Beobachtung einzelner mRNA Transkripte im Zellkern und während des Kernporenkomplex vermittelten Exports in lebenden Zellen

DNA的遗传信息被转录成mRNA（信使RNA），并输出到细胞质中。在那里，mRNA被一种叫做核糖体的特殊机器翻译成蛋白质。mRNA可用性的控制是细胞代谢的中心调控机制。在这个项目中，单个mRNA分子将从其诞生到出现在细胞质中。在转录过程中，RNA被修饰和包装。RNA确实以信使核糖核粒子（mRNP）的形式存在于细胞核中。目前尚不清楚这些mrnp如何以及如何被修改以被出口机制识别。一次输出事件的时间，mRNA在输出过程中的方向，mRNA是否展开以及运输如何终止的问题对于阐明我们对细胞功能的理解很重要。在此之前，为活细胞内的显微应用标记mRNA是不可能的。利用一种新的病毒系统（MS2），可以修饰一个基因，使其mRNA包含一个被MS2结合蛋白（MBP）高度特异性结合的序列。MBP对哺乳动物细胞来说不是内源性的，可以与荧光蛋白结合作为mRNA的标记物。为了在活细胞中跟踪这种ms2标记的mrna，将建立一种新的多路复用高速显微镜，允许以毫秒时间分辨率观察至少三种不同的荧光通道。

The genetic information of DNA is transcribed into mRNA (messenger RNA), which gets exported into the cytoplasm. There mRNA is translated into proteins by a special machinery, called Ribosome. The control of mRNA availability is a central regulatory mechanism for cellular metabolism. Within this project single mRNA molecules shall be followed from their birth to their appearance in the cytoplasm. While transcribed, RNA gets modified and packed. RNA does exist as mRNP (messenger Ribo-Nucleo-Particle) in the nucleus. How and than these mRNPs gets modified to be recognized by the export machinery is currently not known. The time of one export event, the orientation of the mRNA during export, the question if the mRNA gets unfolded and how transport is terminated are important to elucidate our understanding of cellular function. Before it was impossible to label mRNA for microscopic applications inside living cells. Using a new viral system (MS2), it is possible to modify a gene so its mRNA contains a sequence which is highly specifically bound by the MS2 binding protein (MBP). MBP is not endogenous for mammalian cells and can be conjugated to a fluorescent protein to be used as marker for the mRNA. To track such MS2-tagged mRNAs in living cells a new multiplexing high speed microscope will be built, allowing the observation of at least three different fluorescence channels with millisecond time resolution.