The function of UBEIL2, a novel ubiquitin specific activating enzyme (E1)

• Applicants: Professor Dr. Marcus Groettrup (†); Privatdozent Dr. Klaus-Peter Knobeloch
• Subject Area: Cell Biology
• Term: from 2008 to 2012
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 65246642

Ubiquitin needs to be activated at its C-terminus in order to be covalently linked to target proteins. Since the discovery of the ubiquitin specific activation enzyme UBE1 in 1981 experimental evidence has suggested that only a single El activating enzyme for ubiquitin (named UBE1) exists. While sear-ching for the El activating enzyme of the ubiquitin-like protein FAT 10 we characterized a so far barely investigated El enzyme named MOP4 or UBE1L2. This enzyme activated ubiquitin as potently as U-BE1 in vitro and in vivo and was able to transfer activated ubiquitin onto the E2 enzyme UBCHSb and further onto E3 enzymes and the substrate p53. These findings pose the question why a second El enzyme for ubiquitin has evolved and whether it serves a function that UBE1 can not fulfil. In this pro-ject we will investigate the expression levels of UBE1L2 on mRNA and protein level, we will establish a knock down of UBE1L2 and determine its effects on protein degradation, polyubiquitylation, proliferation, survival, radiation resistance, and stress resistance in cells. In addition, we will try to identify ubiquitylation substrates in UBE1 deficient cells and investigate if these rely on UBE1L2 to become ubiquitylated. Moreover, proteins that accumulate in UBE1L2 deficient cells will be investiga-ted by 2D-DIGE technology. A yeast two hybrid screen will be performed with UBE1L2 and the UBE1L2 specific E2 enzyme Usel in order to find E2 enzymes that bind to UBE1L2 and E3 enzymes that interact with Usel. Finally we will generate organ specific and inducible knock out mice in order to elucidate the in vivo functions of UBE1L2.

• DFG Programme: Research Grants