Elucidating the mechanisms of chromosome length sensing by the synaptonemal complex

在本财政年度，我们通过对体内条件进行几项调整，改进了成像技术。我们已经成功地跟踪了mMaple 3标记的SYP-3蛋白在C. elegans，同时通过使用eGFP标记的COSA-1蛋白可视化交叉指定位点的位置。虽然COSA-1和未活化的mMaple 3-SYP-3在绿色通道中都是荧光的，但我们发现了标记交叉的COSA-1焦点在表征SYP-3染色的线性信号背景下明确可见的条件。因此，我们能够克服仅具有两个荧光通道来测量三个所需信号（未活化的mMaple 3-SYP-3、活化的mMaple 3-SYP-3和COSA-1）的限制。此外，我们还开发了一套更新的图像分析工具（https：//github.com/carltonlab/carltonlab_imagej_tools）和模拟软件（https：//github.com/carltonlab/scsim），用于生成和测试我们的扩散和积累模型的预测。领导该项目的研究生根据研究计划，在2022财年的Meiosis Gordon会议上介绍了我们的工作进展，并收到了研究界的宝贵反馈。

In this fiscal year we have improved our imaging techniques by making several adjustments to the in vivo conditions. We have succeeded in following the dynamic movement of mMaple3-labeled SYP-3 protein within and between meiotic chromosomes of C. elegans, while simultaneously visualizing the position of crossover designation sites by using eGFP-labeled COSA-1 protein. While both COSA-1 and non-activated mMaple3-SYP-3 are fluorescent in the green channel, we have found conditions where the COSA-1 foci that mark crossovers are unambiguously visible against the background of the linear signal characterizing SYP-3 staining. Thus, we were able to overcome the limitation of having only two fluorescent channels to measure three desired signals (non-activated mMaple3-SYP-3, activated mMaple3-SYP-3, and COSA-1). Further, we have developed an updated set of image analysis tools (https://github.com/carltonlab/carltonlab_imagej_tools) and simulation software (https://github.com/carltonlab/scsim) that we use to generate and test predictions of our diffusion and accumulation model.The graduate student leading this project has presented our work-in-progress at the Meiosis Gordon Conference in FY2022, in accordance with the research plan, and received valuable feedback from the research community.