基92处)有一个20bp的缺失,导致提前终止密码子。对血影蛋白缺陷幼虫的检查显示,肠道上皮细胞之间失去接触,细胞与基质的相互作用受到破坏。最明显的形态学变化出现在细胞结构复杂的组织中,如中肠中部,杯状嗜铜细胞与相邻间质细胞之间失去细胞接触,同时嗜铜细胞刷状缘紊乱。我们对血影蛋白缺陷幼虫的检查表明,非红细胞血影蛋白的一个重要作用是稳定细胞间相互作用,这对维持组织内细胞形状和亚细胞组织至关重要。