Live cell assays to identify regulators of ER to Golgi trafficking

• Autoři: LISAUSKAS Tautvydas; MATULA Petr; CLAAS Christoph; REUSING Susanne; WIEMANN Stefan; ERFLE Holger; LEHMANN Lars; FISCHER Peter; EILS Roland; ROHR Karl; STORRIE Brian; STARKUVIENE Vytaute
• Rok publikování: 2012
• Druh: Článek v odborném periodiku
• Časopis / Zdroj: Traffic
• Fakulta / Pracoviště MU: Fakulta informatiky
• 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.

Související projekty

• Vysoce paralelní a distribuované výpočetní systémy
• Vytipování markerů, screening a časná diagnostika nádorových onemocnění pomocí vysoce automatizovaného zpracování multidimenzionálních biomedicínských obrazů