Shumei Liang1, Yunyan He2, Yun Xia3, Huijuan Wang1, Lipeng Wang1, Rui Gao1, Meilan Zhang1. 1. Department of Clinical Laboratory, the First Affiliated Hospital of Chongqing Medical University, 1 You Yi Road, Yuzhong District, Chongqing 40016, China. 2. Department of Clinical Laboratory, Chongqing Zhongshan Hospital, Chongqing, China. 3. Department of Clinical Laboratory, the First Affiliated Hospital of Chongqing Medical University, 1 You Yi Road, Yuzhong District, Chongqing 40016, China. Electronic address: xiayun12cn@aliyun.com.
Abstract
BACKGROUND: The increasing emergence of clinical infections caused by methicillin-resistant Staphylococcus aureus (MRSA) challenges existing therapeutic options and highlights the need to develop novel treatment strategies. The ftsZ gene is essential to bacterial cell division. METHODS: In this study, two antisense peptide nucleic acids (PNAs) conjugated to a cell-penetrating peptide were used to inhibit the growth of MRSA. PPNA1, identified with computational prediction and dot-blot hybridization, is complementary to nucleotides 309-323 of the ftsZ mRNA. PPNA2 was designed to target the region that includes the translation initiation site and the ribosomal-binding site (Shine-Dalgarno sequence) of the ftsZ gene. Scrambled PPNA was constructed with mismatches to three bases within the antisense PPNA1 sequence. RESULTS: PPNA1 and PPNA2 caused concentration-dependent growth inhibition and had bactericidal effects. The minimal bactericidal concentrations of antisense PPNA1 and PPNA2 were 30μmol/l and 40μmol/l, respectively. The scrambled PPNA had no effect on bacterial growth, even at higher concentrations, confirming the sequence specificity of the probes. RT-PCR showed that the antisense PPNAs suppressed ftsZ mRNA expression in a dose-dependent manner. CONCLUSION: Our results demonstrate that the potent effects of PNAs on bacterial growth and cell viability were mediated by the down-regulation or even knock-out of ftsZ gene expression. This highlights the utility of ftsZ as a promising target for the development of new antisense antibacterial agents to treat MRSA infections.
BACKGROUND: The increasing emergence of clinical infections caused by methicillin-resistant Staphylococcus aureus (MRSA) challenges existing therapeutic options and highlights the need to develop novel treatment strategies. The ftsZ gene is essential to bacterial cell division. METHODS: In this study, two antisense peptide nucleic acids (PNAs) conjugated to a cell-penetrating peptide were used to inhibit the growth of MRSA. PPNA1, identified with computational prediction and dot-blot hybridization, is complementary to nucleotides 309-323 of the ftsZ mRNA. PPNA2 was designed to target the region that includes the translation initiation site and the ribosomal-binding site (Shine-Dalgarno sequence) of the ftsZ gene. Scrambled PPNA was constructed with mismatches to three bases within the antisense PPNA1 sequence. RESULTS: PPNA1 and PPNA2 caused concentration-dependent growth inhibition and had bactericidal effects. The minimal bactericidal concentrations of antisense PPNA1 and PPNA2 were 30μmol/l and 40μmol/l, respectively. The scrambled PPNA had no effect on bacterial growth, even at higher concentrations, confirming the sequence specificity of the probes. RT-PCR showed that the antisense PPNAs suppressed ftsZ mRNA expression in a dose-dependent manner. CONCLUSION: Our results demonstrate that the potent effects of PNAs on bacterial growth and cell viability were mediated by the down-regulation or even knock-out of ftsZ gene expression. This highlights the utility of ftsZ as a promising target for the development of new antisense antibacterial agents to treat MRSA infections.
Authors: José Rivera-Chávez; Lindsay Caesar; Juan J Garcia-Salazar; Huzefa A Raja; Nadja B Cech; Cedric J Pearce; Nicholas H Oberlies Journal: Tetrahedron Lett Date: 2019-01-17 Impact factor: 2.415