Poster abstracts

Poster number 69 submitted by Sajad Shiekh

FRET-PAINT for Studying Long Telomeric DNA Overhangs and Their Interactions with Shelterin Proteins

Sajad Shiekh (Department of Physics, Kent State University, Kent Ohio), Golam Mustafa, Sineth G Kodikara, Eric Yokie, Prabesh Gyawali (Department of Physics, Kent State University, Kent Ohio), John J. Portmana, Hamza Balcia (Department of Physics, Kent State University, Kent Ohio), Mohammed Enamul Hoque (Department of Chemistry and Biochemistry, Kent State University, Kent Ohio), Amanda Jack, Ahmet Yildiz (Department of Molecular and Cell Biology, University of California, Berkeley, Physics Department, University of California, Berkeley)

I will be presenting the single molecule FRET-PAINT measurements and computational modeling studies where we investigated the accessibility of a range of different human telomeric overhangs. These overhangs can form 1-7 tandem G-quadruplex (GQ) structures, which, to our knowledge, is the most comprehensive range studied to date and covers a significant portion of the physiologically relevant telomeric overhang length scale. Our measurements demonstrate novel accessibility maps where certain regions of the telomeric overhang are significantly more accessible than others. We also observe folding frustration patterns with a well-defined periodicity and constructs with a certain number of telomeric repeats demonstrate elevated levels of frustration compared to others. These patterns have significant implications for telomere organization, protection of free 3’-end against exonuclease activity and telomerase-catalyzed extension, and folding cooperativity between neighboring GQ structures. We further investigated the impact of POT1 and a four-protein Shelterin complex on the accessibility of human telomeric DNA constructs with physiologically relevant overhang lengths (28-150 nt). To quantify telomere accessibility, we monitored transient binding events of a Cy5-labeled, short peptide nucleic acid strand to available sites on the telomere using the FRET-PAINT methodology. We observed approximately 2.5-fold reduced accessibility in the presence of POT1 (compared to DNA-only case) and about 5-fold reduced accessibility in the presence of Shelterin. These results suggest the protection of exposed telomeric segments by POT1 was effective enough to dominate over its mild GQ unfolding activity. The more effective protection in the presence of Shelterin compared to the POT1-only case suggests the restructuring of the junction region between single and double-stranded telomere, which is otherwise the most accessible part of the overhang, serves an important function in telomere maintenance.

1. Sajad Shiekh, G Mustafa, Mohammed Enamul Hoque, Eric Yokie, John J. Portman, H Balci."Emerging Accessibility Patterns in Long Telomeric Overhangs"(

1. G Mustafa, S Shiekh, K Gc, S Abeysirigunawardena, H Balci. "Interrogating accessibility of telomeric sequences with FRET-PAINT: evidence for length-dependent telomere compaction" NAR, Vol 49, Issue 6, Pages 3371–3380

Keywords: FRET-PAINT, G-quadruplexes, Telomere