Systematic transcriptomics reveals a biphasic mode of sarcomere morphogenesis in flight muscles regulated by Spalt

Muscles organise a pseudo-crystalline array of actin, myosin and titin filaments to build force-producing sarcomeres. To study how sarcomeres are built, we performed mRNA-sequencing of developing Drosophila flight muscles and identified 40 distinct expression profile clusters. Strikingly, two clusters are strongly enriched for sarcomeric components. Temporal gene expression together with detailed morphological analysis enabled us to define two distinct phases of sarcomere development, both of which require the transcriptional regulator Spalt major. During the first sarcomere formation phase, 2.0 µm long immature sarcomeres assemble myofibrils that spontaneously contract. In the second sarcomere maturation phase, sarcomeres grow to their final 3.2 µm length and 1.5 µm diameter and acquire stretch-sensitivity. Interestingly, the final number of myofibrils per flight muscle fiber is determined at the onset of the first phase and remains constant. Together, this defines a biphasic mode of sarcomere and myofibril morphogenesis – a new concept which may also apply to vertebrate muscle or heart development. Part I: An 8-point timecourse of wild-type flight muscle development in Drosophila melanogaster was analyzed with duplicates/triplicates for each timepoint Part II: A Mef2-Gal4 x salmIR timecourse in duplicate at 4 timepoints was compared to wild-type flight muscle

肌肉通过组装肌动蛋白、肌球蛋白与肌联蛋白丝的伪晶阵列，构建可产生收缩力的肌节（sarcomere）。为探究肌节的构建机制，我们对发育中的果蝇（Drosophila）飞行肌开展了mRNA测序（mRNA-sequencing），并鉴定出40个不同的表达谱聚类簇。值得注意的是，其中两个聚类簇显著富集肌节相关组分。结合时序基因表达数据与精细形态学分析，我们明确了肌节发育的两个截然不同的阶段，两个阶段均依赖转录调控因子Spalt major。在第一阶段即肌节形成期，长度为2.0 μm的未成熟肌节组装成可自发收缩的肌原纤维（myofibril）。在第二阶段即肌节成熟期，肌节生长至最终的3.2 μm长度与1.5 μm直径，并获得牵张敏感性。有趣的是，每条飞行肌纤维最终含有的肌原纤维数量在第一阶段启动时便已确定，且此后保持恒定。综上，本研究明确了肌节与肌原纤维形态发生的双相模式——这一新概念或许同样适用于脊椎动物肌肉或心脏发育。
第一部分：对黑腹果蝇（Drosophila melanogaster）野生型飞行肌发育的8个时间节点进行了时序分析，每个时间点设置双重复或三重复样本。
第二部分：对4个时间点的Mef2-Gal4 × salmIR样本（双重复）进行时序分析，并与野生型飞行肌进行对比。