RNA-binding activity of TRIM25 is mediated by its PRY/SPRY domain and is required for ubiquitination

Nila Roy Choudhury; Gregory Heikel; Maryia Trubitsyna; Peter Kubik; Jakub Stanislaw Nowak; Shaun Webb; Sander Granneman; Christos Spanos; Juri Rappsilber; Alfredo Castello; Gracjan Michlewski

More than 300 novel RNA-binding proteins have been identified in HeLa (Castello et al., 2012) , HEK293 (Baltz et al., 2012) and mouse ES cells (Kwon et al., 2013) . Furthermore, a recent study identified 393 new RNA-binding proteins unique to cardiomyocytes (Liao et al., 2016) . Strikingly, many of these proteins were not previously known for their RNA-binding properties and do not contain canonical RNA-binding domains. Intriguingly, E3 ubiquitin ligases from the Tripartite Motif (TRIM) family (TRIM25, TRIM56 and TRIM71) are among these newly identified RNA-binding proteins. TRIM proteins have a characteristic architecture that includes RING zinc-finger, B-Box, coiled-coil (CC) and variable C-terminal domains. There are over 80 TRIM proteins in the human genome, and they have been divided into 11 subfamilies based on their structural composition (Versteeg et al., 2013) . The largest subfamily (C-IV) of 33 TRIM proteins is characterized by the PRY/SPRY domain at the Cterminal end. Most the proteins from the TRIM family are involved in innate immune responses (Jefferies et al., 2011; Rajsbaum et al., 2014; van Tol et al., 2017) . For example, TRIM25 plays a role in the RIG-I-mediated interferon response to viral RNAs (Ozato et al., 2008; Versteeg et al., 2013) . RIG-I recognizes viral RNA molecules with a 5'-triphosphate (5'-ppp-RNA) (Yoneyama et al., 2004) and initiates a downstream signaling cascade that culminates with the expression of type I interferons (interferon α and β ) (Loo and Gale, 2011) and an anti-viral response. TRIM25 mediates K63-linked ubiquitination of RIG-I, which leads to efficient recruitment of its downstream partners. Such recruitment in turn triggers the interferon response (Gack et al., 2007) . Recently, TRIM25 was shown to be required for the antiviral activity of RNA-binding Zinc Finger Antiviral Protein (ZAP) Zheng et al., 2017) . Almost nothing is known about the RNA-mediated roles and characteristics of these RNA-binding E3 ligases.