The role of histone 2B monoubiquitination (H2Bub1) in HER2-driven mammary carcinoma
by Evangelos Prokakis née Evangelos
Date of Examination:2021-01-15
Date of issue:2021-03-25
Advisor:Dr. Florian Wegwitz
Referee:Prof. Dr. Matthias Dobbelstein
Referee:Prof. Dr. Jürgen Wienands
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Description:Doctoral Dissertation of Evangelos Prokakis
Abstract
English
Breast cancer (BC) is the most frequent tumor entity in women worldwide with a high chance of therapeutic response in early- and non-metastatic disease stage. Despite the advent of HER2-specific targeted therapies, a considerable fraction of HER2 (Human Epidermal Growth Factor Receptor 2) positive breast cancer (HER2+-BC) patients become refractory or acquire resistance to these therapies. These patients show a poor prognosis, hence, necessitating a better characterization of the molecular pathology of this malignancy. Accumulating studies indicate that cancer disease does not exclusively result from genetic alterations but can be fostered by aberrant epigenetic changes in the physiological gene expression program of a healthy tissue. Cancer cells benefit from the speed and reversible nature of these epigenetic alterations, empowering a rapid adaption to the gene expression program in response to adverse intra- or extracellular conditions. At the same time, this reversibility broadens the avenue to therapeutically interfere with acknowledged epigenetic dependencies in various malignancies. Monoubiquitination of lysine 120 of histone 2B (H2Bub1) is a transcription-promoting histone post-translational modification (hPTM). Gradual loss of H2Bub1 has been numerous times linked to cancer progression in many malignancies including BC. Consequently, H2Bub1 was suspected to harbor universal tumor-suppressive properties. However, its biological function in BC has only been scarcely addressed in vivo with the help of xenograft approaches. To uncover the consequences of H2Bub1 imbalance in BC, we performed a combined approach whereby we compromised or augmented the global H2Bub1 levels by impairing the expression of RNF40 or USP22, the major E3 ligase and deubiquitinase (DUB) of this hPTM, respectively. Specifically, we leveraged the HER2+-BC mouse model MMTV-Erbb2 as well as human HER2+-BC cell lines to shed light on the functional role of the H2Bub1 epigenetic machinery in cancer progression and to underpin its underlying mechanisms. Against our initial expectations, we revealed that tumor-specific deletion of Rnf40 led to a substantially decreased tumor burden in MMTV-Erbb2 mice compared to their control counterparts. Accordingly, we identified that RNF40 silencing impairs the oncogenic properties in human HER2+-BC cell lines. Mechanistically, we discovered an unknown role of the RNF40-mediated H2Bub1/H3K4me3 trans-histone crosstalk in driving the actin cytoskeleton (AC) dynamics and the activity of the downstream focal adhesion kinase (FAK)-driven pathway. Therefore, our data strongly suggest that the RNF40/H2Bub1/H3K4me3 epigenetic axis plays a rather tumor-supportive than -suppressive role in HER2+-BC by controlling the AC/FAK axis to sustain the aggressive behavior of this malignancy. In parallel, the H2Bub1-specific DUB USP22 was multiple times reported to harbor oncogenic properties, among other, in BC in vitro. Specifically, using the same experimental settings, we surprisingly uncovered a global H2Bub1-independent involvement of USP22 in the tumorigenic phenotype of HER2+-BC. Specifically, we confirmed that USP22 actively suppresses the unfolded protein response (UPR) by deubiquitinating and, thereby stabilizing, the ER-chaperone HSPA5 in vivo and in vitro. Accordingly, loss of USP22 activated the apoptotic cascade and significantly increased the efficiency of therapies targeting the ER folding capacity. Consequently, our data suggest that USP22 is not only a promising therapeutic target to combat HER2+-BC, but can substantially improve UPR-based therapeutic schemes that dysregulate proteostasis pathways for cell survival. Collectively, our data introduce novel functions of the RNF40/H2Bub1/H3K4me3 axis in this aggressive BC subtype by significantly updating the current knowledge we possess for the context-dependent role of this axis. Finally, this work sheds light on a new pro-tumorigenic function of USP22 in the suppression of UPR signaling and in the maintenance of the global chaperoning system that protects tumor cells from proteostasis imbalance.
Keywords: HER2; breast cancer; RNF40; USP22; H2Bub1; actin cytoskeleton; UPR