Functional analysis of String/CDC25 phosphatase in post-mitotic neurons

Cell cycle and differentiation are two highly coordinated processes during organ development. Recent studies have demonstrated that core cell cycle regulators also play cell cycle-independent functions in post-mitotic neurons, and are essential for the maintenance of neuronal homeostasis. CDC25 phosphatases are well-established CDK activators and their activity is mainly associated to proliferating tissues. The expression and activity of mammalian CDC25s has been reported in adult brains. However, their physiological relevance and the potential substrates in a non-proliferative context have never been addressed. string (stg) encodes the Drosophila CDC25 homolog. Previous studies from our group showed that stg is expressed in photoreceptors (PRs) and in lamina neurons, which are two differentiated cell types that compose the fly visual system. The aims of this work are to uncover the function of stg and to identify its potential neuronal substrates, using the Drosophila visual system as a model. To gain insight into the function of stg in a non-dividing context we used the GAL4/UAS system to promote downregulation of stg in PR-neurons, through the use of an RNAi transgene. The defects caused by stg loss-of-function were evaluated in the developing eye imaginal disc by immunofluorescence, and during adult stages by scanning electron microscopy. This genetic approach was combined with a specific proteomic method, two-dimensional difference gel electrophoresis (2D-DIGE), to identify the potential substrates in PR-cells. Our results showed that stg downregulation in PRs affects the well-patterned retina organization, inducing the loss of apical maintenance of PR-nuclei on the eye disc, and ommatidia disorganization. We also detected an abnormal accumulation of cytoskeletal proteins and a disruption of the axon structure. As a consequence, the projection of PR-axons into the lamina and medulla neuropils of the optic lobe was impaired. Upon stg downregulation, we also detected that PR-cells accumulate Cyclin B. Although the rough eye phenotype observed upon stg downregulation suggests neurodegeneration, we did not detect neuronal death during larval stages, suggesting that it likely occurs during pupal stages or during adulthood. By 2D-DIGE, we identified seven proteins which were differentially expressed upon stg downregulation, and are potential neuronal substrates of Stg. Altogether, our observations suggest that Stg phosphatase plays an essential role in the Drosophila visual system neurons, regulating several cell components and processes in order to ensure their homeostasis.

Integrated coastal geoengineering approach for maritime environments

This dissertation introduces several methodological approaches which integrate a proposed coastal management model in an interdisciplinary perspective. The research presented herein is displayed as a set of publications comprising different thematic outlooks. The thesis develops an integrated coastal geoengineering approach which is intrinsically linked to the studied maritime environments. From sandy coasts and marine works to rocky platforms and sea cliffs, this study includes field work between Caminha – Figueira da Foz (NW Portugal) and Galicia (NW Spain). The research also involves an analysis and geological-geotechnical characterisation of natural rock (armourstone) and artificial units (concrete blocks) applied to coastal structures. The main goal is to contribute to the characterisation and re-evaluation of georesources and to determine armourstone suitability and availability from its source (quarry). It was also important to diagnose the geomaterials in situ concerning their degradation/deterioration level on the basis of the current status of the coastal protection works in order to facilitate more efficient monitoring and maintenance, with economic benefits. In the rocky coast approach the coastal blocks were studied along the platform, but also the geoforms were studied from a coastal morphodynamics point of view. A shoreline evolution analysis was developed for sandy coasts through Digital Shoreline Analysis System (DSAS) extension. In addition, the spatial and statistical analysis applied to sea cliffs allowed the establishment of susceptibility zones to erosion and hazardous areas. All of these studies have different purposes and results however, there is a common denominator – GIS mapping. Hence, apart from the studied coastal environment, there is an integrated system which includes a sequence of procedures and methodologies that persisted during the research period. This is a step forward in the study of different coastal environments by using almost the same methodologies. This will allow the characterisation, monitoring and assessment of coastal protection works, rocky coasts, and shore platforms. With such data, it is possible to propose or recommend strategies for coastal and shoreline management based on several justifications in terms of social, economic, and environmental questions, or even provide a GIS-based planning support system reinforced by geocartographic decisions. Overall the development of the applied cartography embraces six stages which will allow the production of detailed maps of the maritime environment: (1) high-resolution aerial imagery surveys; (2) visual inspection and systematic monitoring; (3) applied field datasheet; (4) in situ evaluation; (5) scanline surveying; and (6) GIS mapping. This thesis covers fundamental matters that were developed over the course of scientific publication and as a consequence they represent the results obtained and discussed. The subjects directly related to the thesis architecture are: (i) cartography applied to coastal dynamics (including an art historical analysis as a tool to comprehend the coastal evolution and the littoral zone); (ii) georesources assessment (the role of cartography in georesources zoning, assessment and armourstone durability); (iii) coastal geoengineering applications and monitoring (Espinho pilot site in NW Portugal as an experimental field); (iv) rocky coast and shore platform studies and characterisation; (v) sandy and mixed environment approaches; (vi) coastal geosciences GIS mapping and photogrammetric surveying (coastal geoengineering); and (vii) shoreline change mapping and coastal management strategies (the CartGalicia Project as an example – NW Spain). Finally, all of these thematic areas were crucial to generate the conceptual models proposed and to shape the future of integrated coastal coastal geoengineering management.