@article{TzonevaStoyanovaPetrichetal.2020,
  author    = {Tzoneva, Rumiana and Stoyanova, Tihomira and Petrich, Annett and Popova, Desislava and Uzunova, Veselina and Momchilova, Albena and Chiantia, Salvatore},
  title     = {Effect of Erufosine on Membrane Lipid Order in Breast Cancer Cell Models},
  series = {Biomolecules},
  volume    = {10},
  journal   = {Biomolecules},
  number    = {5},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2218-273X},
  doi       = {10.3390/biom10050802},
  pages     = {17},
  year      = {2020},
  abstract  = {Alkylphospholipids are a novel class of antineoplastic drugs showing remarkable therapeutic potential. Among them, erufosine (EPC3) is a promising drug for the treatment of several types of tumors. While EPC3 is supposed to exert its function by interacting with lipid membranes, the exact molecular mechanisms involved are not known yet. In this work, we applied a combination of several fluorescence microscopy and analytical chemistry approaches (i.e., scanning fluorescence correlation spectroscopy, line-scan fluorescence correlation spectroscopy, generalized polarization imaging, as well as thin layer and gas chromatography) to quantify the effect of EPC3 in biophysical models of the plasma membrane, as well as in cancer cell lines. Our results indicate that EPC3 affects lipid-lipid interactions in cellular membranes by decreasing lipid packing and increasing membrane disorder and fluidity. As a consequence of these alterations in the lateral organization of lipid bilayers, the diffusive dynamics of membrane proteins are also significantly increased. Taken together, these findings suggest that the mechanism of action of EPC3 could be linked to its effects on fundamental biophysical properties of lipid membranes, as well as on lipid metabolism in cancer cells.},
  language  = {en}
}
@misc{TzonevaStoyanovaPetrichetal.2020,
  author    = {Tzoneva, Rumiana and Stoyanova, Tihomira and Petrich, Annett and Popova, Desislava and Uzunova, Veselina and Albena, Momchilova and Chiantia, Salvatore},
  title     = {Effect of Erufosine on Membrane Lipid Order in Breast Cancer Cell Models},
  series = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  number    = {1000},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-47705},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-477056},
  pages     = {19},
  year      = {2020},
  abstract  = {Alkylphospholipids are a novel class of antineoplastic drugs showing remarkable therapeutic potential. Among them, erufosine (EPC3) is a promising drug for the treatment of several types of tumors. While EPC3 is supposed to exert its function by interacting with lipid membranes, the exact molecular mechanisms involved are not known yet. In this work, we applied a combination of several fluorescence microscopy and analytical chemistry approaches (i.e., scanning fluorescence correlation spectroscopy, line-scan fluorescence correlation spectroscopy, generalized polarization imaging, as well as thin layer and gas chromatography) to quantify the effect of EPC3 in biophysical models of the plasma membrane, as well as in cancer cell lines. Our results indicate that EPC3 affects lipid-lipid interactions in cellular membranes by decreasing lipid packing and increasing membrane disorder and fluidity. As a consequence of these alterations in the lateral organization of lipid bilayers, the diffusive dynamics of membrane proteins are also significantly increased. Taken together, these findings suggest that the mechanism of action of EPC3 could be linked to its effects on fundamental biophysical properties of lipid membranes, as well as on lipid metabolism in cancer cells.},
  language  = {en}
}
@article{PetrichDunsingBoboneetal.2021,
  author    = {Petrich, Annett and Dunsing, Valentin and Bobone, Sara and Chiantia, Salvatore},
  title     = {Influenza A M2 recruits M1 to the plasma membrane},
  series = {Biophysical journal : BJ / ed. by the Biophysical Society},
  volume    = {120},
  journal   = {Biophysical journal : BJ / ed. by the Biophysical Society},
  number    = {24},
  publisher = {Cell Press},
  address   = {Cambridge},
  issn      = {0006-3495},
  doi       = {10.1016/j.bpj.2021.11.023},
  pages     = {5478 -- 5490},
  year      = {2021},
  abstract  = {Influenza A virus (IAV) is a respiratory pathogen that causes seasonal epidemics with significant mortality. One of the most abundant proteins in IAV particles is the matrix protein 1 (M1), which is essential for the virus structural stability. M1 organizes virion assembly and budding at the plasma membrane (PM), where it interacts with other viral components. The recruitment of M1 to the PM as well as its interaction with the other viral envelope proteins (hemagglutinin [HA], neuraminidase, matrix protein 2 [M2]) is controversially discussed in previous studies. Therefore, we used fluorescence fluctuation microscopy techniques (i.e., scanning fluorescence cross-correlation spectroscopy and number and brightness) to quantify the oligomeric state of M1 and its interactions with other viral proteins in co-transfected as well as infected cells. Our results indicate that M1 is recruited to the PM by M2, as a consequence of the strong interaction between the two proteins. In contrast, only a weak interaction between M1 and HA was observed. M1-HA interaction occurred only in the event that M1 was already bound to the PM. We therefore conclude that M2 initiates the assembly of IAV by recruiting M1 to the PM, possibly allowing its further interaction with other viral proteins.},
  language  = {en}
}
@phdthesis{Petrich2023,
  author    = {Petrich, Annett},
  title     = {Quantitative fluorescence microscopy methods to investigate molecular interactions and dynamics in living cells},
  doi       = {10.25932/publishup-61180},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-611800},
  school      = {Universit{\"a}t Potsdam},
  pages     = {244},
  year      = {2023},
  abstract  = {Biomolecules such as proteins and lipids have vital roles in numerous cellular functions, including biomolecule transport, protein functions, cellular homeostasis and biomembrane integrity. Traditional biochemistry methods do not provide precise information about cellular biomolecule distribution and behavior under native environmental conditions since they are not transferable to live cell samples. Consequently, this can lead to inaccuracies in quantifying biomolecule interactions due to potential complexities arising from the heterogeneity of native biomembranes. To overcome these limitations, minimal invasive microscopic techniques, such as fluorescence fluctuation spectroscopy (FFS) in combination with fluorescence proteins (FPs) and fluorescence lipid analogs, have been developed. FFS techniques and membrane property sensors enable the quantification of various parameters, including concentration, dynamics, oligomerization, and interaction of biomolecules in live cell samples. In this work, several FFS approaches and membrane property sensors were implemented and employed to examine biological processes of diverse context. Multi-color scanning fluorescence fluctuation spectroscopy (sFCS) was used the examine protein oligomerization, protein-protein interactions (PPIs) and protein dynamics at the cellular plasma membrane (PM). Additionally, two-color number and brightness (N\&B) analysis was extended with the cross-correlation analysis in order to quantify hetero-interactions of proteins in the PM with very slow motion, which would not accessible with sFCS due strong initial photobleaching. Furthermore, two semi-automatic analysis pipelines were designed: spectral F{\"o}rster resonance energy transfer (FRET) analysis to study changes in membrane charge at the inner leaflet of the PM, and spectral generalized polarization (GP) imaging and spectral phasor analysis to monitor changes in membrane fluidity and order. An important parameter for studying PPIs is molecular brightness, which directly determines oligomerization and can be extracted from FFS data. However, FPs often display complex photophysical transitions, including dark states. Therefore, it is crucial to characterize FPs for their dark-states to ensure reliable oligomerization measurements. In this study, N\&B and sFCS analysis were applied to determine photophysical properties of novel green FPs under different conditions (i.e., excitation power and pH) in living cells. The results showed that the new FPs, mGreenLantern (mGL) and Gamillus, exhibited the highest molecular brightness at the cost of lower photostability. The well-established monomeric enhanced green fluorescent protein (mEGFP) remained the best option to investigate PPIs at lower pH, while mGL was best suited for neutral pH, and Gamillus for high pH. These findings provide guidance for selecting an appropriate FP to quantify PPIs via FFS under different environmental conditions. Next, several biophysical fluorescence microscopy approaches (i.e., sFCS, GP imaging, membrane charge FRET) were employed to monitor changes in lipid-lipid-packing in biomembranes in different biological context. Lipid metabolism in cancer cells is known to support rapid proliferation and metastasis. Therefore, targeting lipid synthesis or membrane integrity holds immense promise as an anticancer strategy. However, the mechanism of action of the novel agent erufosine (EPC3) on membrane stability is not fully under stood. The present work revealed that EPC3 reduces lipid packing and composition as well as increased membrane fluidity and dynamic, hence, modifies lipid-lipid-interaction. These effects on membrane integrity were likely triggered by modulations in lipid metabolism and membrane organization. In the case of influenza A virus (IAV) infection, regulation of lipid metabolism is crucial for multiple steps in IAV replication and is related to the pathogenicity of IAV. Here, it is shown for the first time that IAV infection triggers a local enrichment of negatively charged lipids at the inner leaflet of the PM, which decreases membrane fluidity and dynamic, as well as increases lipid packing at the assembly site in living cells. This suggests that IAV alters lipid-lipid interactions and organization at the PM. Overall, this work highlights the potential of biophysical techniques as a screening platform for studying membrane properties in living cells at the single-cell level. Finally, this study addressed remaining questions about the early stage of IAV assembly. The recruitment of matrix protein 1 (M1) and its interaction with other viral surface proteins, hemagglutinin (HA), neuraminidase (NA), and matrix protein 2 (M2), has been a subject of debate due to conflicting results. In this study, different FFS approaches were performed in transfected cells to investigate interactions between IAV proteins themselves and host factors at the PM. FFS measurements revealed that M2 interacts strongly with M1, leading to the translocation of M1 to the PM. This interaction likely took place along the non-canonical pathway, as evidenced by the detection of an interaction between M2 and the host factor LC3-II, leading to the recruitment of LC3-II to the PM. Moreover, weaker interaction was observed between HA and membrane-bound M1, and no interaction between NA and M1. Interestingly, higher oligomeric states of M1 were only detectable in infected cells. These results indicate that M2 initiates virion assembly by recruiting M1 to the PM, which may serve as a platform for further interactions with viral proteins and host factors.},
  language  = {en}
}
@article{FangHolzmuellerMatulaitisetal.2016,
  author    = {Fang, Lijia and Holzmueller, Felix and Matulaitis, Tomas and Baasner, Anne and Hauenstein, Christoph and Benduhn, Johannes and Schwarze, Martin and Petrich, Annett and Piersimoni, Fortunato and Scholz, Reinhard and Zeika, Olaf and Koerner, Christian and Neher, Dieter and Vandewal, Koen and Leo, Karl},
  title     = {Fluorine-containing low-energy-gap organic dyes with low voltage losses for organic solar cells},
  series = {Synthetic metals : the journal of electronic polymers and electronic molecular materials},
  volume    = {222},
  journal   = {Synthetic metals : the journal of electronic polymers and electronic molecular materials},
  publisher = {Elsevier},
  address   = {Lausanne},
  issn      = {0379-6779},
  doi       = {10.1016/j.synthmet.2016.10.025},
  pages     = {232 -- 239},
  year      = {2016},
  abstract  = {Fluorine-containing donor molecules TFTF, CNTF and PRTF are designed and isomer selectively synthesized for application in vacuum-deposited organic solar cells. These molecules comprise a donor acceptor molecular architecture incorporating thiophene and benzothiadiazole derivatives as the electron-donating and electron-withdrawing moieties, respectively. As opposed to previously reported materials from this class, PRTF can be purified by vacuum sublimation at moderate to high yields because of its higher volatility and better stabilization due to a stronger intramolecular hydrogen bond, as compared to TFTF and CNTF. The UV-vis absorption spectra of the three donors show an intense broadband absorption between 500 nm and 800 nm with, similar positions of their frontier energy levels. The photophysical properties of the three donor molecules are thoroughly tested and optimized in bulk heterojunction solar cells with C-60 as acceptor. PRTF shows the best performance, yielding power conversion efficiencies of up to 3.8\%. Moreover, the voltage loss for the PRTF device due to the non radiative recombination of free charge carriers is exceptionally low (0.26 V) as compared to typical values for organic solar cells (>0.34V). (C) 2016 Published by Elsevier B.V.},
  language  = {en}
}
@article{DunsingPetrichChiantia2021,
  author    = {Dunsing, Valentin and Petrich, Annett and Chiantia, Salvatore},
  title     = {Multicolor fluorescence fluctuation spectroscopy in living cells via spectral detection},
  series = {eLife},
  volume    = {10},
  journal   = {eLife},
  publisher = {eLife Sciences Publications},
  address   = {Cambridge},
  issn      = {2050-084X},
  doi       = {10.7554/eLife.69687},
  pages     = {33},
  year      = {2021},
  abstract  = {Signaling pathways in biological systems rely on specific interactions between multiple biomolecules. Fluorescence fluctuation spectroscopy provides a powerful toolbox to quantify such interactions directly in living cells. Cross-correlation analysis of spectrally separated fluctuations provides information about intermolecular interactions but is usually limited to two fluorophore species. Here, we present scanning fluorescence spectral correlation spectroscopy (SFSCS), a versatile approach that can be implemented on commercial confocal microscopes, allowing the investigation of interactions between multiple protein species at the plasma membrane. We demonstrate that SFSCS enables cross-talk-free cross-correlation, diffusion, and oligomerization analysis of up to four protein species labeled with strongly overlapping fluorophores. As an example, we investigate the interactions of influenza A virus (IAV) matrix protein 2 with two cellular host factors simultaneously. We furthermore apply raster spectral image correlation spectroscopy for the simultaneous analysis of up to four species and determine the stoichiometry of ternary IAV polymerase complexes in the cell nucleus.},
  language  = {en}
}
@inproceedings{DunsingPetrichChiantia2021,
  author    = {Dunsing, Valentin and Petrich, Annett and Chiantia, Salvatore},
  title     = {Spectral detection enables multi-color fluorescence fluctuation spectroscopy studies in living cells},
  series = {Biophysical journal : BJ / ed. by the Biophysical Society},
  volume    = {120},
  booktitle = {Biophysical journal : BJ / ed. by the Biophysical Society},
  number    = {3, Suppl. 1},
  publisher = {Cell Press},
  address   = {Cambridge},
  issn      = {0006-3495},
  doi       = {10.1016/j.bpj.2020.11.2206},
  pages     = {356A -- 356A},
  year      = {2021},
  language  = {en}
}