
@article {128089,
	title = {Quantitative fluorescence emission anisotropy microscopy for implementing homo-fluorescence resonance energy transfer measurements in living cells},
	journal = {Mol Biol Cell.},
	volume = {34},
	year = {2023},
	month = {05/2023},
	abstract = {<p>Quantitative fluorescence emission anisotropy microscopy reveals the organization of fluorescently labeled cellular components and allows their characterization in terms of changes in either rotational diffusion or homo-F{\"o}rster{\textquoteright}s energy transfer characteristics in living cells. These properties provide insights into molecular organization, such as orientation, confinement, and oligomerization in situ. Here we elucidate how quantitative measurements of anisotropy using multiple microscope systems may be made by bringing out the main parameters that influence the quantification of fluorescence emission anisotropy. We focus on a variety of parameters that contribute to errors associated with the measurement of emission anisotropy in a microscope. These include the requirement for adequate photon counts for the necessary discrimination of anisotropy values, the influence of extinction ratios of the illumination source, the detector system, the role of numerical aperture, and excitation wavelength. All these parameters also affect the ability to capture the dynamic range of emission anisotropy necessary for quantifying its reduction due to homo-FRET and other processes. Finally, we provide easily implementable tests to assess whether homo-FRET is a cause for the observed emission depolarization.</p>
},
	doi = {10.1091/mbc.E22-09-0446.},
	author = {van Zanten, Thomas and Greeshma, Pradeep S and Mayor, S}
}
@article {91469,
	title = {Plk4 triggers autonomous de novo centriole biogenesis and maturation.},
	journal = {J Cell Biol},
	volume = {220},
	year = {2021},
	month = {2021 May 03},
	abstract = {<p>Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. Herein, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, our results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and found that proteins of the pericentriolar material (PCM), and in particular γ-tubulin, promote biogenesis, likely by locally concentrating critical components.</p>
},
	issn = {1540-8140},
	doi = {10.1083/jcb.202008090},
	author = {Nabais, Catarina and Pessoa, Delphine and de-Carvalho, Jorge and van Zanten, Thomas and Duarte, Paulo and Mayor, Satyajit and Carneiro, Jorge and Telley, Ivo A and Bettencourt-Dias, M{\'o}nica}
}
