Distribution of α-synuclein in normal human jejunum and its relations with the chemosensory and neuroendocrine system

Alpha-synuclein (α-syn) is a presynaptic neuronal protein and its structural alterations play an important role in the pathogenesis of neurodegenerative diseases, such as Parkinson’s disease (PD). It has been originally described in the brain and aggregated α-syn has also been found in the peripheral nerves including the enteric nervous system (ENS) of PD patients. ENS is a network of neurons and glia found in the gut wall which controls gastrointestinal function independently from the central nervous system. Moreover, two types of epithelial cells are crucial in the creation of an interface between the lumen and the ENS: they are the tuft cells and the enteroendocrine cells (EECs). In addition, the abundant enteric glial cells (EGCs) in the intestinal mucosa play a key role in controlling the intestinal epithelial barrier. Our aim was to localize and characterize the presence of α-syn in the normal human jejunal wall. Surgical specimens of proximal jejunum were collected from patients submitted to pancreaticoduodenectomy and intestinal sections underwent immunohistochemical procedure. Alpha-syn has been found both at the level of the ENS and the epithelial cells. To characterize α-syn immunoreactive epithelial cells, we used markers such as choline acetyltransferase (ChAT), useful for the identification of tuft cells. Then we evaluated the co-presence of α-syn with serotonin (5-HT), expressed in EECs. Finally, we used the low-affinity nerve growth factor receptor (p75NTR), to detect peripheral EGCs. The presence of α-syn has been demonstrated in EECs, but not in the tuft cells. Additionally, p75NTR has been highlighted in EECs of the mucosal layer and co-localized with α-syn in EECs but not with ChAT-positive cells. These findings suggest that α-syn could play a possible role in synaptic transmission of the ENS and may contribute to maintain the integrity of the epithelial barrier of the small intestine through EECs.Key words: Small intestine, jejunum, tuft cells, enteroendocrine cells (EECs), α-synuclein (α-syn), enteric     

The self-assembly and liquid crystal formation of d(GpGpApGpG)

d(GpGpApGpG) dissolved in water forms liquid crystalline phases of the cholesteric and hexagonal type. The building blocks of the phases are columnar four-stranded aggregates composed of G quartets. The diameter of the columnar aggregates is larger and the melting temperature is lower than for homoguanylic derivatives, indicating a distorsion and loss of stability due to the presence of adenines, which do not form a hydrogen-bonded quartet. The present study shows that, as often observed for single crystals, it is possible also in the liquid crystalline phase to construct long columnar aggregates composed of shorter segments of the blunt-end type in a “closed architecture.” © 1997 John Wiley & Sons, Inc. Biopoly 42: 561–574, 1997     

The IGF-1-IGF-1 Receptor System Modulates Myocyte Proliferation but Not Myocyte Cellular Hypertrophy in Vitro

In preliminary experiments it was established that the hypertrophic and hyperplastic responses of neonatal cardiac myocytes in culture were associated with enhanced expression of IGF-1 and IGF-1 receptors in these cells. Therefore, to determine the role of IGF-1 receptors on myocyte growth, cells were exposed to antisense oligodeoxynucleotides to IGF-1 receptor mRNA and the effects of this intervention on DNA synthesis, nuclear mitotic division, and changes in the number of myocytes were measured. Moreover, the influence of this procedure on ANF induction and myocyte cell volume was examined. Inhibition of the formation of IGF-1 receptors on myocytes suppressed DNA replication, mitosis, and cell proliferation. In contrast, the antisense treatment did not alter the expression of ANF in myocytes or cellular hypertrophy. Finally, IGF-1 stimulated DNA synthesis in myocytes cultured in serum-free medium, without inducing cellular hypertrophy. In conclusion, ligand activation of IGF-1 receptors on myocytes appears to be coupled with cell proliferation, whereas myocyte cellular hypertrophy seems to be independent from this effector pathway.     

Cloning, Expression, and Chromosomal Assignment of the Human Mitochondrial Intermediate Peptidase Gene (MIPEP)

The mitochondrial intermediate peptidase ofSaccharomyces cerevisiae(YMIP) is a component of the yeast mitochondrial protein import machinery critically involved in the biogenesis of the oxidative phosphorylation (OXPHOS) system. This leader peptidase removes specific octapeptides from the amino terminus of nuclear-encoded OXPHOS subunits and components of the mitochondrial genetic apparatus. To address the biologic role of the human peptidase [MIPEP gene, HMIP polypeptide], we have initiated its molecular and functional characterization. A full-length cDNA was isolated by screening a human liver library using a rat MIP (RMIP) cDNA as a probe. The encoded protein contained a typical mitochondrial leader peptide and showed 92 and 54% homology to RMIP and YMIP, respectively. A survey of human mitochondrial protein precursors revealed that, similar to YMIP, HMIP is primarily involved in the maturation of OXPHOS-related proteins. Northern analysis showed that the MIPEP gene is differentially expressed in human tissues, with the highest levels of expression in the heart, skeletal muscle, and pancreas, three organ systems that are frequently affected in OXPHOS disorders. Using fluorescencein situhybridization, the MIPEP locus was assigned to 13q12. This information offers the possibility of testing the potential involvement of HMIP in the pathophysiology of nuclear-driven OXPHOS disorders.     

The involvement of Alu repeats in recombination events at the α-globin gene cluster: characterization of two α°-thalassaemia deletion breakpoints

Alu repetitive sequences are frequently involved in homologous and non-homologous recombination events in the α-cluster. Possible mechanisms involved in Alu-mediated recombination events are strand exchange, promoted by DNA pairing between highly homologous Alu repeats, and subsequent strand invasion. Alternatively, Alu sequences might play a more active role in recombinogenic processes in the α-cluster. We describe a novel 33-kb α°-thalassaemia deletion ––^DUTCH encompassing the α- and zeta-globin genes and pseudogenes in a kindred of Dutch-Caucasian origin. This deletion appears similar, although not identical, to the previously described ––^MEDII deletion. Cloning and sequencing of both the ––^DUTCH and ––^MEDII deletion breakpoints clearly indicate that the mechanism leading to these α°-thalassaemia deletions involves misalignment between the highly homologous tandemly arranged Alu repeats at both parental sides, which are normally 33 kb apart. Comparison of breakpoint positions along the Alu consensus sequence indicate the involvement of a 26-bp core sequence in two out of five α°-thalassaemia deletions. This sequence has been identified by others as a possible hotspot of recombination. These findings favour the idea that Alu repeats stimulate recombination events not only by homologous pairing, but also by providing binding sites for recombinogenic proteins. Received: 14 October 1996 / Revised: 14 November 1996     

Novel, fluorescent, SSB protein chimeras with broad utility

The Escherichia coli single-stranded DNA binding protein (SSB) is a central player in DNA metabolism where it organizes genome maintenance complexes and stabilizes single-stranded DNA (ssDNA) intermediates generated during DNA processing. Due to the importance of SSB and to facilitate real-time studies, we developed a dual plasmid expression system to produce novel, chimeric SSB proteins. These chimeras, which contain mixtures of histidine-tagged and fluorescent protein(FP)-fusion subunits, are easily purified in milligram quantities and used without further modification, a significant enhancement over previous methods to produce fluorescent SSB. Chimeras retain the functionality of wild type in all assays, demonstrating that SSB function is unaffected by the FPs. We demonstrate the power and utility of these chimeras in single molecule studies providing a great level of insight into the biochemical mechanism of RecBCD. We also utilized the chimeras to show for the first time that RecG and SSB interact in vivo. Consequently, we anticipate that the chimeras described herein will facilitate in vivo, in vitro and single DNA molecule studies using proteins that do not require further modification prior to use.     

Governance factors in the identification of global conservation priorities for mammals

Global conservation priorities have often been identified based on the combination of species richness and threat information. With the development of the field of systematic conservation planning, more attention has been given to conservation costs. This leads to prioritizing developing countries, where costs are generally low and biodiversity is high. But many of these countries have poor governance, which may result in ineffective conservation or in larger costs than initially expected. We explore how the consideration of governance affects the selection of global conservation priorities for the world's mammals in a complementarity-based conservation prioritization. We use data on Control of Corruption (Worldwide Governance Indicators project) as an indicator of governance effectiveness, and gross domestic product per capita as an indicator of cost. We show that, while core areas with high levels of endemism are always selected as important regardless of governance and cost values, there are clear regional differences in selected sites when biodiversity, cost or governance are taken into account separately. Overall, the analysis supports the concentration of conservation efforts in most of the regions generally considered of high priority, but stresses the need for different conservation approaches in different continents owing to spatial patterns of governance and economic development.     

Reconciling global mammal prioritization schemes into a strategy

The huge conservation interest that mammals attract and the large datasets that have been collected on them have propelled a diversity of global mammal prioritization schemes, but no comprehensive global mammal conservation strategy. We highlight some of the potential discrepancies between the schemes presented in this theme issue, including: conservation of species or areas, reactive and proactive conservation approaches, conservation knowledge and action, levels of aggregation of indicators of trend and scale issues. We propose that recently collected global mammal data and many of the mammal prioritization schemes now available could be incorporated into a comprehensive global strategy for the conservation of mammals. The task of developing such a strategy should be coordinated by a super-partes, authoritative institution (e.g. the International Union for Conservation of Nature, IUCN). The strategy would facilitate funding agencies, conservation organizations and national institutions to rapidly identify a number of short-term and long-term global conservation priorities, and act complementarily to achieve them.