Relationship between plasma aldosterone and left ventricular structure and function in patients with essential hypertension.

Recent experimental studies demonstrated that elevation of plasma aldosterone is associated with an increased collagen accumulation resulting in myocardial fibrosis. The aim of this study was to evaluate the association of circulating aldosterone with cardiac structural and functional changes in patients with essential hypertension. The authors examined 39 patients (aged 51 +/- 14 years, 13 male, 26 female) with essential hypertension. M-mode and two-dimensional echocardiography was performed in each subject. Plasma renin activity (PRA) and plasma aldosterone (PA) at baseline and after 4 hours upright posture were measured. Mean levels of baseline and stimulated PRA were similar in hypertensives and control group of normotensives. Baseline and stimulated PA levels were mildly but not significantly higher compared to control group. Using Tukey Kramer multiple analysis we found a significant positive correlation of both baseline and stimulated PA with echocardiographic parameters of left ventricle in hypertensive subjects: IVSd (p<0.001), PWd (p<0.001), LVIDd (p<0.001) and E/A (p<0.001). However, no correlation between PRA and left ventricular structure and function was found in this group of patients. The authors conclude that plasma aldosterone levels are related to the structure and function of left ventricle. (Tab. 3, Ref. 20.).

A small domain of CBP/p300 binds diverse proteins: solution structure and functional studies.

The transcriptional coactivators CBP and p300 are critical regulators of metazoan gene expression. They associate with many different DNA-bound transcription factors through small, conserved domains. We have identified a compactly folded 46 residue domain in CBP and p300, the IRF-3 binding domain (IBiD), and we have determined its structure by NMR. It has a helical framework containing an apparently flexible polyglutamine loop that participates in ligand binding. Spectroscopic data indicate that induced folding accompanies association of IBiD with its partners, which exhibit no evident sequence similarities. We demonstrate the significance both in vitro and in vivo of interactions between IBiD and a number of diverse partners. Thus, IBiD is an important contributor to signal integration by CBP and p300.

Assembly of a functional beta interferon enhanceosome is dependent on ATF-2-c-jun heterodimer orientation.

Heterodimeric transcription factors, including the basic region-leucine zipper (bZIP) protein ATF-2-c-jun, are well-characterized components of an enhanceosome that mediates virus induction of the human beta interferon (IFN-beta) gene. Here we report that within the IFN-beta enhanceosome the ATF-2-c-jun heterodimer binds in a specific orientation, which is required for assembly of a complex between ATF-2-c-jun and interferon regulatory factor 3 (IRF-3). We demonstrate that correct orientation of the ATF-2-c-jun binding site is required for virus induction of the IFN-beta gene and for IRF-3-dependent activation of a composite ATF-2- c-jun-IRF site in the IFN-beta promoter. We also show that in vitro the DNA-bound ATF-2-c-jun heterodimer adopts a fixed orientation upon the binding of IRF-3 at an adjacent site in the IFN-beta enhancer and that the DNA-binding domain of IRF-3 is sufficient to mediate this effect. In addition, we show that the DNA-binding domain of ATF-2 is necessary and sufficient for selective protein-protein interactions with IRF-3. Strikingly, in vivo chromatin immunoprecipitation experiments with IFN-beta reporter constructs reveal that recruitment of IRF-3 to the IFN-beta promoter upon virus infection is dependent on the orientation of the ATF-2-c-jun heterodimer binding site. These observations demonstrate functional and physical cooperativity between the bZIP and IRF transcription factor families and illustrate the critical role of heterodimeric transcription factors in formation of the IFN-beta enhanceosome.

Structural basis of DNA recognition by the heterodimeric cell cycle transcription factor E2F-DP.

The E2F and DP protein families form heterodimeric transcription factors that play a central role in the expression of cell cycle-regulated genes. The crystal structure of an E2F4-DP2-DNA complex shows that the DNA-binding domains of the E2F and DP proteins both have a fold related to the winged-helix DNA-binding motif. Recognition of the central c/gGCGCg/c sequence of the consensus DNA-binding site is symmetric, and amino acids that contact these bases are conserved among all known E2F and DP proteins. The asymmetry in the extended binding site TTTc/gGCGCc/g is associated with an amino-terminal extension of E2F4, in which an arginine binds in the minor groove near the TTT stretch. This arginine is invariant among E2Fs but not present in DPs. E2F4 and DP2 interact through an extensive protein-protein interface, and structural features of this interface suggest it contributes to the preference for heterodimers over homodimers in DNA binding.

Engrailed homeodomain-DNA complex at 2.2 A resolution: a detailed view of the interface and comparison with other engrailed structures.

We report the 2.2 A resolution structure of the Drosophila engrailed homeodomain bound to its optimal DNA site. The original 2.8 A resolution structure of this complex provided the first detailed three-dimensional view of how homeodomains recognize DNA, and has served as the basis for biochemical studies, structural studies and molecular modeling. Our refined structure confirms the principal conclusions of the original structure, but provides important new details about the recognition interface. Biochemical and NMR studies of other homeodomains had led to the notion that Gln50 was an especially important determinant of specificity. However, our refined structure shows that this side-chain makes no direct hydrogen bonds to the DNA. The structure does reveal an extensive network of ordered water molecules which mediate contacts to several bases and phosphates (including contacts from Gln50), and our model provides a basis for detailed comparison with the structure of an engrailed Q50K altered-specificity variant. Comparing our structure with the crystal structure of the free protein confirms that the N and C termini of the homeodomain become ordered upon DNA-binding. However, we also find that several key DNA contact residues in the recognition helix have the same conformation in the free and bound protein, and that several water molecules also are "preorganized" to contact the DNA. Our structure helps provide a more complete basis for the detailed analysis of homeodomain-DNA interactions.

Comparison of X-ray and NMR structures for the Antennapedia homeodomain-DNA complex.

Homeodomains are one of the key families of eukaryotic DNA-binding motifs and provide an important model system for studying protein-DNA interactions. We have crystallized the Antennapedia homeodomain-DNA complex and solved this structure at 2.4 A resolution. NMR and molecular dynamics studies had implied that this homeodomain achieves specificity through an ensemble of rapidly fluctuating DNA contacts. The crystal structure is in agreement with the underlying NMR data, but our structure reveals a well-defined set of contacts and also reveals the locations and roles of water molecules at the protein-DNA interface. The synthesis of X-ray and NMR studies provides a unified, general model for homeodomain-DNA interactions.

Aromatic-aromatic interactions in the zinc finger motif. Analysis of the two-dimensional nuclear magnetic resonance structure of a mutant domain.

The folding and stability of globular proteins are determined by a variety of chemical mechanisms, including hydrogen bonds, salt bridges and the hydrophobic effect. Of particular interest are weakly polar interactions involving aromatic rings, which are proposed to regulate the geometry of closely packed protein interiors. Such interactions reflect the electrostatic contribution of pi-electrons and, unlike van der Waals' interactions and the hydrophobic effect, may, in principle, introduce a directional force in a protein's hydrophobic core. Although the weakly polar hypothesis is supported by a statistical analysis of protein structures, the general importance of such contributions to protein folding and stability is unclear. Here, we show the presence of alternative aromatic-aromatic interactions in the two-dimensional nuclear magnetic resonance structure of a mutant Zn finger. Changes in aromatic packing lead in turn to local and non-local differences between the structures of a wild-type and mutant domain. The results provide insight into the evolution of Zn finger sequences and have implications for understanding how geometric relationships may be chemically encoded in a simple sequence template.