Seasonal Adaptation of a Circadian Clock

描述(由申请人提供)：本申请是对最近授予的2 R01-NS42088资助的竞争性补充。2 R01-NS42088的总体目标是了解昼夜节律起搏器如何适应环境温度和日长(光周期)的季节性变化而适应新的相位对齐。提出的特定目标的一个主要基础是基于我们先前的工作，即从黑腹果蝇时期(PER)RNA(称为dmpi8；D.Blackogaster dper内含子8)的3‘端内含子的温度和光敏剪接起到传达日历信息的多模式“季节传感器”的作用。我们还发现编码磷脂酶C(PLC？)的NORPA基因调控温度和时钟控制dmpi8的剪接效率。这些发现确定了Norpa的一个新的非光作用。我们建议确定温度、光、时钟因素和Norpa/脂质代谢如何共同调节dmpi8的剪接效率，以及这一分子事件对节律行为的季节性适应的贡献。在此，我们请求获得更多资金，用于购买急需的设备，这些设备对实现核定的具体目标至关重要，并将大大扩大项目的范围。该项目的主要实验策略包括测量RNA水平、荧光素酶活性和果蝇运动活动节律。最初申请中没有预料到的最近事件也为扩大特定目标的范围提供了新的可能性。例如，我们将评估新描述的温度偏好突变体在季节性适应方面的变化。然而，我们目前实现关键目标的能力严重有限。为了增强我们的研究能力，我们要求提供资金购买以下仪器：(1)实时聚合酶链式反应系统，(2)微板发光仪，(3)环境孵化器和(4)运动活动监测器。此外，我们要求为一名研究技术员提供工资支持，他将100%地致力于这项提议(目前正在投入50%的努力)。设备和人员的要求将对我们的研究产生显着影响，这些研究旨在了解时钟功能的季节性适应，以及Norpa如何参与校准黑腹毛虫的分子和行为热反应。

DESCRIPTION (provided by applicant): This application is a competitive supplement to a recently awarded grant, 2 R01-NS42088. The overall goal of 2 R01-NS42088 is to understand how a circadian pacemaker accommodates new phase alignments in response to seasonal changes in ambient temperature and day length (photoperiod). A major foundation for the proposed specific aims is based on our prior work showing that the thermosensitive and photosensitive splicing of the 3'-terminal intron from the D. melanogaster period (per) RNA (termed dmpi8; D. melanogaster dper intron 8) acts as a multi-modal "seasonal sensor" conveying calendar information. We also showed that the norpA (no receptor potential A) gene encoding phospholipase C (PLC?), modulates the temperature and clock control of dmpi8 splicing efficiency. These findings identified a novel non-photic role for norpA. We proposed to determine how temperature, light, clock factors and NORPA/lipid metabolism co-regulate the splicing efficiency of dmpi8, and the contribution of this molecular event to seasonal adaptation in rhythmic behavior. Herein, we are requesting additional funds to purchase much needed equipment that is critical for progress on the approved specific aims and will significantly enhance the scope of the project. Major experimental strategies for this project involve measuring RNA levels, luciferase activity and Drosophila locomotor activity rhythms. Recent events not anticipated in the original application also offer novel possibilities to expand the scope of the specific aims. For example, we will evaluate newly described temperature-preference mutants for alterations in seasonal adaptation. However, our current capabilities in pursuing key objectives are severely limited. To enhance our research capabilities we request funding to purchase the following instruments; (1) Real-Time PCR system, (2) Microplate luminometer, (3) Environmental incubators and (4) Locomotor activity monitors. In addition, we request salary support for a Research Technician who will devote 100% effort to this proposal (currently devoting 50% effort). The equipment and personnel requests will have a marked impact on our studies aimed at understanding seasonal adaptation of clock function and how norpA participates in calibrating molecular and behavioral thermoresponses in D. melanogaster.