TY  - JOUR
A1  - Groeneweg, Femke L.
A1  - van Royen, Martin E.
A1  - Fenz, Susanne
A1  - Keizer, Veer I. P.
A1  - Geverts, Bart
A1  - Prins, Jurrien
A1  - de Kloet, E. Ron
A1  - Houtsmuller, Adriaan B.
A1  - Schmidt, Thomas S.
A1  - Schaaf, Marcel J. M.
T1  - Quantitation of Glucocorticoid Receptor DNA-Binding Dynamics by Single-Molecule Microscopy and FRAP
JF  - PLOS ONE
N2  - Recent advances in live cell imaging have provided a wealth of data on the dynamics of transcription factors. However, a consistent quantitative description of these dynamics, explaining how transcription factors find their target sequences in the vast amount of DNA inside the nucleus, is still lacking. In the present study, we have combined two quantitative imaging methods, single-molecule microscopy and fluorescence recovery after photobleaching, to determine the mobility pattern of the glucocorticoid receptor (GR) and the mineralocorticoid receptor (MR), two ligand-activated transcription factors. For dexamethasone-activated GR, both techniques showed that approximately half of the population is freely diffusing, while the remaining population is bound to DNA. Of this DNA-bound population about half the GRs appeared to be bound for short periods of time (similar to 0.7 s) and the other half for longer time periods (similar to 2.3 s). A similar pattern of mobility was seen for the MR activated by aldosterone. Inactive receptors (mutant or antagonist-bound receptors) show a decreased DNA binding frequency and duration, but also a higher mobility for the diffusing population. Likely, very brief (<= 1 ms) interactions with DNA induced by the agonists underlie this difference in diffusion behavior. Surprisingly, different agonists also induce different mobilities of both receptors, presumably due to differences in ligand-induced conformational changes and receptor complex formation. In summary, our data provide a consistent quantitative model of the dynamics of GR and MR, indicating three types of interactions with DNA, which fit into a model in which frequent low-affinity DNA binding facilitates the search for high-affinity target sequences.
KW  - NF-KAPPA-B
KW  - image correlation spectroscopy
KW  - human mineralocorticoid receptor
KW  - nuclear-pore complexes
KW  - in-vivo
KW  - living cells
KW  - mobility
KW  - transcription
KW  - protein
KW  - reveals
Y1  - 2014
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-117085
VL  - 9
IS  - 3
ER  - 
TY  - JOUR
A1  - Serfling, Edgar
A1  - Rudolf, Ronald
A1  - Busch, Rhoda
A1  - Patra, Amiya K.
A1  - Muhammad, Khalid
A1  - Avots, Andris
A1  - Andrau, Jean-Christophe
A1  - Klein-Hessling, Stefan
T1  - Architecture and expression of the Nfatc1 gene in lymphocytes
N2  - In lymphocytes, the three NFAT factors NFATc1 (also designated as NFAT2), NFATc2 (NFAT1), and NFATc3 (NFAT4 or NFATx) are expressed and are the targets of immune receptor signals, which lead to a rapid rise of intracellular Ca++, the activation of phosphatase calcineurin, and to the activation of cytosolic NFATc proteins. In addition to rapid activation of NFAT factors, immune receptor signals lead to accumulation of the short NFATc1/αA isoform in lymphocytes which controls their proliferation and survival. In this mini-review, we summarize our current knowledge on the structure and transcription of the Nfatc1 gene in lymphocytes, which is controlled by two promoters, two poly A addition sites and a remote downstream enhancer. The Nfatc1 gene resembles numerous primary response genes (PRGs) induced by LPS in macrophages. Similar to the PRG promoters, the Nfatc1 promoter region is organized in CpG islands, forms DNase I hypersensitive sites, and is marked by histone tail modifications before induction. By studying gene induction in lymphocytes in detail, it will be important to elucidate whether the properties of the Nfatc1 induction are not only typical for the Nfatc1 gene but also for other transcription factor genes expressed in lymphocytes.
KW  - transcription
KW  - chromatin
KW  - induction
KW  - lymphocytes
KW  - Nfatc1
Y1  - 2014
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-112718
ER  -