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This work was carried out in the lab of Dr Andrew Turnell (University of Birmingham)

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The transcriptional repressor, E3 ubiquitin ligase and putative tumour suppressor TIF1γ/TRIM33/Ectodermin is involved in embryological development and haematopoiesis. It antagonises the TGF-β pathway by preventing downstream signalling through SMAD4. Recent observations suggested that absolute cellular levels of TIF1γ fall dramatically post-infection with different adenoviral serotypes, including Ad5 and Ad12. Given the functional similarities between the usurpation of the cellular machinery by adenovirus and the cellular reprogramming that occurs in cancer, it was considered useful to investigate adenoviral regulation of TIF1γ.

Western blot findings confirmed a reduction in cellular TIF1γ levels following Ad5 or Ad12 infection. TIF1β/TRIM28 levels did not fall, however, suggesting selective regulation of TIF1γ. TIF1γ interacted with the adenoviral E1B-55kDa oncoprotein in adenoviral transformed cells and immunofluorescence microscopy implied that they co-localised in cytoplasmic aggresome-like structures.

As E1B-55kDa and E4orf3 can cooperatively mediate protein degradation, further work examined the role of Ad5 E4orf3 in the regulation of TIF1γ. Confocal microscopy revealed that TIF1γ co-localised with PML in nuclear tracts in adenovirus-infected cells. Similar nuclear co-localisation of TIF1γ with Ad5 E4orf3 was shown 18 hours post-infection and with Ad12 E4orf3 in transfected HeLa cells.

E4orf3- mutant Ad5 infection failed to abate TIF1γ levels, implying a necessary role for E4orf3 in the process. Interestingly, the adenoviral E3 ubiquitin ligase remained functional as p53 was still degraded. Ad5 E4orf3 transcriptional repression of p53-dependent gene expression was shown not to be required to attenuate TIF1γ levels.

To identify novel E4orf3-interactors, immunoprecipitation followed by mass spectrometry was performed. The data suggested UBE4A as a promising potential regulator of TIF1γ.

To conclude, E4orf3 plays a necessary role reducing cellular TIF1γ levels during Ad5 infection, although it remains to be seen if this phenomenon involves the Ad5 E3 ubiquitin ligase or a novel mechanism.

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Dr Pól Roibeárd Ó Catnaigh