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001-es BibID:	BIBFORM060655
Első szerző:	Szalóki Nikoletta (biológus)
Cím:	Evidence for Homodimerization of the c-Fos Transcription Factor in Live Cells Revealed by Fluorescence Microscopy and Computer Modeling / Nikoletta Szalóki, Jan Wolfgang Krieger, István Komáromi, Katalin Tóth, György Vámosi
Dátum:	2015
ISSN:	0270-7306
Megjegyzések:	The c-Fos and c-Jun transcription factors, members of the activator protein 1 (AP-1) complex, form heterodimers and bind to DNA via a basic leucine zipper and regulate the cell cycle, apoptosis, differentiation, etc. Purified c-Jun leucine zipper fragments could also form stable homodimers, whereas c-Fos leucine zipper homodimers were found to be much less stable in earlier in vitro studies. The importance of c-Fos overexpression in tumors and the controversy in the literature concerning c-Fos homodimerization prompted us to investigate Fos homodimerization. Förster resonance energy transfer (FRET) and molecular brightness analysis of fluorescence correlation spectroscopy data from live HeLa cells transfected with fluorescent-protein-tagged c-Fos indicated that c-Fos formed homodimers. We developed a method to determine the absolute concentrations of transfected and endogenous c-Fos and c-Jun, which allowed us to determine dissociation constants of c-Fos homodimers (Kd = 6.7 ? 1.7 ?M) and c-Fos?c-Jun heterodimers (on the order of 10 to 100 nM) from FRET titrations. Imaging fluorescence cross-correlation spectroscopy (SPIM-FCCS) and molecular dynamics modeling confirmed that c-Fos homodimers were stably associated and could bind to the chromatin. Our results establish c-Fos homodimers as a novel form of the AP-1 complex that may be an autonomous transcription factor in c-Fos-overexpressing tissues and could contribute to tumor development.
Tárgyszavak:	Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban c-Fos homodimerization dissociation constant fluorescence correlation spectroscopy SPIM-FCCS FRET live cell imaging oncogene protein-protein interactions protein-DNA interactions
Megjelenés:	Molecular And Cellular Biology. - 35 : 21 (2015), p. 3785-3798. -
További szerzők:	Krieger, Jan Wolfgang Komáromi István (1957-) (vegyész, molekuláris biológus, biokémikus) Tóth Katalin (Heidelberg) Vámosi György (1967-) (biofizikus)
Pályázati támogatás:	K103965 OTKA TÁMOP-4.2.2.A-11/1/KONV-2012-0023 TÁMOP MÖB/21-1/2013 Egyéb TÁMOP-4.2.4.A/2- 11/1-2012-0001 TÁMOP
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001-es BibID:	BIBFORM065088
035-os BibID:	(Cikkazonosító)33022 (WOS)000382917700001 (Scopus)85047916061
Első szerző:	Vámosi György (biofizikus)
Cím:	EGFP oligomers as natural fluorescence and hydrodynamic standards / György Vámosi, Norbert Mücke, Gabriele Müller, Jan Wolfgang Krieger, Ute Curth, Jörg Langowski, Katalin Tóth
Dátum:	2016
ISSN:	2045-2322
Megjegyzések:	EGFP oligomers are convenient standards for experiments on fluorescent protein-taggedbiomolecules. In this study, we characterized their hydrodynamic and fluorescence properties.Diffusion coefficients D of EGFP1-4 were determined by analytical ultracentrifugation withfluorescence detection and by fluorescence correlation spectroscopy (FCS), yielding83.4?48.2 ?m2/s and 97.3?54.8 ?m2/s from monomer to tetramer. A "barrels standing in arow" model agreed best with the sedimentation data. Oligomerization red-shifted EGFPemission spectra without any shift in absorption. Fluorescence anisotropy increased,indicating homoFRET between the subunits. Fluorescence lifetime decreased only slightly(4%) indicating insignificant quenching by FRET to subunits in non-emitting states. FCSmeasuredD, particle number and molecular brightness depended on dark states and lightinducedprocesses in distinct subunits, resulting in a dependence on illumination powerdifferent for monomers and oligomers. Since subunits may be in "on" (bright) or "off" (dark)states, FCS-determined apparent brightness is not proportional to that of the monomer. Fromits dependence on the number of subunits, the probability of the "on" state for a subunit wasdetermined to 96% at pH8 and 77% at pH6.38, i.e., protonation increases the dark state. Thesefluorescence properties of EGFP oligomeric standards can assist interpreting results fromoligomerized EGFP fusion proteins of biological interest.
Tárgyszavak:	Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban folyóiratcikk oligo-EGFP FP photophysics diffusion coefficient molecular brightness protein oligomerization
Megjelenés:	Scientific Reports. - 6 (2016), p. 1-12. -
További szerzők:	Mücke, Norbert Müller, Gabriele Krieger, Jan Wolfgang Curth, Ute Langowski, Jörg Tóth Katalin (Heidelberg)
Internet cím:	Intézményi repozitóriumban (DEA) tárolt változat
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