DROSOPHILA-MELANOGASTER TROPONIN-T MUTATIONS ENGENDER 3 DISTINCT SYNDROMES OF MYOFIBRILLAR ABNORMALITIES
DROSOPHILA-MELANOGASTER TROPONIN-T MUTATIONS ENGENDER 3 DISTINCT SYNDROMES OF MYOFIBRILLAR ABNORMALITIES
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DOI：
10.1016/0022-2836(90)90390-8
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发表时间：
1990-12-05
影响因子：
5.6
通讯作者：
SAVILLE, DL
中科院分区：
文献类型：
--
作者：
FYRBERG, E;FYRBERG, CC;SAVILLE, DL
在脊椎动物中，肌钙蛋白复合物与原肌球蛋白二聚体相互作用以调节骨骼肌收缩。为了进一步研究肌钙蛋白在体内的组装和功能，我们正在开发分子遗传学方法。在这里，我们报告表征的基因编码果蝇肌钙蛋白-T和分析肌肉缺陷所产生的几个突变等位基因。我们发现，果蝇肌钙蛋白-T基因座指定至少三种蛋白质具有类似于脊椎动物肌钙蛋白-T的序列。然而，由于高度酸性的羧基末端延伸，所有这些都明显大于任何鸟类或哺乳动物的同种型。脊椎动物和果蝇肌钙蛋白-T基因的染色体排列的比较显示，一个内含子-外显子边界的位置是保守的。这一观察结果以及脊椎动物和果蝇肌钙蛋白-T一级序列的相似性表明，各自的蛋白质是同源的，并且肌钙蛋白-T早于脊椎动物和无脊椎动物生物体的分歧。果蝇肌钙蛋白-T基因多线染色体的原位杂交表明，它位于亚部12 A的X染色体，正是在支持和缩进胸飞行肌突变已映射以前。我们确定了五个现存突变体的肌钙蛋白-T基因的核苷酸序列。所有这些都有有害的改变，直接确定了支撑和缩进的胸部肌肉异常是由于肌钙蛋白T缺陷。其中两个等位基因upheld 2和upheld 3明显破坏RNA剪接并从飞行和跳跃肌肉中消除大部分或全部肌钙蛋白T，而其余三个等位基因改变肌钙蛋白T的单个氨基酸的身份。突变体肌肉的电子显微镜显示，两个无效等位基因消除细丝，除非它们被电子致密材料结合，推测是Z盘蛋白。其中两个点突变，upheld 101和indented thorax 3，不会干扰肌原纤维的组装，但会在肌肉开始使用后的几天内导致它们的变性。最后一个突变upheldwhu使肌原纤维晶格的直径减小了大约一半。我们提出假说来解释每个肌钙蛋白T突变如何产生所观察到的肌原纤维缺陷。
In vertebrates troponin complexes interact co-operatively with tropomyosin dimers to modulate skeletal muscle contraction. In order further to investigate troponin assembly and function in vivo, we are developing molecular genetic approaches. Here we report characterization of the gene that encodes Drosophila troponin-T and analyses of muscle defects engendered by several mutant alleles. We found that the Drosophila troponin-T locus specifies at least three proteins having sequences similar to vertebrate troponin-T. All are significantly larger than any avian or mammalian isoforms, however, due to a highly acidic carboxy-terminal extension. Comparisons of the chromosomal arrangements of vertebrate and Drosophila troponin-T genes revealed that the location of one intron-exon boundary is conserved. This observation and the similarity of vertebrate and Drosophila troponin-T primary sequences suggest that the respective proteins are homologous, and that troponin-T pre-dates the divergence of vertebrate and invertebrate organisms. In situ hybridization of the Drosophila troponin-T gene to polytene chromosomes demonstrated that it resides within subdivision 12A of the X chromosome, precisely where upheld and indented thorax flight muscle mutations have been mapped previously. We determined the nucleotide sequences of troponin-T genes in five extant mutants. All have deleterious alterations, directly establishing that upheld and indented thorax muscle abnormalities are due to defective troponin-T. Two of the alleles, upheld2 and upheld3, apparently disrupt RNA splicing and eliminate most or all troponin-T from flight and jump muscles, while the remaining three alleles change the identities of single amino acids of troponin-T. Electron microscopy of mutant muscles revealed that the two null alleles eliminate thin filaments, except where they are bound by electron-dense material presumed to be Z-disc proteins. Two of the point mutations, upheld101 and indented thorax3, do not perturb assembly of myofibrils, but cause their degeneration within days after muscles begin to be utilized. The final mutation, upheldwhu, reduces the diameter of the myofibril lattice by approximately one-half. We propose hypotheses to explain how each troponin-T mutation engenders the observed myofibrillar defects.