Live cell assays to identify regulators of ER to Golgi trafficking
| Autoři | |
|---|---|
| Rok publikování | 2012 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | Traffic |
| Fakulta / Pracoviště MU | |
| Citace | |
| Doi | http://dx.doi.org/10.1111/j.1600-0854.2011.01318.x |
| Obor | Morfologické obory a cytologie |
| Klíčová slova | BFA; GalT; ER to Golgi trafficking; YIPF; GOT1B; USE1; SACM1L |
| Popis | We used fluorescence microscopy based quantitative assays in living cells to identify regulators of ER to Golgi trafficking and/or Golgi complex maintenance. We first validated an automated procedure to identify factors, which influence Golgi to ER re-localization of GalT-CFP after BFA addition and wash-out. We then tested 14 proteins that potentially play a role in ER to Golgi trafficking, and localize to the ER and/or Golgi complex when over-expressed. 9 of them interfered with the rate of BFA induced redistribution of GalT-CFP to the ER, 6 of them interfered with GalT-CFP reassembly rate to a juxtanuclear region after BFA wash-out, and 6 of them were positive effectors in both assays. Notably, our live cell approach captures functions of those regulators of ER to Golgi trafficking, which were missed in previous fixed cell assays; as well as assigns respective roles for yet incompletely characterized proteins. Moreover, the assays can be extended to work under the conditions of RNAi and for testing chemical compounds. |
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