Targeting and Biological Implications of Guanine Quadruplexes
Creators: name: Kormuth, Karen A.
Publisher: name: Carnegie Mellon University
Year: 2016
Subjects: subject: 69999 Biological Sciences not elsewhere classified
subject: FOS: Biological sciences
subject: FOS: Biological sciences

description: The multiplicity of nucleic acid structural conformations allows these molecules to drive intermolecular interactions and support essential processes within biological systems. Of these structures, the guanine (G) quadruplex has been demonstrated to function in the majority of cellular processes involving nucleic acids. Driven by a goal of developing a better understanding of the role of G-quadruplexes in ribosomal genes, we have identified new, noncanonical Gquadruplex motifs. Characterization of these structures has allowed us to learn about how they interact with synthetic nucleic acid analogs in vitro and has provided clues to the in vivo functions of these quadruplexes in yeast. Chapter 2 is a summary of our analysis of homologous, G-rich, peptide nucleic acid (PNA) hybridization to noncanonical, long-looped G-quadruplexes. We have confirmed that these PNAs, initially developed in the Armitage group, are compatible with long-looped quadruplexes. In recent years, the acceptance of noncanonical quadruplex motifs, including those with long loops, has been established within the field, enhancing the relevance of our studies. Surprisingly, we discovered that long-looped quadruplexes accommodate higher order heteroquadruplex formation with homologous PNA, wherein &gt;2 PNA strands hybridize to 1 DNA strand. In the future, we intend to explore strategies to exploit the unique features of long-looped quadruplexes to allow for the design of more selective and efficacious PNA-based probes. Chapter 3 is an expansion of previous studies in our group focused on analysis of γ-modified PNA hybridization to DNA G-quadruplex targets. We have demonstrated the compatibility of homologous, diethylene glycol-modified γPNAs with DNA G-quadruplex invasion. We have also explored PNA/DNA interactions under in vitro conditions that mimic the crowded intracellular environment, determining that molecular crowding enhances thermodynamics of PNA/DNA hybridization. These results help to confirm the applicability of similar PNAs to targets within biological systems. In Chapter 4, we initiated studies of our noncanonical G-quadruplex motifs in yeast ribosomal (r) DNA and rRNA. Our efforts also included the development of better systems with which to introduce and assay rDNA quadruplex mutations. Our results support the importance of quadruplex motifs in ribosome biogenesis, as well as the unprecedented hypothesis that specific regions of the rRNA may take on a quadruplex fold at certain points during assembly or translation. Taken together, this thesis contributes to our understanding of quadruplex biology and the interactions between these structures and synthetic PNA probes. Future development of selective and biologically compatible PNAs will be required to assist verification of quadruplex folding and function in cells. The human genome contains &gt;700,000 G-quadruplexes, which hints at the potential for exciting discoveries in G-quadruplex biology yet to come.