Kinetoplastid organisms include the parasites Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. which cause the neglected tropical human diseases Sleeping Sickness (aka Human African Trypanosomiasis), Chagas? disease, and Leishmaniasis, respectively. A common feature of these parasites is their streamlined genomes. Protein-coding genes are arranged in dense directional arrays in which the reading frames are typically separated by only a few hundred base pairs. In addition, these genes have undisrupted reading frames, e.g. do not encode introns. Transcriptome studies determined that the genome of T. brucei harbors only two introns. One intron disrupts the PAP1 gene which encodes a specialized poly(A) polymerase that is involved in the maturation of small nucleolar (sno)RNAs. snoRNAs are essential for ribosome biogenesis, guiding processing and post-transcriptional modification of ribosomal RNA. The second intron is located in the DPB2B gene which encodes a DEAD box RNA helicase. While such helicases are important factors in several molecular processes including transcription, RNA splicing and ribosome biogenesis, the specific function of DBP2B is not yet known. We found that all kinetoplastids analyzed, including the distantly related Bodo saltans, encode PAP1 and DBP2B orthologs with the intron present in a conserved position, disrupting the same codon in each gene. Since it is generally accepted that all branches of extant eukaryotes originated from intron-rich progenitors, the question arises why these introns have been conserved over an estimated 500 million years of evolutionary time in kinetoplastids and have not been eliminated as other introns. We predict that the introns are fundamentally important, preventing kinetoplastids, free-living or parasitic, to eliminate them from their genes. We will investigate the basic roles of the two introns by an array of genetic manipulations. Preliminary data suggest that the PAP1 intron negatively impacts the expression of its gene, raising the possibility that uncontrolled or overexpression of this gene affects parasite fitness. For DBP2B we also strive to understand the helicase?s specific function which will be important in evaluating the intron?s regulatory role. Given the fact that we could not find a kinetoplastid genome without the genes and introns present, this project may [eventually] provide an intervention target against which it will be difficult for the parasites to develop resistance.

Public Health Relevance

Kinetoplastid parasites cause devastating human diseases, affecting millions of people worldwide. Given the rising resistance of parasites against available drugs, new anti-parasitic strategies are needed. As a new line of investigation and based on the finding that these organisms have retained in their genomes [only] the same two introns (non-coding DNA sequences that disrupts the coding region of a gene) for ~500 million years of evolutionary time, this project is to identify the anticipated fundamentally important roles of these introns.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21AI142149-01
Application #
9651461
Study Section
Pathogenic Eukaryotes Study Section (PTHE)
Program Officer
Mcgugan, Glen C
Project Start
2018-11-16
Project End
2020-10-31
Budget Start
2018-11-16
Budget End
2019-10-31
Support Year
1
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Connecticut
Department
Genetics
Type
Schools of Medicine
DUNS #
022254226
City
Farmington
State
CT
Country
United States
Zip Code
06032