Structure and activation of the RING E3 ubiquitin ligase TRIM72 on the membrane

Abstract

Defects in plasma membrane repair can lead to muscle and heart diseases in humans. Tripartite motif-containing protein (TRIM)72 (mitsugumin 53; MG53) has been determined to rapidly nucleate vesicles at the site of membrane damage, but the underlying molecular mechanisms remain poorly understood. Here we present the structure of Mus musculus TRIM72, a complete model of a TRIM E3 ubiquitin ligase. We demonstrated that the interaction between TRIM72 and phosphatidylserine-enriched membranes is necessary for its oligomeric assembly and ubiquitination activity. Using cryogenic electron tomography and subtomogram averaging, we elucidated a higher-order model of TRIM72 assembly on the phospholipid bilayer. Combining structural and biochemical techniques, we developed a working molecular model of TRIM72, providing insights into the regulation of RING-type E3 ligases through the cooperation of multiple domains in higher-order assemblies. Our findings establish a fundamental basis for the study of TRIM E3 ligases and have therapeutic implications for diseases associated with membrane repair.

Deciphering the role of the hendecad-repeat coiled-coil domain of TRIM72 in membrane curvature recognition

“…Details of sample preparation, data collection, and parameters for the SAXS experiments used have been described in a previous report 33 . Ab initio structure determination was performed using DAMMIN software in ATSAS online (https://www.embl-hamburg.de/biosaxs/atsas-online/).…”

“…Details of sample preparation and cryo-TEM data collection methods used have been described in a previous report 33 . For cryo-experiments, protein-80 mol% PS-SUV complexes were loaded onto glowdischarged Quantifoil grids and frozen in a plunge freezer with a Vitrobot (Thermo Fisher Scienti c, Waltham, MA, USA).…”

“…Furthermore, recombinant TRIM72 has shown therapeutic potential for a variety of muscle and non-muscle tissue injuries 36 . In previous studies, the oligomerization of TRIM72 was shown to be a key process for membrane repair and activation 31,33 . TRIM72 facilitates negatively charged small repair vesicles to injury sites on the plasma membrane.…”

“…Despite the increasing number of genetic and cellular studies, biochemical and structural evidence are still limited, and only domain structures are available 26,27,28,29,30,31,32 . We recently presented dimeric and oligomeric structures of TRIM72, in which oligomerization was found to be coupled to ubiquitylation and phospholipid membrane recognition 33 . TRIM72, also known as mitsugumin 53 (MG53), is expressed mainly in muscles 34,35,36 and is a key initiator of the plasma membrane repair machinery following acute membrane damage, such as that induced by ischemia/reperfusion injury 2,37,38 .…”

“…TRIM72 facilitates negatively charged small repair vesicles to injury sites on the plasma membrane. The TRIM72-bound repair vesicles are assumed to accumulate at the injury site where they form a membrane patch to seal holes in the plasma membrane 33,39 . In addition to membrane repair, TRIM72 participates in diverse cellular processes that occur in the membrane environment, such as lopodia-like structures, exocytosis, caveolar structures, and lipid rafts 34,40,41 .…”