Live-Cell Superresolution Imaging Reveals Intraflagellar Transport Dynamics in Primary Cilia
Jung-Chi Liao1*
1IAMS, Academia Sinica, Taipei, Taiwan
* Presenter:Jung-Chi Liao, email:jcliao@iams.sinica.edu.tw
Primary cilia play a vital role in cellular sensing and signaling. An essential component of ciliogenesis is intraflagellar transport (IFT), which first requires IFT-protein recruitment, IFT-protein–motor-protein assembly, axonemal engagement of IFT-protein complexes, and transition zone (TZ) gating. The mechanistic understanding of these processes at the ciliary base was largely missing, because it is exceedingly challenging to observe the motion of IFT proteins in this crowded region using conventional microscopy. Here, we report short trajectory tracking of IFT proteins at the base of mammalian primary cilia by optimizing single-particle tracking photoactivated localization microscopy (sptPALM), balancing the imaging requirements of tracking speed, tracking duration, and localization precision for IFT88-mEOS4b in live human retinal pigment epithelial (hTERT-RPE-1) cells. Intriguingly, we found that mobile IFT proteins “switched gears” multiple times at the ciliary base. Our live-cell superresolution imaging revealed region-dependent slowdown of IFT proteins at the ciliary base, shedding light on staged control of intraflagellar transport.


Keywords: superresolution, sptPALM, primary cilia, live-cell imaging, intraflagellar transport