Study on the Roles of RNF2 and UCHL5 in Wnt/ß-catenin Signaling

Abstract

Wnt/ß-catenin signaling is a fundamental pathway that plays a crucial role in various developmental processes, tissue homeostasis, and cell fate determination. It involves the regulation of β-catenin, a key protein that acts as a transcriptional co-activator and is tightly controlled by a destruction complex. Hence, the complete understanding of its molecular mechanism has been the biggest challenge in the Wnt research field. In this study, we identified two key regulators. Firstly, RNF2 was found to regulate TCF7L1, a transcription factor known for its negative function. Secondly, UCHL5 was discovered to regulate Axin1, which is a crucial component of the destruction complex.

1. RNF2 regulates Wnt/ß-catenin signaling via TCF7L1 destabilization TCF7L1, a transcription factor, serves as a critical mediator of canonical Wnt signaling. TCF7L1 interacts with β-catenin and binds to Wnt target gene promoters, thereby initiating transcriptional activation or repression, which ultimately influences cell fate determination and tissue homeostasis. TCF7L1 is known to possess an inhibitory function; however, the underlying regulatory mechanism remains unclear. In this study, we identified the E3 ligase, RNF2, as a novel positive regulator of the Wnt pathway. Here, we demonstrate that RNF2 promotes the degradation of TCF7L1 through its ubiquitination upon the activation of Wnt signaling. Loss-of-function studies have shown that RNF2 consistently destabilizes nuclear TCF7L1 and is required for proper Wnt target gene transcription in response to Wnt activation. Furthermore, our results revealed that RNF2 controls the threshold, persistence, and termination of Wnt signaling by regulating TCF7L1. Overall, our study sheds light on the previously unknown degradation mechanism of TCF7L1 by a specific E3 ligase, RNF2, and provides new insights into the variability in cellular responses to Wnt activation.

2. UCHL5 controls ß-catenin destruction complex function through Axin1 regulation Axin1 is a crucial component of the β-catenin destruction complex, which plays a vital role in the canonical Wnt signaling pathway. As a scaffold protein, Axin1 acts as a key regulator by facilitating the assembly of the destruction complex and promoting the degradation of β-catenin. By controlling the levels of β-catenin, Axin1 regulates the activation of downstream target genes involved in cellular processes such as development, proliferation, and differentiation. Here, we identified ubiquitin C-terminal hydrolase like 5 (UCHL5), a deubiquitinating enzyme, as a novel negative regulator of Wnt signaling, upstream of ß-catenin. The study further revealed that UCHL5 plays an important role in the ß-catenin destruction complex, as it physically interacts with multiple domains of Axin1 protein. Our functional analyses also elucidated that UCHL5 is required for both the stabilization and the polymerization of Axin1 proteins. Interestingly, although these events are governed by deubiquitination in the DIX domain of Axin1 protein, they do not require the deubiquitinating activity of UCHL5. The study proposes a novel molecular mechanism of UCHL5 potentiating the functional activity of Axin1, a scaffold protein of the ß-catenin destruction complex.