HUWE1 controls tristetraprolin proteasomal degradation by regulating its phosphorylation

Scinicariello, S.; Soederholm, A.; Schaefer, M.; Shulkina, A.; Schwartz, I.; Hacker, K.; Gogova, R.; Kalis, R.; Froussios, K.; Budroni, V.; Bestehorn, A.; Clausen, T.; Kovarik, P.; Zuber, J.; Versteeg, G. A.

doi:10.1101/2022.08.29.505645

bioRxiv · 10.1101/2022.08.29.505645

HUWE1 controls tristetraprolin proteasomal degradation by regulating its phosphorylation

Scinicariello, S.·Soederholm, A.·Schaefer, M.·Shulkina, A.·Schwartz, I.·Hacker, K.·Gogova, R.·Kalis, R.·Froussios, K.·Budroni, V.·Bestehorn, A.·Clausen, T.·Kovarik, P.·Zuber, J.·Versteeg, G. A.

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Abstract

Tristetraprolin (TTP) is a critical negative immune regulator. It binds AU-rich elements in the untranslated-regions of many mRNAs encoding pro-inflammatory mediators, thereby accelerating their decay. A key but poorly understood mechanism of TTP regulation is its timely proteolytic removal: TTP is degraded by the proteasome through yet unidentified phosphorylation-controlled drivers. In this study, we set out to identify factors controlling TTP stability. Cellular assays showed that TTP is strongly lysine-ubiquitinated, which is required for its turnover. A genetic screen identified the ubiquitin E3 ligase HUWE1 as a strong regulator of TTP proteasomal degradation, which we found to control TTP stability indirectly by regulating its phosphorylation. Pharmacological assessment of multiple kinases revealed that HUWE1-regulated TTP phosphorylation and stability was independent of the previously characterized effects of MAPK-mediated S52/S178 phosphorylation. HUWE1 function was dependent on phosphatase and E3 ligase binding sites identified in the TTP C-terminus. Our findings indicate that while phosphorylation of S52/S178 is critical for TTP stabilization at earlier times after pro-inflammatory stimulation, phosphorylation of the TTP C-terminus controls its stability at later stages.

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Scinicariello, S., Soederholm, A., Schaefer, M., Shulkina, A., Schwartz, I., Hacker, K., Gogova, R., Kalis, R., Froussios, K., Budroni, V., Bestehorn, A., Clausen, T., Kovarik, P., Zuber, J., Versteeg, G. A.. 2022-08-29. HUWE1 controls tristetraprolin proteasomal degradation by regulating its phosphorylation. https://doi.org/10.1101/2022.08.29.505645

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