Association of neuregulin 1 with schizophrenia confirmed in a Scottish population

Recently, we identified neuregulin 1 (NRG1) as a susceptibility gene for schizophrenia in the Icelandic population, by a combined linkage and association approach. Here, we report the first study evaluating the relevance of NRG1 to schizophrenia in a population outside Iceland. Markers representing a core at-risk haplotype found in Icelanders at the 5' end of the NRG1 gene were genotyped in 609 unrelated Scottish patients and 618 unrelated Scottish control individuals. This haplotype consisted of five SNP markers and two microsatellites, which all appear to be in strong linkage disequilibrium. For the Scottish patients and control subjects, haplotype frequencies were estimated by maximum likelihood, using the expectation-maximization algorithm. The frequency of the seven-marker haplotype among the Scottish patients was significantly greater than that among the control subjects (10.2% vs. 5.9%, P=.00031). The estimated risk ratio was 1.8, which is in keeping with our report of unrelated Icelandic patients (2.1). Three of the seven markers in the haplotype gave single-point P values ranging from .000064 to .0021 for the allele contributing to the at-risk haplotype. This direct replication of haplotype association in a second population further implicates NRG1 as a factor that contributes to the etiology of schizophrenia.     

Vav1 and Ly-GDI two regulators of Rho GTPases,function cooperatively as signal transducers in T cell antigen receptor-induced pathways

The Rho family GTPases are pivotal for T cell signaling; however, the regulation of these proteins is not fully known. One well studied regulator of Rho GTPases is Vav1; a hematopoietic cell-specific guanine nucleotide exchange factor critical for signaling in T cells, including stimulation of the nuclear factor of activated T cells (NFAT). Surprisingly, Vav1 associates with Ly-GDI, a hematopoietic cell-specific guanine nucleotide dissociation inhibitor of Rac. Here, we studied the functional significance of the interaction between Vav1 and Ly-GDI in T cells. Upon organization of the immunological synapse, both Ly-GDI and Vav1 relocalize to T cell extensions in contact with the antigen-presenting cell. Ly-GDI is phosphorylated on tyrosine residues following T cell receptor stimulation, and it associates with the Src homology 2 region of an adapter protein, Shc. In addition, the interaction between Ly-GDI and Vav1 requires tyrosine phosphorylation. Overexpression of Ly-GDI alone is inhibitory to NFAT stimulation and calcium mobilization. However, when co-expressed with Vav1, Ly-GDI enhances Vav1 induction of NFAT activation, phospholipase Cgamma phosphorylation, and calcium mobilization. Moreover, Ly-GDI does not alter the regulation of these phenomena when coexpressed with oncogenic Vav1. Since oncogenic Vav1 does not bind Ly-GDI, this suggests that the functional cooperativity of Ly-GDI and Vav1 is dependent upon their association. Thus, our data suggest that the interaction of Vav1 and Ly-GDI creates a fine tuning mechanism for the regulation of intracellular signaling pathways leading to NFAT stimulation.     

A review of inner ear fate maps and cell lineage studies

A renewed interest in the development of the inner ear has provided more data on the fate and cell lineage relationships of the tissues making up this complex structure. The inner ear develops from a simple ectodermal thickening of the head called the otic placode, which undergoes a great deal of growth and differentiation to form a multichambered nonsensory epithelium that houses the six to nine sensory organs of the inner ear. Despite a large number of studies examining otic development, there have been surprisingly few fate maps generated. The published fate maps encompass four species and range from preotic to otocyst stages. Although some of these studies were consistent with a compartment and boundary model, other studies reveal extensive cell mixing during development. Cell lineage studies have been done in fewer species. At the single cell level the resulting clones in both chicks and frogs appear somewhat restricted in terms of distribution. We conclude that up until late placode stages there are no clear lineage restriction boundaries, meaning that cells seem to mix extensively at these early stages. At late placode stages, when the otic cup has formed, there are at least two boundaries located dorsally in the forming otocyst but none ventrally. These conclusions are consistent with all the fate maps and reconciles the chick and frog data. These results suggest that genes involved in patterning the inner ear may have dynamic and complex expression patterns.     

Effect of exercise at three exercise intensities on salivary cortisol

Changes in cortisol concentration in response to exercise at 3 different intensities were quantified. Ten apparently healthy, recreationally active males participated. On 4 separate occasions, subjects were assigned a random order of 1-hour cycle ergometer bouts of exercise at 44.5 +/- 5.5%, 62.3 +/- 3.8%, and 76.0 +/- 6.0% (mean +/- SD) of VO2peak and a resting control session. Saliva samples were collected before exercise at 10, 20, 40, and 59 minutes of exercise and at 20 minutes of recovery. Differences in cortisol concentration were assessed via multivariate analysis of variance (alpha = 0.05) Tukey post hoc analysis when indicated. During the highest-intensity exercise session, cortisol was significantly higher at 59 minutes of exercise (p = 0.004) and at 20 minutes of recovery (p = 0.016) than at those same time points during the resting control session. No significant differences in cortisol concentration were noted among resting, low-, and moderate-intensity exercise. Exercise <40 minutes in duration elicited no significant differences at any intensity. These data indicate that only exercise of high intensity and long duration results in significant elevations of salivary cortisol.     

The role of a clinically important mutation in the fold and RNA-binding properties of KH motifs

We have investigated the role in the fold and RNA-binding properties of the KH modules of a hydrophobic to asparagine mutation of clinical importance in the fragile X syndrome. The mutation involves a well-conserved hydrophobic residue close to the N terminus of the second helix of the KH fold (alpha2(3) position). The effect of the mutation has been long debated: Although the mutant has been shown to disrupt the three-dimensional fold of several KH domains, the residue seems also to be directly involved in RNA binding, the main function of the KH module. Here we have used the KH3 of Nova-1, whose structure is known both in isolation and in an RNA complex, to study in detail the role of the alpha2(3) position. A detailed comparison of Nova KH3 structure with its RNA/KH complex and with other KH structures suggests a dual role for the alpha2(3) residue, which is involved both in stabilizing the hydrophobic core and in RNA contacts. We further show by nuclear magnetic resonance (NMR) studies in solution that L447 of Nova-1 in position alpha2(3) is in exchange in the absence of RNA, and becomes locked in a more rigid conformation only upon formation of an RNA complex. This implies that position alpha2(3) functions as a "gate" in the mechanism of RNA recognition of KH motifs based on the rigidification of the fold upon RNA binding.     

The mechanical stability of ubiquitin is linkage dependent

Ubiquitin chains are formed through the action of a set of enzymes that covalently link ubiquitin either through peptide bonds or through isopeptide bonds between their C terminus and any of four lysine residues. These naturally occurring polyproteins allow one to study the mechanical stability of a protein, when force is applied through different linkages. Here we used single-molecule force spectroscopy techniques to examine the mechanical stability of N-C-linked and Lys48-C-linked ubiquitin chains. We combined these experiments with steered molecular dynamics (SMD) simulations and found that the mechanical stability and unfolding pathway of ubiquitin strongly depend on the linkage through which the mechanical force is applied to the protein. Hence, a protein that is otherwise very stable may be easily unfolded by a relatively weak mechanical force applied through the right linkage. This may be a widespread mechanism in biological systems.     

Combined histomorphometric and gene-expression profiling applied to toxicology

We have developed a unique methodology for the combined analysis of histomorphometric and gene-expression profiles amenable to intensive data mining and multisample comparison for a comprehensive approach to toxicology. This hybrid technology, termed extensible morphometric relational gene-expression analysis (EMeRGE), is applied in a toxicological study of time-varied vehicle- and carbon-tetrachloride (CCl₄)-treated rats, and demonstrates correlations between specific genes and tissue structures that can augment interpretation of biological observations and diagnosis.