Abstract 2900: PRAMEF2 ubiquitylates and degrades LATS1 leading to enhanced nuclear accumulation of YAP and tumorigenesis

Abstract

Abstract PRAMEF2 (PRAME Family Member 2) is a Cancer Testis Antigen belonging to the PRAME multi-gene family. All Cancer Testis Antigens are expressed in male testicular germ cells while remaining absent from other adult tissues. Conversely, they show aberrant re-expression in several human malignancies and also elicit immunogenic responses in cancer patients. This makes Cancer Testis Antigens and thereby PRAMEF2, effective and novel targets for cancer vaccine development. However, the physiological function of PRAMEF2 underlying tumorigenesis remains unspecified. Hence, we performed a proteomic screen to explore its unique interactome in the cell. Subsequent size exclusion chromatography and co-immunoprecipitation assays confirmed the role of PRAMEF2 as a BC-box containing substrate recognition subunit of a Cullin 2 based-E3 ubiquitin ligase complex. In the proteomic screen, several proteins were identified, however in-vivo assays revealed LATS1 to be a specific and novel interacting partner of PRAMEF2. LATS1 is a Ser/Thr Kinase of the Hippo Pathway, known to aid tumor suppression. We observed PRAMEF2 mediated ubiquitylation and subsequent proteasomal degradation of LATS1 through ubiquitylation assay. The site for ubiquitylation was mapped to the highly conserved Lysine 860 residue on LATS1. Upon activation of the Hippo Pathway, LATS1 results in phosphorylation of the transcriptional co-activator YAP, leading to its cytosolic translocation. Consequently, PRAMEF2 mediated LATS1 regulation resulted in decreased YAP phosphorylation and its increased accumulation in the nucleus. Furthermore, RTqPCR array and ChIP analysis showed the enhanced expression of YAP target genes implicated in proliferation and metastasis. Moreover, PRAMEF2 depletion also diminished the proliferative capacity, migration potential and invasiveness of tumor cells. Studies in mouse tumor models confirmed that PRAMEF2 abrogates the tumor suppressive function of LATS1 leading to ameliorated tumor growth and metastasis. Taken together, our findings help establish the role of PRAMEF2, an unique Cancer Testis Antigen, as a substrate recognition subunit of a Cullin 2- based E3 ubiquitin ligase complex which ubquitylates and degrades LATS1. Hence, PRAMEF2 realizes its oncogenic potential by increasing YAP nuclear accumulation and promoting tumor cell proliferation and metastasis. Citation Format: Madhurima Ghosh, Sanjeev Das. PRAMEF2 ubiquitylates and degrades LATS1 leading to enhanced nuclear accumulation of YAP and tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2900.