# Evolution, composition and functions of cullin E3 ubiquitin ligases in trypanosomes

## TL;DR

This study explores the evolution and function of cullin E3 ubiquitin ligases in trypanosomes, revealing their complexity and role in protein regulation.

## Contribution

The paper provides new insights into the evolutionary history and functional diversity of cullin-RING ligases in trypanosomatids.

## Key findings

- Cullin-RING ligases show considerable diversity and at least four ancestral subfamilies in eukaryotes.
- Expansions and novel client adaptors in trypanosomatids suggest increasing complexity in adapter architecture.
- TbCul-A/CUL-1 mediates the turnover of ornithine decarboxylase, a key drug target in trypanosomes.

## Abstract

Post-translational modifications (PTMs) modulate protein functions, with ubiquitylation a pre-eminent example, and playing major roles in protein turnover. Ubiquitylation utilises a ligase enzyme cascade for conjugation of ubiquitin to client proteins, of which there are a large number in humans and lesser numbers in unicellular eukaryotes. The Cullin-RING ligases are amongst the most complex ligase subfamily and are present across the eukaryote lineage. We have reconstructed the evolution of cullin-RING E3 ubiquitin ligases across eukaryotes and experimentally determined the composition of six of seven cullin complexes in trypanosomatids. We find considerable diversity within cullins and reconstruct at least four ancestral pan-eukaryotic subfamilies. Furthermore, we identify expansions of cullin client adaptor protein families, novel client adaptors and demonstrate client specificity in trypanosomatids. We also find evidence for increasing complexity within client adaptors, suggesting ongoing expansion of adapter architecture. Finally, we show that turnover of ornithine decarboxylase (TbODC), an important target of the trypanocide eflornithine, is mediated by TbCul-A/CUL-1. These studies highlight lineage-specific aspects of cullin E3 ligases and their contributions towards eukaryotic complexity.

The online version contains supplementary material available at 10.1038/s41598-025-32077-9.