Supporting Online Material Materials and Methods Som Text Figs. S1 to S8 Table S1 Movies S1 to S3 a " Silent " Polymorphism in the Mdr1 Gene Changes Substrate Specificity Corrected 30 November 2007; See Last Page-猫眼课题宝

DOI：

发表时间：

期刊：

影响因子：

通讯作者：

中科院分区：

其他

文献类型：

作者：

E Piddini;F. Marshall;L. Dubois;E. Hirst;J.-P Vincent;J Lippincott-Schwartz;N. Altan-Bonnet

在这些体内条件下，Wg产生l = 5.8 ± 2.04 mm的短程梯度（图1A和1B）。3 I和4A）。哪个动力学参数可以解释这种差异？由于Dpp和Wg具有固有的不同性质-Wg是脂质修饰的分子（20）; Dpp不是（21）-它们可能显示出不同的通过上皮扩散的机制和动力学。Wingless梯度的衰减长度较短是由于GFP-Wingless的降解速率较高，为5倍，并且在较小程度上其扩散系数较小（图4）。虽然在Dpp和Wg实验中Gal 4驱动剂是相同的，但Wg的生产速率是Dpp的约7倍，这意味着它们的成熟和分泌受到不同的控制。此外，虽然62%的Dpp分子是不移动的，但Wg库在25°C下几乎是完全移动的（y = 9.2 ± 13%），尽管与Dpp不同，在较高的实验温度下出现了显著的不移动的部分（图3，K和N）。在25°C下Dpp和Wg的不同不动级分验证了Dpp不动级分的特异性。因此，不动部分不是病态细胞中不完全恢复的伪影。最后，与Dpp相反，Wg的转运和降解不依赖于发动蛋白的内吞作用（图3，K至O）。事实上，Wg运动已经被认为是动力素独立的（6，22）。此外，显性负性发动蛋白和/或长期热敏性shibire阻滞的表达导致机翼中梯度的延伸（5，22，23），这归因于降解的减少（5，22）。我们的FRAP方法研究的结果表明，急性块内吞作用是不需要的Wg运输和降解，或者，Wg的内吞作用是动力蛋白独立的。总之，GFP-无翼FRAP实验（i）验证了我们的FRAP测定和shibire-拯救实验;（ii）表明可以通过独立地微调D、k、n和y来产生不同的形态发生梯度;以及（iii）表明不同的形态发生可以使用不同的转运机制和细胞机制（例如，动力蛋白依赖性转运与动力蛋白非依赖性转运）以实现形态发生剂梯度的形成。同义单核苷酸多态性（SNP）不会改变编码序列，因此预计不会改变其所在蛋白质的功能。我们报告了多药耐药1（MDR 1）基因中的同义SNP，该基因是先前与MDR 1基因功能改变相关的单倍型的一部分。

these in vivo conditions, Wg made a short-range gradient with l = 5.8 ± 2.04 mm (Figs. 3I and 4A). Which kinetic parameter could account for this difference? Because Dpp and Wg have inherently different properties—Wg is a lipid-modified molecule (20); Dpp is not (21)—they are likely to display different mechanisms and kinetics of spreading through the epithelium. The shorter decay length of the Wingless gradient was due to a higher degradation rate of GFP-Wingless, by a factor of 5, and to a lesser extent its smaller diffusion coefficient (Fig. 4). Although the Gal4 driver was the same in the Dpp and Wg experiments, the production rate of Wg was about seven times that of Dpp, which implied that their maturation and secretion were controlled differently. In addition, while 62% of the Dpp molecules were immobile, the Wg pool was almost fully mobile at 25°C (y = 9.2 ± 13%), although, unlike Dpp, a significant immobile fraction appeared at higher experimental temperatures (Fig. 3, K and N). The different immobile fractions of Dpp and Wg at 25°C validated the specificity of the Dpp immobile fraction. Thus, the immobile fraction was not an artifact of incomplete recovery in sick cells. Finally, in contrast to Dpp, Wg transport and degradation were independent of Dynamin endocytosis (Fig. 3, K to O). Indeed, Wg movement has been suggested to be Dynamin-independent (6, 22). In addition , expression of dominant-negative Dynamin and/or long-term thermosensitive shibire block caused an extension of the gradient in the wing (5, 22, 23), which was attributed to decreased degradation (5, 22). Our FRAP approach studying the results of an acute block suggests that endocytosis is not required for Wg transport and degradation or, alternatively, that endocytosis of Wg is Dynamin-independent. Altogether, the GFP-Wingless FRAP experiments (i) validated our FRAP assay and shibire-rescue experiment; (ii) indicated that different morphogen gradients can be generated by independently fine-tuning D, k, n, and y; and (iii) showed that different morphogens may use different mechanisms of transport and cellular machineries (e.g., Dynamin-dependent versus Dynamin-independent transport) to achieve the formation of morphogen gradients. Synonymous single-nucleotide polymorphisms (SNPs) do not produce altered coding sequences, and therefore they are not expected to change the function of the protein in which they occur. We report that a synonymous SNP in the Multidrug Resistance 1 (MDR1) gene, part of a haplotype previously linked to altered function of the MDR1 gene …

Supporting Online Material Materials and Methods Som Text Figs. S1 to S8 Table S1 Movies S1 to S3 a " Silent " Polymorphism in the Mdr1 Gene Changes Substrate Specificity Corrected 30 November 2007; See Last Page

文献摘要

被引文献

相似文献