New work defines molecular players and principles of attaching UFM1 to ribosomes

Published on 14 October 2022

Exciting new work just published by the Kulathu Lab describes the molecular players and biochemical principles that govern attachment of UFM1 onto ribosomes.

On this page

Joshua Peter

Discovered only in 2004, UFM1 is the most recently identified Ubiquitin-like protein. Like ubiquitin, UFM1 is post-translationally attached to proteins via an enzymatic cascade. Ribosomes located at the endoplasmic reticulum, the large molecular machines that carry out protein synthesis, are the main cellular targets of UFM1 attachment. This modification, UFMylation, is crucial for membrane and secretory protein biogenesis and failure in UFMylation has been linked to several developmental and neurological disorders. While UFM1 is important for cellular homeostasis, the mechanisms of how UFM1 is attached onto protein targets have remained enigmatic.

Exciting new work just published by the Kulathu Lab describes the molecular players and biochemical principles that govern attachment of UFM1 onto ribosomes.

This work led by Joshua Peter, a PhD student in Yogesh Kulathu’s lab employed a reductionist approach that involved rebuilding UFMylation in a test tube using purified components. Using this approach, he found that the described E3 ligase UFL1 is inactive on its own. He uncovered two additional factors namely UFBP1 and CDK5RAP3, which bind to UFL1. Surprisingly, UFL1 and UFBP1 together form a functional ligase complex capable of modifying ribosomes. To understand the mechanism, they used the Artificial Intelligence-based structure prediction program AlphaFold (Deepmind) which showed that UFL1 and UFBP1 were made up of several Winged Helix Binding (WHB) domains that are generally found in transcription factors and lacked structural elements typically found in E3 ligases. In addition, this structural prediction revealed a unique mechanism for the assembly of UFL1 and UFBP1 into a complex where two half WHB domains from each protein come together to form a composite domain. The second factor called CDK5RAP3 functions to inhibit the ligase complex in the absence of its substrate, the ribosome. In the presence of the ribosome, this inhibition is relieved and CDK5RAP3 is thought to function as a specificity factor restricting the activity of the E3 ligase complex towards ribosomes. Hence CDK5RAP3 may work like an on-off switch to regulate activity and specificity of the ligase complex.

Diagram showing the work involved rebuilding UFMylation in a test tube


This work also uncovered novel sequence motifs and structural features conserved in the E2 enzyme UFC1 that are crucial for catalyzing UFMylation. To highlight one, a biallelic mutation on UFC1 (T106I) was earlier discovered in patients with early onset encephalopathy. While the exact functional consequence of this mutation remained unclear, by using the fully reconstituted in vitro UFMylation assays, this work identifies that this mutation abolishes UFMylation of ribosomes, which may potentially explain the underlying cause of the disease.

Overall, this work published in the EMBO Journal defines the minimal requirements, regulatory principles and roles of the molecular players involved in the UFMylation pathway and lays the foundation for future biochemical and structural studies to understand the mechanism of this enigmatic modification system.

This work was supported by the European Research Council (ERC), UKRI BBSRC and MRC, and the Lister institute of preventive medicine.

Story category Research