MECHANICAL STABILITY EVALUATION OF I-MOTIF AND G-QUADRUPLEX STRUCTURES UNDER DIVERSE CIRCUMSTANCES
‘ILPR (Insulin Linked Polymorphic Region) is a promoter region located upstream of the human insulin gene. This region facilitates the transcription and hence regulates the production of insulin, a hormone responsible for the metabolism of glucose. When the control of insulin level fails, it leads to insulin related health problems such as diabetes. One of the scenarios that influence the transcription of the insulin gene can be the formation of secondary structures in the promoter. Due to the prevalence of the guanine (G)- and cytosine (C)-rich repeats in the ILPR promoter, it is likely to form unusual structures such as G-quadruplex and i-motif in the respective strands.
G-quadruplex and i-motif structures are the most widely known four-stranded nucleic acid structures which are shown/proposed to alter gene regulation. Although there is significant understanding on the folding topologies of G-quadruplex and i-motif structures, their mechanical stability which determines the interaction with motor proteins, such as DNA/RNA polymerases, are poorly studied. Using laser tweezers based single-molecule study; we investigated, for the first time, the mechanical stability of an i-motif structure in the predominant variant of human ILPR. Similarly, using ensemble and single-molecule approaches, we show that a dsDNA fragment in ILPR can fold into G-quadruplex or i-motif structure under specific conditions. Under a condition that favors the formation of both G-quadruplex and i-motif, we provide compelling evidence that only one species is present in each dsDNA. We propose that G-quadruplex and i-motif are mutually exclusive in human ILPR. Due to the fact that these two species have an unfolding force comparable to the stall force of RNA polymerase, these DNA structures may play significant biological rules in the expression of human insulin inside cells.’
Below are some selected publications:
Dhakal et al., Structural and mechanical properties of individual human telomeric G-quadruplexes in molecularly crowded solutions, Nucleic Acids Research, 2013, Accepted
Dhakal et al., Intramolecular folding in human ILPR fragment with three C-rich repeats PLoS ONE 7(6): e39271. doi:10.1371/journal.pone.0039271
Dhakal et al., G-quadruplex and i-motif are mutually exclusive in double stranded ILPR DNA, Biophysical Journal, 102, 2575–2584
Dhakal et al., Formation of human ILPR G-quadruplex in dsDNA, International Review of Biophysical Chemistry, 2011, 2, N. 6.
Dhakal et al., Coexistence of an ILPR i-motif and a partially folded structure with comparable mechanical stability revealed at the single molecular level, J. Am. Chem. Soc. 2010, 132, 8991–8997
Book Chapter:
Dhakal, S.; Mao, H.; Rajendran, A.; Endo, M.; Sugiyama, H., Eds: Spindler L, Fritzsche W., “G-quadruplex Nanostructures Probed at The Single Molecular Level By Force-Based Methods” in Guanine quartets: Structure and Application, RSC Publishing, 2013, 73-85.
