Heat shock genes and the response to environmental stress during zebrafish development.

热休克蛋白在暴露于环境压力(如高温或有毒金属)后在细胞中表达，并帮助细胞度过这些应激期。几个热休克蛋白还扮演着重要的分子伴侣角色，通常帮助其他蛋白质折叠并到达它们在细胞中的正确目的地。对于细胞来说，能够调控热休克蛋白在哪里、何时以及表达多少是很重要的，这在很大程度上是通过调控编码热休克蛋白的基因的表达来实现的。 汞和镉等有毒金属是一个非常令人担忧的问题，对胚胎发育有负面影响。水银会导致一系列问题，包括人类和鱼类的视觉缺陷。镉会损害人类和鱼类的嗅觉器官，导致嗅觉丧失。关于有毒金属的一个主要问题是，它们是否在某些细胞和组织中比其他细胞和组织积累得更多，以及金属的化学形态是否影响这一过程。 我们的工作以斑马鱼胚胎为模型，研究环境压力对发育的影响。斑马鱼是一个受欢迎的模型系统，因为它们的大部分发育过程与人类发育相似，此外还是观察鱼类发育本身的一个很好的模型。我们使用同步加速器x射线检查了有毒金属汞在斑马鱼胚胎中的积聚位置。我们发现，其中一个主要的目标是感觉系统的细胞，特别是眼睛、嗅觉器官和鱼类中被称为侧线的特殊感觉器官的细胞。我们还检测了一种名为HSP70的特定热休克蛋白。有趣的是，HSP70在眼睛发育过程中正常表达，是正常眼睛形成所必需的。在接触另一种有毒金属镉后，该基因也在嗅觉器官和侧线中表达。 这项提议中的实验将检查斑马鱼胚胎暴露于不同类型汞的影响，以及吸收大量汞的细胞是否也是最敏感的。这将包括观察感觉器官中的细胞是否更容易死亡，以及器官的正常功能是否被破坏。我们还将研究HSP70基因在汞和镉处理的胚胎中是如何打开和关闭的，以及在眼睛晶状体正常发育期间是如何打开和关闭的。其中一些工作将使用我们开发的转基因鱼类模型，该模型可以在表达HSP70的细胞中发出明亮的绿色荧光，从而能够非常快速地检测到应激细胞。综上所述，这项工作将使我们能够开发一个完整的模型，了解在正常发育期间和胚胎暴露于环境应激之后，HSP70基因的调控是如何进行微调的，以及这与靶细胞中发生的其他事件(从有毒金属的摄取和积累到靶器官的功能缺陷)有何关系。 这项工作将对我们在分子和细胞水平上理解胚胎中的细胞如何响应和生存于环境应激具有重要意义。我们将能够将对感觉细胞及其功能的影响与i)这些细胞中有毒金属的实际数量，ii)HSP70基因如何在这种应激反应中被激活，以及iii)这种激活如何帮助胚胎细胞在应激事件中存活相关。这也将使我们更好地了解HSP70基因在眼睛正常发育过程中是如何表达的，并进一步阐明这一事件的重要性。这项工作将进一步确定我们的转基因鱼品系是否可以用作某些感官器官阴性结果的快速荧光预测。

Heat shock proteins are expressed in cells following exposure to environmental stress such as elevated temperature or toxic metals, and help cells survive these periods of stress. Several hsps also play important roles as molecular chaperones that normally help other proteins fold and reach their correct destination in the cell. It is important for a cell to be able to regulate where, when, and how much a heat shock protein is expressed, and it does this largely be regulating the expression of the gene that codes for the hsp. Toxic metals such as mercury and cadmium are a significant concern, and have negative effects on developing embryos. Mercury causes a range of problems, including visual defects in both humans and fish. Cadmium can damage the olfactory organ of both humans and fish, causing a loss of the sense of smell. A major question regarding toxic metals is whether they accumulate more in some cells and tissues than others, and whether the chemical form of the metal affects this process. Our work uses the zebrafish embryo as a model to examine the effects of environmental stress on development. Zebrafish are a popular model system since much of their development occurs in a similar way to human development, in addition to being an excellent model to look at fish development itself. We have used synchrotron x-rays to examine where in zebrafish embryos the toxic metal mercury accumulates. We found that one of the major targets is cells of sensory systems, in particular parts of the eye, olfactory organ, and a specialized sensory organ in fish known as the lateral line. We have also examined a specific heat shock protein known as hsp70. Interestingly, hsp70 is expressed normally during eye development, and is required for normal eye formation. This gene is also expressed in both the olfactory organ and lateral line following exposure to another toxic metal, cadmium. Experiments in this proposal will examine the impact of exposing zebrafish embryos to different types of mercury, and whether the cells that take up large amounts of mercury are also the most sensitive. This will include looking at whether cells in the sensory organs are more prone to death, and whether the proper function of the organs is disrupted. We will also examine how the hsp70 gene is turned on and off in mercury and cadmium treated embryos, and during normal development of the eye lens. Some of the work will use a transgenic fish model we have developed that fluoresces bright green in cells that express hsp70, allowing for very rapid detection of stressed cells. In summary, the work will allow us to develop an integrated model of how regulation of the hsp70 is gene is fine-tuned during normal development and following exposure of embryos to environmental stress, and how this relates to other events occurring in target cells ranging from toxic metal uptake and accumulation to functional defects in target organs. This work will be important to our understanding of how cells in an embryo respond to and survive exposure to environmental stress at the molecular and cellular level. We will be able to correlate impacts on sensory cells and their function with i) the actual amount of a toxic metal in these cells, ii) how the hsp70 gene is activated in response to this stress, and iii) how this activation may assist embryonic cells in surviving the stress event. It will also allow us to develop a better understanding of how the hsp70 gene is expressed during normal development of the eye, and further elucidate the importance of this event. This work will further determine if our transgenic fish strain can be used as a rapid fluorescent predictor of negative outcomes in some sensory organs.