Block of kainate receptor desensitization uncovers a key trafficking checkpoint

A prominent feature of ionotropic glutamate receptors from the AMPA and kainate subtypes is their profound desensitization in response to glutamate-a process thought to protect the neuron from overexcitation. In AMPA receptors, it is well established that desensitization results from rearrangements of the interface formed between agonist-binding domains of adjacent subunits; however, it is unclear how this mechanism applies to kainate receptors. Here we show that stabilization of the binding domain dimer by the generation of intermolecular disulfide bonds apparently blocked desensitization of the kainate receptor GluR6. This result establishes a common desensitization mechanism in both AMPA and kainate receptors. Surprisingly, however, surface expression of these nondesensitizing mutants was drastically reduced and did not depend on channel activity. Therefore, in addition to its role at the synapse, we now propose an intracellular role for desensitization in controlling maturation and trafficking of glutamate receptors.     

Genetic counseling for sex chromosome anomalies (SCAs) in Israel and Germany: assessing medical risks according to the importance of fertility in two cultures

In this article, I report findings from a comparative study of Israeli and German genetic counselors. Specifically, it concerns counselors' attitudes and risk assessments relating to prenatal diagnosis of sex chromosome anomalies (SCAs) such as Klinefelter and Turner syndromes. Data collected through in-depth interviews with counselors in both countries (N = 32) are presented, and the types of claims experts deploy in their personal and professional estimation of the risks involved in SCAs are analyzed. The article concludes by suggesting that the counselors rhetoric concerning SCAs, whose major manifestation is the future infertility of the unborn child as well as their estimations of the related risks, should be situated in a broader cultural context, that of local Israeli and German understandings of the importance of fertility, and not in their professional nondirective ethos. Hence, to understand the practice of genetic counselors in two late-modern societies, one must understand the unique relationship between the individual bodies of pregnant women and the body politics of their nations, a relationship mediated by the counselors, who are the bearers of knowledge and expertise in this field.     

Learning from stargazin: the mouse, the phenotype and the unexpected

The stargazin gene (also referred to as Cacng2) has been identified by forward genetics in a spontaneous mouse mutant with ataxic gait, upward head-elevating movements (hence the name stargazer for the mouse) and episodes of spike-wave discharges. Stargazin is related to the gamma-1 subunit of skeletal muscle voltage-dependent calcium channel (VDCC), and a deficit in its role as auxiliary VDCC subunit was proposed to underlie the epileptic phenotype of the mouse; yet, a conclusive demonstration of stargazin function in VDCC regulation is still lacking. In contrast, stargazin and its three closely related isoforms gamma-3, gamma-4 and gamma-8 were shown to function as auxiliary subunits for a very different ion channel - the AMPA-type glutamate receptor - prominently regulating early intracellular transport, synaptic targeting and anchoring, and ion channel functions of this major excitatory receptor in the brain.     

Matching Biochemical Reaction Kinetics to the Timescales of Life: Structural Determinants That Influence the Autodephosphorylation Rate of Response Regulator Proteins

In two-component regulatory systems, covalent phosphorylation typically activates the response regulator signaling protein, and hydrolysis of the phosphoryl group reestablishes the inactive state. Despite highly conserved three-dimensional structures and active-site features, the rates of catalytic autodephosphorylation for different response regulators vary by a factor of almost 10⁶. Previous studies identified two variable active-site residues, corresponding to Escherichia coli CheY residues 59 and 89, that modulate response regulator autodephosphorylation rates about 100-fold. Here, a set of five CheY mutants, which match other “model” response regulators (ArcA, CusR, DctD, FixJ, PhoB, or Spo0F) at variable active-site positions corresponding to CheY residues 14, 59, and 89, were characterized functionally and structurally in an attempt to identify mechanisms that modulate autodephosphorylation rate. As expected, the autodephosphorylation rates of the CheY mutants were reduced 6- to 40-fold relative to wild-type CheY, but all still autodephosphorylated 12- to 80-fold faster than their respective model response regulators. Comparison of X-ray crystal structures of the five CheY mutants (complexed with the phosphoryl group analogue BeF₃⁻) to wild-type CheY or corresponding model response regulator structures gave strong evidence for steric obstruction of the phosphoryl group from the attacking water molecule as one mechanism to enhance phosphoryl group stability. Structural data also suggested that impeding the change of a response regulator from the active to the inactive conformation might retard the autodephosphorylation reaction if the two processes are coupled, and that the residue at position ‘58’ may contribute to rate modulation. A given combination of amino acids at positions ‘14’, ‘59’, and ‘89’ adopted similar conformations regardless of protein context (CheY or model response regulator), suggesting that knowledge of residue identity may be sufficient to predict autodephosphorylation rate, and hence the kinetics of the signaling response, in the response regulator family of proteins.     

Over-expression of <Emphasis Type="Italic">CONSTANS-LIKE 5</Emphasis> can induce flowering in short-day grown <Emphasis Type="Italic">Arabidopsis</Emphasis>

To ensure that the initiation of flowering occurs at the correct time of year, plants need to integrate a diverse range of external and internal signals. In Arabidopsis, the photoperiodic flowering pathway is controlled by a set of regulators that include CONSTANS (CO). In addition, Arabidopsis plants also have a family of genes with homologies to CO known as CO-LIKE (COL) about which relatively little is known. In this paper, we describe the regulation and interactions of a novel member of the family, COL5. The expression of COL5 is under circadian and diurnal regulation, but COL5 itself does not appear to affect circadian rhythms. COL5, like CO, is regulated by GIGANTEA. Furthermore, COL5 is expressed in the vascular tissue. Using COL5 over-expressing lines we show that, under short days, constitutive expression of COL5 affects flowering time and the expression of the floral integrator genes, FLOWERING LOCUS T and SUPPRESSOR OF OVEREXPRESSION OF CO 1. Constitutive expression of COL5 partially suppresses the late flowering phenotype of the co-mutant plants. However, plants with loss of COL5 function do not show altered flowering. Taken together, our results suggest that COL5 has COL activity, but may either not have a role in regulating flowering in wild-type plants or may act redundantly with other flowering regulators. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A genome-wide screen for essential yeast genes that affect telomere length maintenance

Telomeres are structures composed of repetitive DNA and proteins that protect the chromosomal ends in eukaryotic cells from fusion or degradation, thus contributing to genomic stability. Although telomere length varies between species, in all organisms studied telomere length appears to be controlled by a dynamic equilibrium between elongating mechanisms (mainly addition of repeats by the enzyme telomerase) and nucleases that shorten the telomeric sequences. Two previous studies have analyzed a collection of yeast deletion strains (deleted for nonessential genes) and found over 270 genes that affect telomere length (Telomere Length Maintenance or TLM genes). Here we complete the list of TLM by analyzing a collection of strains carrying hypomorphic alleles of most essential genes (DAmP collection). We identify 87 essential genes that affect telomere length in yeast. These genes interact with the nonessential TLM genes in a significant manner, and provide new insights on the mechanisms involved in telomere length maintenance. The newly identified genes span a variety of cellular processes, including protein degradation, pre-mRNA splicing and DNA replication.     

Innexin genes and gap junction proteins in the locust frontal ganglion

Gap junctions (GJs) belong to one of the most conserved cellular structures in multicellular organisms. They probably serve similar functions in all Metazoa, providing one of the most common forms of intercellular communication. GJs are widely distributed in embryonic cells and tissues and have been attributed an important role in development, modulating cell growth and differentiation. These channels have been also implicated in mediating electrical synaptic signaling; Coupling through GJs is now accepted as a major pathway that supports network behavior and contributes to physiological rhythms. Here we focus on the physiology and molecular biology of GJs in a recently established model for the study of rhythm-generating networks and their role in behavior: the frontal ganglion (FG) of the desert locust, Schistocerca gregaria. Four novel genes of the invertebrate GJs (innexin) gene family were found to be expressed in the FG: Sg-inx1, Sg-inx2, Sg-inx3 and Sg-inx4. Immunohistochemistry revealed that some of the neurons in the FG express at least one innexin protein, INX1. We also established the presence of functional gap junction proteins in the FG and demonstrated functional electrical coupling between the neurons in the FG. This study forms the basis for further investigation of the role of GJs in network development and behavior.     

Complementary medicine in the primary care setting: Results of a survey of gender and cultural patterns in Israel

Objective: The purpose of this study was to examine the use of complementary and alternative medicine (CAM) in a primary care practice in Israel to determine prevalence and patterns of use.Methods: Trained research assistants invited all patients attending the administrative, medical, pharmaceutical, or nursing services of 7 clinics in urban and rural areas of northern Israel over a 16-month period, from April 1, 2005, through August 1, 2006, to complete a 13-item written questionnaire about CAM use and beliefs about CAM safety and efficacy. CAM was defined as therapies often referred to as alternative, complementary, natural, or folk/traditional medicine, and which are not usually offered as part of the medical treatment in the clinic, including herbal medicine, Chinese medicine (including acupuncture), homeopathy, folk and traditional remedies, dietary/nutritional therapy (including nutritional supplements), chiropractic, movement/manual healing therapies (including massage, reflexology, yoga, and Alexander and Feldenkrais techniques), mind-body techniques (including meditation, guided imagery, and relaxation), energy and healing therapies, and other naturopathic therapies. The Pearson χ² test and multivariate logistic regression were used to assess univariate associations with the odds ratios of CAM use among Arab and Jewish women. A t test was performed to determine whether there were any differences in the continuous variables between the 2 groups.Results: Of 3972 consecutive patients who received the questionnaire, 3447 responded; 2139 respondents (62%) were women. Of the female respondents, 2121 reported their religion (1238 respondents [58%] self-identified as being Arab, and 883 [41.6%] as being Jewish). Compared with men, more women used CAM during the previous year (46.4% vs 39.4%; P < 0.001). Women were more likely to use CAM and to be interested in receiving CAM at primary care clinics. Arab women reported less CAM use than Jewish women but were more interested in experiencing CAM, had a higher degree of confidence in CAM efficacy and safety, and more frequently supported the integration of CAM practitioners in primary care clinics.Conclusions: In this study, women visiting primary care clinics in northern Israel used CAM more often than men did. Arab women reported less use of CAM than did Jewish women but also reported greater confidence in CAM efficacy and safety.     

Na(+)- and H(+)-dependent motility in the coral pathogen Vibrio shilonii

In this work, we analyzed motility and the flagellar systems of the marine bacterium Vibrio shilonii. We show that this bacterium produces lateral flagella when seeded on soft agar plates at concentrations of 0.5% or 0.6%. However, at agar concentrations of 0.7%, cells become round and lose their flagella. The sodium channel blocker amiloride inhibits swimming of V. shilonii with the sheathed polar flagellum, but not swarming with lateral flagella. We also isolated and characterized the filament–hook–basal body of the polar flagellum. The proteins in this structure were analyzed by MS. Eight internal sequences matched with known flagellar proteins. The comparison of these sequences with the protein database from the complete genome of V. shilonii allows us to conclude that some components of the polar flagellum are encoded in two different clusters of flagellar genes, suggesting that this bacterium has a complex flagellar system, more complex possibly than other Vibrio species reported so far.