The metal ion binding protein Cnnm4 is mutated in rod-cone dystrophy/amelogenesis imperfecta syndrome

Purpose:To identify the gene causing rod-cone dystrophy/amelogenesis imperfecta Methods:Homozygosity mapping was performed using the Affymetrix 50K XbaI array in one family and candidate genes in the linked interval were sequenced with ABI Dye Terminator, vers. 1 in the index patient of 3 families. The identified mutations were screened in normal control individuals. Expression analyses were performed on RNA extracted from the brain, various parts of the eye and teeth; immunostaining was done on mouse eyes and jaw and knock-down experiments were carried out in zebrafish embroys. Results:Sequencing the coding regions of ancient conserved domain protein 4 (CNNM4), a metal ions transporter, revealed a 1-base pair duplication (p.L438fs) in family A, a p.R236Q mutation in family B and a p.L324P in family C. All these mutations were homozygous and involved very conserved amino acids in paralogs and orthologs. Immunostaining and RT-PCR confirmed that CNNM4 was strongly expressed in various parts of the eye and in the teeth. Morpholino experiments in zebrafish showed a loss of ganglion cells at 5 days post fertilization. Conclusions:The rod-cone dystrophy/amelogenesis imperfecta syndrome is caused by mutation in CNNM4 and is due to aberrant metal ion homeostasis.

Translating from India and the Moving Space of Translation (Illustrated by the Works of Ajñeya)

The inadequacies and obsolescence of Eurocentric theories based on a binaryand static worldview have become a staple topic of postcolonial studies, and tosome extent also of translation studies. Nonetheless, the literary texts that arecalled upon in order to show the dynamism and hybridity of (post)modern worksbelong for the most part to the languages of the former colonial powers, especiallyEnglish, and remain inserted in a system that construes literatures interms of opposition. As a consequence, there is outside India a doubly misleadingunderstanding of Indian literatures other than those written in English:firstly, that translations of works in Hindi and in the Indian bhāṣā seem to belacking, if not inexistent, and secondly, that these "minor" literatures - as theyare regularly termed - are still often viewed as being highly dependent on theidea of "tradition," in opposition to the "postmodern" hybridity of the literatureswritten in the "dominant" languages, such as English or French. Againstthese views and supported by the analysis of Ajñeya's works in Hindi togetherwith their English translations, this paper aims to show: 1) that translationsfrom Hindi, which are not in fact non-existent, are mainly carried out in India,and 2) that Ajñeya's works, while representing a significant instance of the effectivehybridity present in Indian literatures, help to illustrate the moving spaceof translation. This demonstration effectively invalidates the above-mentionedoppositional standpoint.

Effect of added dead space on sleep disordered breathing at high altitude.

OBJECTIVE: Sleep disordered breathing with central apnea or hypopnea frequently occurs at high altitude and is thought to be caused by a decrease in blood CO(2) level. The aim of this study was to assess the effects of added respiratory dead space on sleep disordered breathing.¦METHODS: Full polysomnographies were performed on 12 unacclimatized swiss mountaineers (11 males, 1 female, mean age 39±12 y.o.) in Leh, Ladakh (3500m). In random order, half of the night was spent with a 500ml increase in dead space through a custom designed full face mask and the other half without it.¦RESULTS: Baseline data revealed two clearly distinct groups: one with severe sleep disordered breathing (n=5, AHI>30) and the other with moderate to no disordered breathing (n=7, AHI<30). DS markedly improved breathing in the first group (baseline vs DS): apnea hypopnea index (AHI) 70.3±25.8 vs 29.4±6.9 (p=0.013), oxygen desaturation index (ODI): 72.9±24.1/h vs 42.5±14.4 (p=0.031), whereas it had no significant effect in the second group or in the total population. Respiratory events were almost exclusively central apnea or hypopnea. Microarousal index, sleep efficiency, and sleep architecture remained unchanged with DS. A minor increase in mean PtcCO(2) (n=3) was observed with DS.¦CONCLUSION: A 500ml increase in dead space through a fitted mask may improve nocturnal breathing in mountaineers with severe altitude-induced sleep disordered breathing.

Bax-induced apoptosis in Leber's congenital amaurosis : a dual role in rod and cone degeneration

Purpose:We previously observed that anti- and pro-apoptotic genes of the Bcl-2 family were differentially expressed during the development of LCA in the Rpe65-/- mouse model (Cottet et al. 2006). Moreover, we reported that activation and translocation of pro-apoptotic Bax to the mitochondria was associated with apoptosis of rod photoreceptors as the disease progressed (Cottet et al. 2008). In this study we challenged whether disruption of the pro-apoptotic pro-apoptotic Bax protein is sufficient to protect photoreceptor cells against apoptosis. Methods:Apoptosis of photoreceptor cells was addressed by TUNEL assay on flatmounted retinas. Counting of the rod nuclei within the ONL was performed following hematoxylin/eosin histological staining of retina sections. Expression level and localization of photoreceptor gene markers were assessed by quantitative PCR and immunohistological analyses. Results:While expression of rod photoreceptor genes was decreased in Rpe65-deficient retina, expression level remained unchanged in Rpe65-/- / Bax-/- mice. Moreover, OS dysorganization and shortening as well as decrease in ONL thickness observed in diseased retina were prevented in mice lacking functional Bax protein. TUNEL assay confirmed that Bax-dependent rod photoreceptor apoptosis was abolished in Rpe65-/- / Bax-/- mice. However, early and fast degeneration of cone cells was not prevented in Rpe65-/- / Bax-/- mice, indicating that Bax-induced apoptotic pathway was not involved in the degenerating process of cones in Rpe65-deficient retina. Conclusions:Altogether, these data show for the first time that a single genetic mutation can trigger two independent apoptotic pathways in rod and cone photoreceptors in LCA disease. While pro-apoptotic Bax is essential to trigger rod photoreceptor apoptosis, early degeneration of cones is not dependent on Bax-mediated apoptotic pathway in Rpe65-deficientmice.

The immune space: a concept and template for rationalizing vaccine development.

Abstract Empirical testing of candidate vaccines has led to the successful development of a number of lifesaving vaccines. The advent of new tools to manipulate antigens and new methods and vectors for vaccine delivery has led to a veritable explosion of potential vaccine designs. As a result, selection of candidate vaccines suitable for large-scale efficacy testing has become more challenging. This is especially true for diseases such as dengue, HIV, and tuberculosis where there is no validated animal model or correlate of immune protection. Establishing guidelines for the selection of vaccine candidates for advanced testing has become a necessity. A number of factors could be considered in making these decisions, including, for example, safety in animal and human studies, immune profile, protection in animal studies, production processes with product quality and stability, availability of resources, and estimated cost of goods. The "immune space template" proposed here provides a standardized approach by which the quality, level, and durability of immune responses elicited in early human trials by a candidate vaccine can be described. The immune response profile will demonstrate if and how the candidate is unique relative to other candidates, especially those that have preceded it into efficacy testing and, thus, what new information concerning potential immune correlates could be learned from an efficacy trial. A thorough characterization of immune responses should also provide insight into a developer's rationale for the vaccine's proposed mechanism of action. HIV vaccine researchers plan to include this general approach in up-selecting candidates for the next large efficacy trial. This "immune space" approach may also be applicable to other vaccine development endeavors where correlates of vaccine-induced immune protection remain unknown.

Rapid cohort generation and analysis of disease spectrum of large animal model of cone dystrophy.

Large animal models are an important resource for the understanding of human disease and for evaluating the applicability of new therapies to human patients. For many diseases, such as cone dystrophy, research effort is hampered by the lack of such models. Lentiviral transgenesis is a methodology broadly applicable to animals from many different species. When conjugated to the expression of a dominant mutant protein, this technology offers an attractive approach to generate new large animal models in a heterogeneous background. We adopted this strategy to mimic the phenotype diversity encounter in humans and generate a cohort of pigs for cone dystrophy by expressing a dominant mutant allele of the guanylate cyclase 2D (GUCY2D) gene. Sixty percent of the piglets were transgenic, with mutant GUCY2D mRNA detected in the retina of all animals tested. Functional impairment of vision was observed among the transgenic pigs at 3 months of age, with a follow-up at 1 year indicating a subsequent slower progression of phenotype. Abnormal retina morphology, notably among the cone photoreceptor cell population, was observed exclusively amongst the transgenic animals. Of particular note, these transgenic animals were characterized by a range in the severity of the phenotype, reflecting the human clinical situation. We demonstrate that a transgenic approach using lentiviral vectors offers a powerful tool for large animal model development. Not only is the efficiency of transgenesis higher than conventional transgenic methodology but this technique also produces a heterogeneous cohort of transgenic animals that mimics the genetic variation encountered in human patients.

A Novel Homozygous RPE65 Mutation in Leber Congenital Amaurosis Associated With Cone-Rod Dystrophy

Purpose: To report the clinical and genetic study of a family with Leber congenital amaurosis (LCA). Methods: We studied a consanguineous family from Yemen in which three individuals were affected with LCA. Genomic DNA was prepared from venous leukocytes. Linkage analysis of all family members using polymorphic markers flanking the known LCA genes was performed, followed by direct sequencing of all the exons and intron-exon junctions of the RPE65 gene. Results: The three affected were 5, 8 and 12 years old. Severe visual impairment and night blindness were noticed during infancy. Nystagmus was not a feature. Photophobia was only observed in the 8-year-old patient. The 5-year old youngest affected had a bilateral hyperopia of +3.50 and a visual acuity of 1/60. The oldest two had mild myopia and visual acuity limited to hand movements RE and counting fingers LE for the oldest and of 5/60 OD, 6/60 OS for the other. On fundus examination, they harbored common clinical features such as disc pallor, attenuated vessels, white flecks in the retina mid-periphery and bull's eye maculopathy. Electroretinograms of the oldest child were completely extinguished while residual scotopic responses with abolished photopic and flicker responses were observed in the two youngest. Sequencing identified a novel missense mutation, IVS2-3C>G, in the second RPE65 intron. The mutation was not detected in 80 ethnically matched normal individuals. Conclusion: We have identified a novel LCA-related homozygous RPE65 mutation associated with a severe clinical presentation including an early and severe cone dysfunction. This is in contrast with the presentation associated with other RPE65 mutations predominantly causing a rod-cone dystrophy with residual cone function. The identified mutation potentially affects splicing of the third exon and could result in a loss of function. Definite functional consequences of this change still need to be characterized.

Partition coefficient and molecular flexibility: the concept of lipophilicity space.

The rationale of this study was to investigate molecular flexibility and its influence on physicochemical properties with a view to uncovering additional information on the fuzzy concept of dynamic molecular structure. Indeed, it is now known that computed molecular interaction fields (MIFs) such as molecular electrostatic potentials (MEPs) and lipophilicity potentials (MLPs) are conformation-dependent, as are dipole moments. A database of 125 compounds was used whose conformational space was explored, while conformation-dependent parameters were computed for each non-redundant conformer found in the conformational space of the compounds. These parameters were the virtual log P (log P(MLP), calculated by a MLP approach), the apolar surface area (ASA), polar surface area (PSA), and solvent-accessible surface (SAS). For each compound, the range taken by each parameter (its property space) was divided by the number of rotors taken as an index of flexibility, yielding a parameter termed 'molecular sensitivity'. This parameter was poorly correlated with others (i.e., it contains novel information) and showed the compounds to fall into two broad classes. 'Sensitive' molecules are those whose computed property ranges are markedly sensitive to conformational effects, whereas 'insensitive' (in fact, less sensitive) molecules have property ranges which are comparatively less affected by conformational fluctuations. A pharmacokinetic application is presented.

Shift in cytotype frequency and niche space in the invasive plant Centaurea maculosa.

Polyploidy is often assumed to increase the spread and thus the success of alien plant species, but few empirical studies exist. We tested this hypothesis with Centaurea maculosa Lam., a species native to Europe and introduced into North America approximately 120 years ago where it became highly invasive. We analyzed the ploidy level of more than 2000 plants from 93 native and 48 invasive C. maculosa populations and found a pronounced shift in the relative frequency of diploid and tetraploid cytotypes. In Europe diploid populations occur in higher frequencies than tetraploids and only four populations had both cytotypes, while in North America diploid plants were found in only one mixed population and thus tetraploids clearly dominated. Our results showed a pronounced shift in the climatic niche between tetraploid populations in the native and introduced range toward drier climate in North America and a similar albeit smaller shift between diploids and tetraploids in the native range. The field data indicate that diploids have a predominately monocarpic life cycle, while tetraploids are often polycarpic. Additionally, the polycarpic life-form seems to be more prevalent among tetraploids in the introduced range than among tetraploids in the native range. Our study suggests that both ploidy types of C. maculosa were introduced into North America, but tetraploids became the dominant cytotype with invasion. We suggest that the invasive success of C. maculosa is partly due to preadaptation of the tetraploid cytotype in Europe to drier climate and possibly further adaptation to these conditions in the introduced range. The potential for earlier and longer seed production associated with the polycarpic life cycle constitutes an additional factor that may have led to the dominance of tetraploids over diploids in the introduced range.