Ligand- and mutation-induced conformational selection in the CCR5 chemokine G protein-coupled receptor.

We predicted the structural basis for pleiotropic signaling of the C-C chemokine type 5 (CCR5) G protein-coupled receptor (GPCR) by predicting the binding of several ligands to the lower-energy conformations of the CCR5 receptor and 11 mutants. For each case, we predicted the ∼ 20 most stable conformations for the receptor along with the binding sites for four anti-HIV ligands. We found that none of the ligands bind to the lowest-energy apo-receptor conformation.

Regulation of dendritic development by neuron-specific chromatin remodeling complexes.

The diversity of dendritic patterns is one of the fundamental characteristics of neurons and is in part regulated by transcriptional programs initiated by electrical activity. We show that dendritic outgrowth requires a family of combinatorially assembled, neuron-specific chromatin remodeling complexes (nBAF complexes) distinguished by the actin-related protein BAF53b and based on the Brg/Brm ATPases. nBAF complexes bind tightly to the Ca(2+)-responsive dendritic regulator CREST and directly regulate genes essential for dendritic outgrowth.

Purification and identification of proteins that bind to the hereditary persistence of fetal hemoglobin -198 mutation in the gamma-globin gene promoter.

Expression of the gamma-globin gene is silenced in adult humans. However, certain point mutations in the gamma-globin gene promoter are capable of maintaining expression of this gene during adult erythropoiesis, a condition called non-deletion hereditary persistence of fetal hemoglobin (HPFH). Among these, the British form of HPFH carrying a T-->C point mutation at position -198 of the Agamma-globin gene promoter results in 4-10% fetal hemoglobin in heterozygotes.

Transcriptional potential of the gamma-globin gene is dependent on the CACCC box in a developmental stage-specific manner.

To test the role of CACCC box on gamma-globin gene activation, the CACCC box was deleted or mutated and gamma-gene expression was monitored in transgenic mice. Disruption of the CACCC box had no effect on gamma-gene expression in the cells of embryonic erythropoiesis but it strikingly reduced gamma-gene expression in fetal erythropoiesis, and abolished gamma-gene expression in adult erythroid cells. The CACCC mutation diminished HS formation, as well as TBP and polII recruitment at the gamma-gene promoter; however, it only resulted in slight or no effects on histone H3 and H4 acetylation in adult erythropoiesis.

Juxtaposition of the HPFH2 enhancer is not sufficient to reactivate the gamma-globin gene in adult erythropoiesis.

Previous studies have suggested that juxtaposition of a downstream enhancer to the fetal gamma-globin gene results in reactivation of the gamma-gene in adult erythrocytes of individuals with hereditary persistence of fetal hemoglobin (HPFH). To test the hypothesis in a much stricter basis, we produced beta locus YAC transgenic mice carrying an exact beta locus replicate of a deletional HPFH mutation, HPFH 2. Although the gamma-globin gene was expressed in the HPFH 2/beta locus YAC (HPFH2/YAC) transgenic mice in the early stage of development, it was completely silenced in the adult mice.

Identification of a polymorphic, neuron-specific chromatin remodeling complex.

A variety of chromatin remodeling complexes are thought to assist sequence-specific transcription factors. The complexes described to date are expressed ubiquitously, suggesting that they have general transcriptional functions. We show that vertebrate neurons have a specialized chromatin remodeling complex, bBAF, specifically containing the actin-related protein, BAF53b, which is first expressed in postmitotic neurons at about murine embryonic day 12.

Nuclear actin and actin-related proteins in chromatin remodeling.

The existence and function of actin in the nucleus has been hotly debated for forty years. Recently, beta-actin was found to be a component of mammalian SWI/SNF-like BAF chromatin remodeling complexes and still more recently other SWI/SNF-related chromatin remodeling complexes in yeast, flies, and man. Although the function of actin in these chromatin remodeling complexes is only starting to be explored, the fact that actin is one of the most regulated proteins in the cell suggests that control of nuclear actin may be a critical regulatory point in the control of chromatin remodeling.

Cloning and characterization of hELD/OSA1, a novel BRG1 interacting protein.

A highly conserved multisubunit enzymic complex, SWI/SNF, participates in the regulation of eukaryote gene expression through its ability to remodel chromatin. While a single component of SWI/SNF, Swi2 or a related protein, can perform this function in vitro, the other components appear to modulate the activity and specificity of the complex in vivo. Here we describe the cloning of hELD/OSA1, a 189 KDa human homologue of Drosophila Eld/Osa protein, a constituent of Drosophila SWI/SNF.