Plants use light as a signal to degrade transcription factors altering gene expression to maximise their photosynthetic capabilities. Protein degradation is typically initiated by the ubiquitin ligase pathway consisting of E1, E2 and E3 enzymes that modify substrates with ubiquitin to target them to the proteosome. The ubiquitin-conjugating E2 enzyme COP10 is unique to plants and lacks a key active site residue that renders it catalytically inactive. COP10 is essential for the ubiquitination and subsequent degradation of transcription factors and ultimately the repression of photomorphogenesis. To understand how COP10 is able to manipulate protein regulation via the ubiquitin ligase pathway, we have expressed, purified and crystalised COP10 from which we have obtained a 1.26Å structure. This structure revels a typical UBC-domain with an intact ubiquitin binding pocket and another possible protein interaction site on its backside β-sheet surface.
COP10 is a member of the CDDD complex (COP10, DDB1, DET1 and DDA1) referred to as a “photomorphogenic switch”. Surprisingly the inactive COP10 is required for normal light-dependent gene regulation via the CDDD complex and the ubiquitin-proteasome system. It is understood that COP10 can enhance the activity of other E2 enzymes, however it is unclear if this activity is essential in the CDDD complex or if COP10 has additional roles independent of the complex. Recently we heterologously expressed and purified the Arabidopsis CDDD complex from Trichoplusia ni cells. This enabled affinity-purification of the CDDD complex, regular structures could be visualised by negative stain. Ongoing experiments include further structural studies including sample preparation for cryo-electron microscopy and ubiquitination assays aiming to uncover a complete mechanism for ubiquitin-based regulation of photomorphogenesis.