Inhibiting transcription of chromosomal DNA with antigene peptide nucleic acids

Abstract

Synthetic molecules that recognize specific sequences within cellular DNA are potentially powerful tools for investigating chromosome structure and function. Here, we designed antigene peptide nucleic acids (agPNAs) to target the transcriptional start sites for the human progesterone receptor B (hPR-B) and A (hPR-A) isoforms at sequences predicted to be single-stranded within the open complex of chromosomal DNA. We found that the agPNAs were potent inhibitors of transcription, showing for the first time that synthetic molecules can recognize transcription start sites inside cells. Breast cancer cells treated with agPNAs showed marked changes in morphology and an unexpected relationship between the strictly regulated levels of hPR-B and hPR-A. We confirmed these phenotypes using siRNAs and antisense PNAs, demonstrating the power of combining antigene and antisense strategies for gene silencing. agPNAs provide a general approach for controlling transcription initiation and a distinct option for target validation and therapeutic development.

Closed Chromatin Architecture Is Induced by an RNA Duplex Targeting the HIV-1 Promoter Region

“…Specific target gene activation has been demonstrated using promoter-targeted dsRNAs to several mammalian genes (31,70). As yet the characteristics differentiating activating from inhibitory promoter-targeted dsRNAs have not been defined; however, these are likely to be subtle and complex (13,16,70). Furthermore, although nonspecific "off target" activation has been described by certain antisense and dsRNAs targeting the 5Ј-LTR of HIV-1 at sites over 100 bases upstream of the prom-A siRNA target site (22), the determinants of these effects are unclear at this point.…”

“…These directly target the transcription start site and induce repressive changes in chromatin architecture of the promoter region, which extend through this site (13). It is also becoming clear that miRNAs can also induce both changes in the histone code and chromatin remodeling resulting in repressive chromatin structures.…”

Progress and prospects: RNA-based therapies for treatment of HIV infection

“…Exogenous siRNAs targeting specific promoters in mammalian cells have been shown to affect expression of the corresponding gene. 44,[57][58][59][60][61][62][63][64] Although the mechanism is not fully understood, the possibility of using TGS to intervene earlier in the viral replication cycle by repressing transcription is very attractive, especially because it avoids the opportunities for viral escape that active replication provides. Also, strategies targeting HIV cellular cofactors that by themselves mediate only moderate suppression of HIV, but avoid toxicity, may be more effective in combination with TGS.…”

Experimental non-ATP-competitive therapies for chronic myelogenous leukemia

“…39 PNAs are nonionic DNA mimics that have outstanding potential for recognizing duplex DNA. 40 PNAs are hybrid molecules formed by both a peptide and a nucleic acid and consisting of N-(2-aminoethyl)glycine monomers linked by amide bonds. 40,41 The purine (A and G) and pyrimidine (C and T) nucleobases are attached to this polyamide backbone via a methylene carbonyl linkage.…”

“…40 PNAs are hybrid molecules formed by both a peptide and a nucleic acid and consisting of N-(2-aminoethyl)glycine monomers linked by amide bonds. 40,41 The purine (A and G) and pyrimidine (C and T) nucleobases are attached to this polyamide backbone via a methylene carbonyl linkage. 42 On the basis of these characteristics, PNAs are considered as synthetic DNA analogues that are resistant to nucleases and proteases and have the ability to bind with very high affinity to RNA.…”