Poster abstracts
Poster number 68 submitted by Mohammad Lutful Kabir
Exploring the Interplay of G-Quadruplexes and CRISPR/dCas9 in Gene Expression Regulation
Mohammad Lutful kabir (Chemistry and Biochemistry, Kent State University), Hamza Balci (Physics, Kent State University), Soumitra basu (Chemistry and Biochemistry, Kent State University)
Abstract:
Gene expression control mechanisms play a critical role in molecular biology. Both G-quadruplexes (GQ) and the CRISPR/dCas9 complex have emerged as intriguing tools for gene regulation by impeding the progression of RNA Polymerase. Unlike alternative methods, the CRISPR/dCas9 complex offers specificity, transience, and does not necessitate sequence mutation. However, the comparative behavior of the CRISPR/dCas9 complex while in proximity to GQ remains unresolved. This study aims to elucidate the underlying mechanism and assess the combined impact of these two independent molecular systems on gene expression inhibition. Using c-MYC transcription as a model system, we focus on the nuclease hypersensitivity element (NHE) III, a critical regulator of 85–90% of c-MYC transcription. Situated 142 to 115 base pairs upstream from the P1 promoter, NHE III is comprised of a 27-bp sequence known to form two distinct intramolecular GQ structures within the purine-rich DNA strand. Our findings reveal that when employed together, CRISPR/dCas9 and selective GQ targeting exhibit a remarkable capacity to suppress in vitro transcription. Notably, the CRISPR/dCas9 complex may facilitate and/or stabilize the GQ structure, which results in an inhibitory effect on gene expression. However, the extent of the impediment is contingent on the target DNA strand and its specific position with respect to the GQ. Moreover, it demonstrates the ability to suppress more than 50% in-vivo c-MYC transcriptional activation, presumably by stabilizing the GQ structure within the c-MYC NHE region of the Ramos cells, which are B-lymphocytes. This study sheds light on the intricate interaction between GQ and CRISPR/dCas9, emphasizing their potential as potent and versatile gene expression modulators.
Keywords: CRISPRdCas9 complex, G-quadruplexes (GQ), Gene Expression