Phosphoinositide cycle in Drosophila

光感受器通过一系列生化步骤将光转化为电信号，每个步骤都涉及特定的蛋白质分子（例如视觉色素和酶）。这种“光导级联”的最终结果是激活细胞周围脂质膜上被称为“离子通道”的蛋白质。一旦激活，离子通道打开，允许带电离子（如钠和钙）进入细胞，从而产生电信号并传输到大脑。我们可以很好地研究果蝇的光转导，因为我们可以很容易地操纵特定的基因（以及蛋白质），因为我们可以用一系列高精度的技术记录它们的光感受器的活动。参与其中的分子并不是飞行光感受器所独有的，在我们身体的细胞中也发现了与之密切相关的分子。其中一个分子就是所谓的TRP通道。在果蝇中，这是光导过程中激活的通道；在哺乳动物中，色氨酸通道对于许多重要过程都是必不可少的，比如激素反应、血压调节、味觉、嗅觉、痛觉、冷热感觉。果蝇光感受器用来激活TRP通道的特殊级联称为磷酸肌苷（PI）循环。这是活细胞中应用最广泛的生化级联反应之一。在人体中，它几乎存在于身体的每个细胞中，负责各种各样的激素反应，例如调节血压，以及大脑神经元与各种感官（如味觉）之间的交流。这个级联反应的激活涉及到所有细胞膜中被称为PIP2的重要脂质分子的分解。为了维持运作，必须不断地重新合成PIP2。这是一个复杂的循环，涉及多个步骤和至少5种不同的中间体。如果这些步骤中的任何一个受到损害，不仅级联反应停止工作，而且受影响的细胞也会死亡或癌变。为了在活细胞中研究这种循环，研究人员开发了荧光标记的分子，这些分子与每种中间体结合，然后可以用特殊的显微镜观察。通常这种实验是在人工条件下在培养细胞中进行的，这些细胞的行为可能与活组织中的细胞不同。在我们的研究中，我们将使用遗传技术在苍蝇的光感受器中表达一系列这样的荧光标记传感器。通过利用苍蝇眼睛的一些独特的光学特征，这将使我们能够在完整的活体动物中成像和测量它们的荧光。这将使我们能够跟踪各种中间体对生理刺激的反应，这在眼睛中可以通过光精确控制。此外，通过在候选基因中制造突变，我们还将能够确定负责调节PI周期每个步骤的基因。这种方法将使我们能够建立一个全面和定量的图像，这个重要的和无处不在的生化循环的功能在完整的动物。我们从这些研究中获得的知识不仅将进一步加深我们对光感受器如何看到的理解，而且由于基本的潜在生化机制被广泛发现，将为许多其他的，通常是临床上重要的身体过程提供新的见解。

Photoreceptors transduce light into electrical signals by a series of biochemical steps, each involving specific protein molecules (e.g. visual pigments and enzymes). The end result of this "phototransduction cascade" is the activation of proteins known as "ion channels", in the lipid membrane surrounding the cell. Once activated, ion channels open to allow charged ions, such as sodium and calcium, into the cell, thereby generating electrical signals for transmission to the brain. Phototransduction can be particularly well studied in the fruitfly Drosophila because of the ease with which we can manipulate specific genes (and hence proteins) and because we can record the activity from their photoreceptors with a range of high precision techniques. The molecules involved are not unique to fly photoreceptors and closely related molecules are found in cells throughout our own bodies. One such molecule is the so-called TRP channel. In flies, this is the channel activated during phototransduction; in mammals, TRP channels are essential for a wide range of vital processes such as hormonal responses, regulation of blood pressure, taste, smell, and sensations of pain, hot and cold. The particular cascade used by the fly photoreceptor to activate the TRP channels is called the phospho-inositide (PI) cycle. This is one of the most widely used biochemical cascades in living cells. In humans it is found in almost every cell in the body and is responsible for a wide range and hormonal responses, such as those involved in regulating blood pressure as well as in communication between neurons in the brain and various senses, such as taste. Activation of this cascade involves the breakdown of an important lipid molecule found in all cell membranes known as PIP2. To maintain operation it is essential that PIP2 is continually resynthesised. This takes place via a complex cycle involving multiple steps and at least 5 distinct intermediates. If any of these steps is compromised, not only does the cascade cease working, but the cells affected can die or become cancerous. In order to study this cycle in living cells, researchers have developed fluorescently labelled molecules which bind to each intermediate which can then be viewed with special microscopes. Normally such experiments are performed under artificial conditions in cultured cells which may differ in their behaviour to cells in living tissue. In our research we will use genetic techniques to express a range of such fluorescently labelled sensors in fly photoreceptors. By exploiting some unique optical features of the fly's eye this will enable us to image and measure their fluorescence in the completely intact living animal. This will allow us to follow the fate of the various intermediates in response to physiological stimulation, which in the eye can be precisely controlled by light. Additionally by making mutations in candidate genes, we will also be able to identify the genes responsible for regulating each step of the PI cycle. This approach will allow us to build up a comprehensive and quantitative picture of the functioning of this important and ubiquitous biochemical cycle in the intact animal. The knowledge we gain from these studies will not only further our understanding of how photoreceptors see but, because the basic underlying biochemical mechanisms are so widely found, will provide new insight into many other, often clinically important processes in the body.