We have developed a new molecular beacon design that requires an additional UV pulse for fluorescence activation. This improves the signal-to-noise ratio tremendously compared to previous approaches and allows for a precise control of the time point and location of RNA labelling.
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Light-Inducible Deformation of Mitochondria in Live Cells.
Methods Mol Biol
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Department of Biomedical Engineering, The Chinese University of Hong Kong, Hong Kong, China.
Mitochondria are dynamic organelles with constantly changing morphologies. Despite recent reports indicating that mechanical cues modulate mitochondrial morphologies and functions, there is a lack of methods that can exclusively and precisely exert mechanical forces to and deform mitochondria in live cells. Therefore, how mitochondria sense and respond to mechanical forces remains largely elusive.
View Article and Find Full Text PDF-targeted, stable expression of ChR2 in human brain organoids for consistent optogenetic control.
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November 2024
School of Biological Sciences, College of Natural Sciences, Seoul National University Seoul Republic of Korea.
Self-organizing brain organoids provide a promising tool for studying human development and disease. Here we created human forebrain organoids with stable and homogeneous expression of channelrhodopsin-2 (ChR2) by generating safe harbor locus-targeted, ChR2 knocked-in human pluripotent stem cells (hPSCs), followed by the differentiation of these genetically engineered hPSCs into forebrain organoids. The resulting ChR2-expressing human forebrain organoids showed homogeneous cellular expression of ChR2 throughout entire regions without any structural and functional perturbations and displayed consistent and robust neural activation upon light stimulation, allowing for the non-virus mediated, spatiotemporal optogenetic control of neural activities.
View Article and Find Full Text PDFIn vivo optogenetic manipulations of endogenous proteins reveal spatiotemporal roles of microtubule and kinesin in dendrite patterning.
Sci Adv
August 2024
Weill Institute for Cell and Molecular Biology, Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
During animal development, the spatiotemporal properties of molecular events largely determine the biological outcomes. Conventional gene analysis methods lack the spatiotemporal resolution for precise dissection of developmental mechanisms. Although optogenetic tools exist for manipulating designer proteins in cultured cells, few have been successfully applied to endogenous proteins in live animals.
View Article and Find Full Text PDFGlucose-Operated Widget (GLOW) for Closed-Loop Optogenetic Glycemic Control.
Adv Mater
November 2024
Department of Biosystems Science and Engineering, ETH Zurich, Klingelbergstrasse 48, Basel, CH-4056, Switzerland.
Closed-loop control systems for precise control of therapeutic gene expression are promising candidates for personalized treatment of chronic ailments such as diabetes. Pancreatic iβ-cells are engineered with blue-light-inducible melanopsin to drive rapid insulin release by vesicular secretion from intracellular stores. In this work, a glucose-operated widget (GLOW) is designed as a component of a closed-loop control system for diabetes treatment by employing a probe that emits blue fluorescence in a glucose-concentration-dependent manner as a real-time glucose sensor to precisely control insulin release from these iβ-cells.
View Article and Find Full Text PDFCondensate interfacial forces reposition DNA loci and probe chromatin viscoelasticity.
Cell
September 2024
Department of Chemical and Biological Engineering, Princeton University, Princeton, NJ 08544, USA; Omenn-Darling Bioengineering Institute, Princeton University, Princeton, NJ 08544, USA; Princeton Materials Institute, Princeton University, Princeton, NJ 08544, USA; Howard Hughes Medical Institute, Chevy Chase, MD 21044, USA. Electronic address:
Biomolecular condensates assemble in living cells through phase separation and related phase transitions. An underappreciated feature of these dynamic molecular assemblies is that they form interfaces with other cellular structures, including membranes, cytoskeleton, DNA and RNA, and other membraneless compartments. These interfaces are expected to give rise to capillary forces, but there are few ways of quantifying and harnessing these forces in living cells.
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