# A cytidine deaminase-like protein modulates pyrimidine nucleotide homeostasis in Trypanosoma brucei

## TL;DR

A unique protein in a parasite helps control nucleotide levels, affecting DNA replication and cell survival, and could be a target for treating trypanosomiasis.

## Contribution

Identifies a trypanosome-specific cytidine deaminase-like protein as a key regulator of pyrimidine nucleotide homeostasis and a potential drug target.

## Key findings

- Overexpression of TbCDA-like reduces dCTP and CTP while increasing dTTP and UTP levels.
- TbCDA-like depletion causes CMP accumulation and reduced dTTP, leading to DNA damage and growth arrest.
- TbCDA-like localizes to stress granules under nutrient deprivation, linking it to RNA metabolism.

## Abstract

Pyrimidine nucleotide homeostasis is critical for DNA replication and cell viability, yet its regulation in Trypanosoma brucei, the causative agent of African trypanosomiasis, remains poorly understood. Here, we characterize a T. brucei cytidine deaminase-like protein (TbCDA-like), a kinetoplastid-specific enzyme absent in mammals, that harbors a deaminase domain and a zinc-finger CCCH motif. Using RNA interference (RNAi) and overexpression approaches, we demonstrate that TbCDA-like modulates pyrimidine nucleotide pools, influencing both ribonucleotide and deoxyribonucleotide profiles. Overexpression of TbCDA-like resulted in a substantial reduction of dCTP and CTP levels while elevating dTTP and UTP pools, suggesting a role in cytidine derivative deamination. Conversely, RNAi-mediated depletion of TbCDA-like caused CMP accumulation and reduced dTTP levels. Notably, overexpression induced severe cytotoxicity, growth arrest, DNA damage and cell cycle defects, evidenced by sub-G1 populations, increased nuclear H2A phosphorylation and aberrant kinetoplast and nuclear morphologies. Localization studies revealed that TbCDA-like is primarily cytosolic and relocalizes to stress granules upon nutrient deprivation, suggesting a role in RNA metabolism. These findings establish TbCDA-like as a key regulator of pyrimidine nucleotide homeostasis in T. brucei, linking nucleotide imbalances to DNA replication stress and genome instability. Given its absence in mammals, TbCDA-like presents an attractive target for therapeutic intervention against trypanosomiasis.

The online version contains supplementary material available at 10.1038/s41598-025-00942-2.