Suge Zhang1, Hongxia Sun2, Hongbo Chen1, Qian Li3, Aijiao Guan4, Lixia Wang3, Yunhua Shi1, Shujuan Xu1, Meirong Liu5, Yalin Tang6. 1. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry Chinese Academy of Sciences, 100190 Beijing, PR China; University of Chinese Academy of Sciences, Beijing, 100049, PR China. 2. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry Chinese Academy of Sciences, 100190 Beijing, PR China. Electronic address: hongxsun@iccas.ac.cn. 3. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry Chinese Academy of Sciences, 100190 Beijing, PR China. 4. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry Chinese Academy of Sciences, 100190 Beijing, PR China. Electronic address: ajguan@iccas.ac.cn. 5. Center for Physiochemical Analysis & Measurement, Institute of Chemistry Chinese Academy of Sciences, Beijing, 100190, PR China. 6. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry Chinese Academy of Sciences, 100190 Beijing, PR China; University of Chinese Academy of Sciences, Beijing, 100049, PR China. Electronic address: tangyl@iccas.ac.cn.
Abstract
BACKGROUND: Direct detection of G-quadruplexes in human cells has become an important issue due to the vital role of G-quadruplex related to biological functions. Despite several probes have been developed for detection of the G-quadruplexes in cytoplasm or whole cells, the probe being used to monitor the nucleolar G-quadruplexes is still lacking. METHODS: Formation of the nucleolar G-quadruplex structures was confirmed by using circular dichroism (CD) spectroscopy. The binding affinity and selectivity of Thioflavin T (ThT) towards various DNA/RNA motifs in solution and gel system were measured by using fluorescence spectroscopy and polyacrylamide gel electrophoresis (PAGE), respectively. G-quadruplex imaging in live cells was directly captured by using confocal laser scanning microscopy (CLSM). RESULTS: Formation of the rDNA and rRNA G-quadruplex structures is demonstrated in vitro. ThT is found to show much higher affinity and selectivity towards these G-quadruplex structures versus other nucleic acid motifs either in solution or in gel system. The nucleolar G-quadruplexes in living cells are visualized by using ThT as a fluorescent probe. G-quadruplex-ligand treatments in live cells lead to sharp decrease of ThT signal. CONCLUSIONS: The natural existence of the G-quadruplexes structure in the nucleoli of living cells is directly visualized by using ThT as an indicator. GENERAL SIGNIFICANCE: The research provides substantive evidence for formation of the rRNA G-quadruplex structures, and also offers an effective probe for direct visualization of the nucleolar G-quadruplexes in living cells.
BACKGROUND: Direct detection of G-quadruplexes in human cells has become an important issue due to the vital role of G-quadruplex related to biological functions. Despite several probes have been developed for detection of the G-quadruplexes in cytoplasm or whole cells, the probe being used to monitor the nucleolar G-quadruplexes is still lacking. METHODS: Formation of the nucleolar G-quadruplex structures was confirmed by using circular dichroism (CD) spectroscopy. The binding affinity and selectivity of Thioflavin T (ThT) towards various DNA/RNA motifs in solution and gel system were measured by using fluorescence spectroscopy and polyacrylamide gel electrophoresis (PAGE), respectively. G-quadruplex imaging in live cells was directly captured by using confocal laser scanning microscopy (CLSM). RESULTS: Formation of the rDNA and rRNA G-quadruplex structures is demonstrated in vitro. ThT is found to show much higher affinity and selectivity towards these G-quadruplex structures versus other nucleic acid motifs either in solution or in gel system. The nucleolar G-quadruplexes in living cells are visualized by using ThT as a fluorescent probe. G-quadruplex-ligand treatments in live cells lead to sharp decrease of ThT signal. CONCLUSIONS: The natural existence of the G-quadruplexes structure in the nucleoli of living cells is directly visualized by using ThT as an indicator. GENERAL SIGNIFICANCE: The research provides substantive evidence for formation of the rRNA G-quadruplex structures, and also offers an effective probe for direct visualization of the nucleolar G-quadruplexes in living cells.
Authors: Santi Mestre-Fos; Petar I Penev; Suttipong Suttapitugsakul; Michael Hu; Chieri Ito; Anton S Petrov; Roger M Wartell; Ronghu Wu; Loren Dean Williams Journal: J Mol Biol Date: 2019-03-15 Impact factor: 5.469
Authors: Adam T Green; Amanda J Pickard; Rongzhong Li; Alexander D MacKerell; Ulrich Bierbach; Samuel S Cho Journal: J Phys Chem B Date: 2022-01-13 Impact factor: 2.991
Authors: Brian Noh; Maria P Blasco-Conesa; Yun-Ju Lai; Bhanu Priya Ganesh; Akihiko Urayama; Ines Moreno-Gonzalez; Sean P Marrelli; Louise D McCullough; Jose Felix Moruno-Manchon Journal: Front Aging Date: 2022-01-12
Authors: Olaya Domarco; Claudia Kieler; Christine Pirker; Carina Dinhof; Bernhard Englinger; Johannes M Reisecker; Gerald Timelthaler; Marcos D García; Carlos Peinador; Bernhard K Keppler; Walter Berger; Alessio Terenzi Journal: Angew Chem Int Ed Engl Date: 2019-05-08 Impact factor: 15.336
Authors: Santi Mestre-Fos; Petar I Penev; John Colin Richards; William L Dean; Robert D Gray; Jonathan B Chaires; Loren Dean Williams Journal: PLoS One Date: 2019-12-13 Impact factor: 3.240