Honors & Awards
Nomination for ETH Medal for outstanding doctoral thesis projects, ETH Zurich (08/2018)
HHMI Janelia Graduate Research Fellowship, Howard Hughes Medical Institute (HHMI) (10/2016)
Honorary Membership in the American National Honor Society, US National Honor Society (2006)
Honorary Membership in the Mu Alpha Theta National Honor Society for mathematics, Mu Alpha Theta (2006)
Online Stipend, e-fellows (2008)
Willi Studer Price for the best Master Student of the year, ETH Zurich (2015)
Doctor of Science, Eidgenossische Technische Hochschule (ETH Zurich) (2018)
Master of Science, Eidgenossische Technische Hochschule (ETH Zurich) (2014)
Bachelor of Science, Technische Universitat Munchen (2011)
jYCaMP: an optimized calcium indicator for two-photon imaging at fiber laser wavelengths.
Femtosecond lasers at fixed wavelengths above 1,000nm are powerful, stable and inexpensive, making them promising sources for two-photon microscopy. Biosensors optimized for these wavelengths are needed for both next-generation microscopes and affordable turn-key systems. Here we report jYCaMP1, a yellow variant of the calcium indicator jGCaMP7 that outperforms its parent in mice and flies at excitation wavelengths above 1,000nm and enables improved two-color calcium imaging with red fluorescent protein-based indicators.
View details for DOI 10.1038/s41592-020-0835-7
View details for PubMedID 32451475
Primed Track, high-fidelity lineage tracing in mouse pre-implantation embryos using primed conversion of photoconvertible proteins
Accurate lineage reconstruction of mammalian pre-implantation development is essential for inferring the earliest cell fate decisions. Lineage tracing using global fluorescence labeling techniques is complicated by increasing cell density and rapid embryo rotation, which hampers automatic alignment and accurate cell tracking of obtained four-dimensional imaging data sets. Here, we exploit the advantageous properties of primed convertible fluorescent proteins (pr-pcFPs) to simultaneously visualize the global green and the photoconverted red population in order to minimize tracking uncertainties over prolonged time windows. Confined primed conversion of H2B-pr-mEosFP-labeled nuclei combined with light-sheet imaging greatly facilitates segmentation, classification, and tracking of individual nuclei from the 4-cell stage up to the blastocyst. Using green and red labels as fiducial markers, we computationally correct for rotational and translational drift, reduce overall data size, and accomplish high-fidelity lineage tracing even for increased imaging time intervals - addressing major concerns in the field of volumetric embryo imaging.
View details for PubMedID 30663981
moxMaple3: a Photoswitchable Fluorescent Protein for PALM and Protein Highlighting in Oxidizing Cellular Environments
2018; 8: 14738
The ability of fluorescent proteins (FPs) to fold robustly is fundamental to the autocatalytic formation of the chromophore. While the importance of the tertiary protein structure is well appreciated, the impact of individual amino acid mutations for FPs is often not intuitive and requires direct testing. In this study, we describe the engineering of a monomeric photoswitchable FP, moxMaple3, for use in oxidizing cellular environments, especially the eukaryotic secretory pathway. Surprisingly, a point mutation to replace a cysteine substantially improved the yield of correctly folded FP capable of chromophore formation, regardless of cellular environment. The improved folding of moxMaple3 increases the fraction of visibly tagged fusion proteins, as well as FP performance in PALM super-resolution microscopy, and thus makes moxMaple3 a robust monomeric FP choice for PALM and optical highlighting applications.
View details for DOI 10.1038/s41598-018-32955-5
View details for Web of Science ID 000446339400022
View details for PubMedID 30283009
View details for PubMedCentralID PMC6170497
Primed Conversion: The New Kid on the Block for Photoconversion
CHEMISTRY-A EUROPEAN JOURNAL
2018; 24 (33): 8268-+
In 2015, a novel way to convert photoconvertible fluorescent proteins was reported that uses the intercept of blue and far-red light instead of traditional violet or near-UV light illumination. This Minireview describes and contrasts this distinct two-step mechanism termed primed conversion with traditional photoconversion. We provide a comprehensive overview of what is known to date about primed conversion and focus on the molecular requirements for it to take place. We provide examples of its application to axially confined photoconversion in complex tissues as well as super-resolution microscopy. Further, we describe why and when it is useful, including its advantages and disadvantages, and give an insight into potential future development in the field.
View details for PubMedID 29430743
Virus stamping for targeted single-cell infection in vitro and in vivo
2018; 36 (1): 81-+
Genetic engineering by viral infection of single cells is useful to study complex systems such as the brain. However, available methods for infecting single cells have drawbacks that limit their applications. Here we describe 'virus stamping', in which viruses are reversibly bound to a delivery vehicle-a functionalized glass pipette tip or magnetic nanoparticles in a pipette-that is brought into physical contact with the target cell on a surface or in tissue, using mechanical or magnetic forces. Different single cells in the same tissue can be infected with different viruses and an individual cell can be simultaneously infected with different viruses. We use rabies, lenti, herpes simplex, and adeno-associated viruses to drive expression of fluorescent markers or a calcium indicator in target cells in cell culture, mouse retina, human brain organoid, and the brains of live mice. Virus stamping provides a versatile solution for targeted single-cell infection of diverse cell types, both in vitro and in vivo.
View details for PubMedID 29251729
Monitoring and manipulating cellular crosstalk during kidney fibrosis inside a 3D in vitro co-culture
2017; 7: 14490
In pharmacological research the development of promising lead compounds requires a detailed understanding of the dynamics of disease progression. However, for many diseases, such as kidney fibrosis, gaining such understanding requires complex real-time, multi-dimensional analysis of diseased and healthy tissue. To allow for such studies with increased throughput we established a dextran hydrogel-based in vitro 3D co-culture as a disease model for kidney fibrosis aimed at the discovery of compounds modulating the epithelial/mesenchymal crosstalk. This platform mimics a simplified pathological renal microenvironment at the interface between tubular epithelial cells and surrounding quiescent fibroblasts. We combined this 3D technology with epithelial reporter cell lines expressing fluorescent biomarkers in order to visualize pathophysiological cell state changes resulting from toxin-mediated chemical injury. Epithelial cell damage onset was robustly detected by image-based monitoring, and injured epithelial spheroids induced myofibroblast differentiation of co-cultured quiescent human fibroblasts. The presented 3D co-culture system therefore provides a unique model system for screening of novel therapeutic molecules capable to interfere and modulate the dialogue between epithelial and mesenchymal cells.
View details for PubMedID 29101326
Rational Engineering of Photoconvertible Fluorescent Proteins for Dual-Color Fluorescence Nanoscopy Enabled by a Triplet-State Mechanism of Primed Conversion
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
2017; 56 (38): 11628–33
Green-to-red photoconvertible fluorescent proteins (pcFPs) are powerful tools for super-resolution localization microscopy and protein tagging. Recently, they have been found to undergo efficient photoconversion not only by the traditional 400-nm illumination but also by an alternative method termed primed conversion, employing dual wavelength illumination with blue and far-red/near-infrared light. Primed conversion has been reported only for Dendra2 and its mechanism has remained elusive. Here, we uncover the molecular mechanism of primed conversion by reporting the intermediate "primed" state to be a triplet dark state formed by intersystem crossing. We show that formation of this state can be influenced by the introduction of serine or threonine at sequence position 69 (Eos notation) and use this knowledge to create "pr"- (for primed convertible) variants of most known green-to-red pcFPs.
View details for PubMedID 28661566
Optogenetic control with a photocleavable protein, PhoCl
2017; 14 (4): 391-+
To expand the range of experiments that are accessible with optogenetics, we developed a photocleavable protein (PhoCl) that spontaneously dissociates into two fragments after violet-light-induced cleavage of a specific bond in the protein backbone. We demonstrated that PhoCl can be used to engineer light-activatable Cre recombinase, Gal4 transcription factor, and a viral protease that in turn was used to activate opening of the large-pore ion channel Pannexin-1.
View details for PubMedID 28288123
Labeling cellular structures in vivo using confined primed conversion of photoconvertible fluorescent proteins
2016; 11 (12): 2419–31
The application of green-to-red photoconvertible fluorescent proteins (PCFPs) for in vivo studies in complex 3D tissue structures has remained limited because traditional near-UV photoconversion is not confined in the axial dimension, and photomodulation using axially confined, pulsed near-IR (NIR) lasers has proven inefficient. Confined primed conversion is a dual-wavelength continuous-wave (CW) illumination method that is capable of axially confined green-to-red photoconversion. Here we present a protocol to implement this technique with a commercial confocal laser-scanning microscope (CLSM); evaluate its performance on an in vitro setup; and apply primed conversion for in vivo labeling of single cells in developing zebrafish and mouse preimplantation embryos expressing the green-to-red photoconvertible protein Dendra2. The implementation requires a basic understanding of laser-scanning microscopy, and it can be performed within a single day once the required filter cube is manufactured.
View details for PubMedID 27809312
Single neuron morphology in vivo with confined primed conversion
ZEBRAFISH: CELLULAR AND DEVELOPMENTAL BIOLOGY, PT A: CELLULAR BIOLOGY, 4TH EDITION
2016; 133: 125–38
Unraveling the structural organization of neurons can provide fundamental insights into brain function. However, visualizing neurite morphology in vivo remains difficult due to the high density and complexity of neural packing in the nervous system. Detailed analysis of neural morphology requires distinction of closely neighboring, highly intricate cellular structures such as neurites with high contrast. Green-to-red photoconvertible fluorescent proteins have become powerful tools to optically highlight molecular and cellular structures for developmental and cell biological studies. Yet, selective labeling of single cells of interest in vivo has been precluded due to inefficient photoconversion when using high intensity, pulsed, near-infrared laser sources that are commonly applied for achieving axially confined two-photon (2P) fluorescence excitation. Here we describe a novel optical mechanism, "confined primed conversion," which employs continuous dual-wave illumination to achieve confined green-to-red photoconversion of single cells in live zebrafish embryos. Confined primed conversion exhibits wide applicability and this chapter specifically elaborates on employing this imaging modality to analyze neural morphology of optically targeted single neurons in the developing zebrafish brain.
View details for DOI 10.1016/bs.mcb.2015.12.005
View details for Web of Science ID 000382401100007
View details for PubMedID 27263411