SELECTIVE DEGRADATION OF HUMAN TIP110/SART3 IN A TRANEGNIC MOUSE MODEL: ROLE OF UBIQUITIN SPECIFIC PEPTIDASE 15

Purpose: HIV Tat-interacting-protein 110 (Tip110) is an RNA binding protein. It is highly elevated and expressed in the nucleus of all malignant tumor cell lines tested and in the majority of cancer tissues. In addition, it regulates or interacts with several oncogenic proteins. Thus, the purpose of this study is to understand the regulatory pathways of Tip110 degradation. Methods: Transgenic human (Tg-h-Tip110) mice were created. Mouse embryonic fibroblast (MEF) cells were obtained from wild-type and Tip110 transgenic (Tg) embryos. Mass Spectroscopy, immunoprecipitation and immunofluorescence assays were performed to study the host cellular proteins interacting with Tip110. To study the physiological significant of this interaction, an in vivo deubiquitination assay was performed. Results: The Tg-h-Tip110 mice did not show any abnormalities during mouse development. Surprisingly, Tg-h-Tip110 protein was not detectable in different Tg tissues although Tg-h-Tip110 mRNA was transcribed. Treatments of Tg-MEF with proteasomal inhibiters led to detection of the Tg-h-Tip110 protein, indicating that loss of Tg-h-Tip110 in mice was due to degradation through the ubiquitin proteasomal pathway. Interestingly, h- but not m-Tip110 protein underwent selective degradation in mouse cells. This difference appeared to be consistent with the finding that h-Tip110 exhibited more ubiquitination modification in comparison to m-Tip110. Our results showed that Tip110 interacted with deubiquitionation enzyme ubiquitin specific peptidase 15 (USP15) and the expression of Tip110 resulted in the translocation of USP15 from the cytoplasm to the nucleus where they co-localized. In vivo deubiquitination assay showed that Tip110 served as a substrate for USP15 and that deubiquitination of h-Tip110 is more pronounced than m-Tip110. Unexpectedly, knock-down but not overexpression of USP15 in Tg-MEF resulted in the accumulation of Tg-h-Tip110. Conclusions: Here, we showed that h-Tip110 protein is degraded in transgenic mice through the ubiquitin proteasomal pathway. We identified the USP15 as an interacting partner with Tip110. Interestingly, the finding that the deubiquitination activity by USP15 protein was found to be more efficient for h-Tip110 in comparison to the m-Tip110 corroborated the degradation behavior of h-Tip110 in transgenic mice. These results revealed that the Tip110 protein level in tissues is under tight control, suggest that Tip110 interaction with USP15 may be responsible for its selective and rapid turnover.