Miyawaki, A. | bioRxiv Science

bioRxiv2018

Moments reconstruction and local dynamic range compression of high order Superresolution Optical Fluctuation Imaging

AbstractSuper-resolution Optical Fluctuation Imaging (SOFI) offers a simple and affordable alternative to other super-resolution (SR) imaging techniques. The theoretical resolution enhancement of SOFI scales linearly with the cumulants order, while imaging conditions are less phototoxic to living samples as compared to other SR methods. High order SOFI could, therefore, be a method of choice for dynamic live cell imaging. However, due to cusp-artifacts and to dynamic range expansion of pixel intensities, this promise has not been materialized as of yet. Here we investigated and compared high order moments vs. high order cumulants SOFI reconstructions. We demonstrate that even-order moments reconstructions are intrinsically free of cusp artifacts, allowing for a subsequent deconvolution operation to be performed, hence enhancing the resolution even further. High order moments reconstructions performance was examined for various (simulated) conditions and applied to (experimental) imaging of QD labeled microtubules in fixed cells, and actin stress fiber dynamics in live cells.

bioengineering↗

bioRxiv2018

In vitro characterization of the human segmentation clock

The vertebral column is characterized by the periodic arrangement of vertebrae along the anterior-posterior (AP) axis. This segmental or metameric organization is established early in embryogenesis when pairs of embryonic segments called somites are rhythmically produced by the presomitic mesoderm (PSM). The tempo of somite formation is controlled by a molecular oscillator known as the segmentation clock 1,2. While this oscillator has been well characterized in model organisms 1,2, whether a similar oscillator exists in humans remains unknown. We have previously shown that human embryonic stem (ES) cells or induced pluripotent stem (iPS) cells can differentiate in vitro into PSM upon activation of the Wnt signaling pathway combined with BMP inhibition3. Here, we show that these human PSM cells exhibit Notch and YAP-dependent oscillations4 of the cyclic gene HES7 with a 5-hour period. Single cell RNA-sequencing comparison of the differentiating iPS cells with mouse PSM reveals that human PSM cells follow a similar differentiation path and exhibit a remarkably coordinated differentiation sequence. We also demonstrate that FGF signaling controls the phase and period of the oscillator. This contrasts with classical segmentation models such as the \"Clock and Wavefront\" 1,2,5, where FGF merely implements a signaling threshold specifying where oscillations stop. Overall, our work identifying the human segmentation clock represents an important breakthrough for human developmental biology.

developmental biology↗