Identification and Characterization of Naturally Derived Splicealtering Compounds and Their Potential Impact on Individuals With Hutchinson-Gilford Progeria Syndrome
Abstract
Hutchinson-Gilford progeria syndrome is a rare autosomal dominant disease primarily caused by a C to T transition in the 1824th coding nucleotide of the LMNA gene. This mutation creates a cryptic splice donor site in exon 11 of LMNA, resulting in the production of a new LMNA transcript that lacks the terminal 150 nucleotides of exon 11. This shorter transcript encodes a protein called progerin that becomes permanently anchored to the inner nuclear membrane, resulting in nuclear malformation, leading to global changes in nuclear structure and function. Increasing the understanding of how small molecules influence the production of the progerin-encoding transcript is paramount in the search for new therapeutic modalities for the HGPS population.I present here two small molecules, trans-resveratrol and luteolin, isolated from plant extracts of commonly consumed foods that exhibit the ability to alter the splicing of LMNA transcripts. Trans-resveratrol modulated the splicing of LMNA transcripts resulting in an increase in the production of the LMNAΔ150 transcript at the expense of the production of the LMNAWT transcript in HEK293 cells transfected with a LMNA minigene carrying the c.C1824T HGPS-causing mutation and in HGPS patient-derived lymphocyte cell lines. Furthermore, I observed an increase in the progerin/lamin A protein ratio in trans-resveratrol treated HGPS lymphocyte cell lines. Trans-resveratrol treatment also suppressed the protein levels of the serine and arginine-rich splicing factor 3 (SRSF3). Suppression of SRSF3 levels using targeting siRNA mimicked the change in splicing of minigene-derived LMNA transcripts I observed in transresveratrol treated HEK293 cells. Suppression of the trans-resveratrol-mediated decrease in SRSF3 levels through overexpressing SRSF3 in the presence of trans-resveratrol inhibited the trans-resveratrol induced increase in LMNAΔ150 transcript levels, indicating that trans-resveratrol mediates its ability to alter LMNA splicing, in part, through suppressing SRSF3 levels.Similar to what I observed with trans-resveratrol, luteolin treatment of HEK293 cells transfected with a LMNA minigene carrying a HGPS-causing mutation and in HGPS lymphocyte cell lines resulted in a change in LMNA splicing to favor the production of the LMNAΔ150 transcript relative to the LMNAWT transcript. Luteolin also induced an increase in the progerin/lamin A protein ratio in HGPS lymphocyte cell lines. I further explored the mechanism by which luteolin exerts its ability to modulate LMNA splicing and determined that luteolin has no impact on nuclear levels of the heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) or the heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1). Furthermore, siRNAmediated suppression of hnRNP A1 and hnRNP A2/B1 resulted in no significant change in LMNA splicing in HEK293 cells transfected with a LMNA minigene carrying the most common HGPS-causing mutation. These results taken together provide no evidence that luteolin modulates the RNA splicing process through impacting the activity of hnRNP A1 or hnRNP A2/B1. The findings of this thesis reveal the underlying importance of understanding whether what the HGPS population is consuming is influencing the progression of their disease.
Subject Area
Molecular biology|Biology|Genetics
Recommended Citation
Evans, Anthony J, "Identification and Characterization of Naturally Derived Splicealtering Compounds and Their Potential Impact on Individuals With Hutchinson-Gilford Progeria Syndrome" (2023). ETD Collection for Fordham University. AAI30486879.
https://research.library.fordham.edu/dissertations/AAI30486879
