# RNA Uridylation in Trypanosomes

> **NIH NIH R56** · BOSTON UNIVERSITY MEDICAL CAMPUS · 2022 · $641,794

## Abstract

ABSTRACT
Parasitic infections by Trypanosoma brucei spp. undermine public health and economy in Sub-Saharan Africa.
These hemoflagellates are distinguished by the kinetoplast, a mitochondrial nucleoprotein body containing
interlinked maxicircle and minicircle DNA. From a basic knowledge perspective, findings of polycistronic
transcription, trans-splicing and RNA editing in trypanosomatids have profoundly influenced the conceptual fabric
of RNA biology. Accordingly, ongoing work on U-insertion/deletion mitochondrial mRNA editing has produced
pivotal breakthroughs, including discoveries of gRNA, the editosome, and the first RNA uridylating enzyme,
KRET1 TUTase. Sustained efforts by the PI and other laboratories have expanded the uridylation’s breadth from
a unique editing reaction into a compound transcriptome-shaping force prominent in most eukaryotes.
Building on recently unraveled mitochondrial and cytosolic mRNA maturation and decay pathways, and progress
in TUTase structural biology, we submit these Specific Aims:
Aim 1 investigates the TUTase–3′-5′ exonuclease nexus underpinning mitochondrial RNA biogenesis and
degradation. It focuses on mechanisms by which KRET1-containing molecular machines recognize diverse
ribonucleoprotein particles, thus governing processing, functions, and metabolic fates of their cargo RNA.
Molecular and Cryo-EM approaches will define transcript type-specific KRET1 complexes and determine their
interactomes, in vivo RNA targets and near-atomic structures. We will analyze a novel RNA helicase, KREH3,
as the potential link between massive antisense transcription of kinetoplast DNA and KRET1-mediated turnover.
Aim 2 examines structural determinants and interactions enabling KRET2 and MEAT1 to recognize double-
stranded RNA substrates, such as gRNA–mRNA hybrids. These programable mitochondrial TUTases execute
U-insertion editing, which constitutes a Kinetoplastea-specific method of genetic information transfer.
Aim 3 explores TbTUT3 and TbTUT4 as potential modulators of cytosolic mRNA decay, a principal conduit
regulating nuclear gene expression in lieu of missing transcriptional control. By assessing transcriptome-wide U-
tailed mRNAs prevalence and decay rates, we will test the coupling between mRNA 3′ uridylation and directional
clearance. Beyond the parasite RNA metabolism, we are inquiring how the divergent auxiliary modalities confer
orthogonal specificities and functions to a conserved and phylogenetically pervasive TUTase catalytic domain.

## Key facts

- **NIH application ID:** 10591650
- **Project number:** 1R56AI168191-01
- **Recipient organization:** BOSTON UNIVERSITY MEDICAL CAMPUS
- **Principal Investigator:** Ruslan Afasizhev
- **Activity code:** R56 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $641,794
- **Award type:** 1
- **Project period:** 2022-05-05 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10591650

## Citation

> US National Institutes of Health, RePORTER application 10591650, RNA Uridylation in Trypanosomes (1R56AI168191-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10591650. Licensed CC0.

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