Browsing by Subject "circular RNA"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Mitochondrial mRNA termini processing in Trypanosoma brucei
    (2023-08) Smoniewski, Clara
    A large and ever-expanding field, RNA biology is the study of the RNA lifecycle, how RNAs function, and their diverse roles in cellular processes. The most well-known RNA type are the protein-coding mRNAs transcribed from genes encoded in DNA. mRNAs progress through multiple steps during their lifecycle, one of which is maturation in which a 3' nonencoded nucleotide tail is added. These tails vary widely across organisms and organelles, both in composition and function. This dissertation examines the 3' tail addition and termini processing of mRNA transcripts in the mitochondrion of a parasitic protozoan Trypanosoma brucei. Trypanosomes have a single mitochondrion in which RNA regulation is crucial for gene expression changes necessary for host-switching. Although a working model for mitochondrial mRNA processing in T. brucei has been developed, transcript-specific gene regulation remains undefined. Differences in mRNA editing, stability, and translation remain unexplained, as do differences in 3' mRNA tail characteristics between transcripts and life stages. Here, I present several studies of the RNA lifecycle and mRNA termini processing in the mitochondrion of T. brucei. Chapter 1 contains an introduction to RNA, 3' mRNA tails, and trypanosomes. In Chapter 2, I explain our refinement of circTAIL-seq, a tool used to high-throughput sequence 3' and 5' termini. I define the circTAIL-seq error rate and describe a newly developed alignment protocol that allows for greater nuance in aligning reads. The discovery and characterization of circular mitochondrial mRNAs (circRNAs) is presented in Chapter 3. We show that circRNAs are present in both T. brucei life stages and are a subpopulation of total mRNAs with distinct characteristics. Chapter 4 contains a description of the effects of manipulating two poly(A) polymerase (PAP)-family proteins found in T. brucei’s mitochondria — KPAP1 and KPAP2. KPAP2 is found in unknown complexes of various sizes, and mutations of KPAP1 arginine methylation sites are shown to have transcript-specific effects on 3' tail characteristics. I then summarize my major conclusions and present a discussion of limitations and future directions of this work. Overall, this dissertation expands our understanding of the RNA lifecycle by further elucidating mitochondrial mRNA termini processing in T. brucei.
  • Thumbnail Image
    Item
    A novel bioinformatics approach to characterize and integrate messenger RNAs, circular RNAs and micro RNAs
    (2018-04) Nair, Asha
    High-throughput Next Generation RNA sequencing (RNA-Seq) technology is affluent with information about the transcriptome, which includes both protein-coding and multiple non-coding regions. In a diseased state, complex interactions between these regions can go awry. Identification of such interactions is critical to translate the underlying biology of the transcriptome, especially for lethal diseases such as cancer. The field of bioinformatics is currently deficient in workflows that can analyze both coding and non-coding regions together efficiently, to identify disease-specific interactions. In this dissertation, I developed three coherent bioinformatics solutions that aim to address these shortcomings in RNA-Seq. First, a comprehensive workflow called MAPR-Seq was developed to analyze and report various features of protein-coding messenger RNAs. Second, a workflow for non-coding circular RNAs, called Circ-Seq, was developed to identify, quantify and annotate expressed circular RNAs. Third, an integration workflow called ReMIx was developed to identify microRNA response elements (MREs) and integrate them with the different types of RNAs (messenger RNAs, circular RNAs, and microRNAs). Collectively, the three workflows were applied to the largest cohort of breast cancer samples (n=885) from The Cancer Genome Atlas (TCGA). Based on the results obtained from these workflows, I present several key findings that are pertinent to breast cancer. I show that circular RNAs may be a marker for tumor proliferation in estrogen response positive (ER+) breast cancer subtype. I also show how triple negative (TN) breast cancer subtype-specific MRE signatures of messenger RNA – microRNA interactions can be obtained using RNA-Seq data, which has not been explored to date and thus, is a novel undertaking. In the end, my results highlight candidate messenger RNAs, circular RNAs and microRNAs that are found to be associated with MAPK and PI3K/AKT signaling cascades in TN breast cancer subtype. In general, the developed bioinformatics solutions can also be applied to RNA-Seq data of other cancer subtypes and diseases to identify unique messenger RNA – microRNA – circular RNA candidates that could be promising diagnostic targets towards improving treatment options for complex diseases.