Identification and study of novel genes expressed in the heart precursor cells

Understanding heart development on a molecular level is a requirement for uncovering the causes of congenital heart diseases. Several genes have been implicated as critical for heart development. However, the inducers of these genes as well as their targets and pathways, remain largely unknown. We have identified a promoter element of chick cCer able to drive EGFP expression in a population of cells that consistently exit from the anterior primitive streak region, from as early as stage HH3+, and that later will populate the heart. Using this promoter element as a tool allowed us to identify novel genes previously not known to potentially play a role in heart development. In order to identify and study genes expressed and involved in the correct development and differentiation of the vertebrate heart precursor cell (HPC) lineages, a differential screening using Affymetrix GeneChip® system technologies was performed. Remarkably, this screening led to the identification of more than 700 transcripts differentially expressed in the heart forming regions (HFR). Bioinformatic tools allowed us to filter the large amount of data generated from this approach and to select a few transcripts for in vivo validation. Five genes were selected for further characterization by whole mount in situ hybridization leading to the validation of their expression in the HPC. From those, Adtk1 and Ccbe1 were selected for functional analysis. Regarding to ccbe1, a more detailed WISH analysis was performed and showed that Ccbe1 is expressed specifically on the cardiac progenitors regions at HH4, more specifically in primary heart field and at later stages is present in the second heart field. Further functional analyses by knockdown and overexpression revealed an important role for Ccbe1 in early heart tube formation. Moreover, the results presented in this thesis suggested that Ccbe1 is a key gene during heart development and might be limited to multipotent and highly proliferative progenitors and downregulated upon cellular commitment into more specific cardiac phenotypes. Other of the genes identified, Adtk1 was also subjected to further functional studies. Knockdown of Adtk1 using morpholino oligonucleotides suggested that it might be necessary for the migration and fusion of the heart tube as well as for neural tube closure.

Bycatch and discards of commercial trawl fisheries in the south coast of Portugal

Bycatch and discards are a cause of great concern in commercial world fisheries, with important ecological, economic and conservation implications. With the recent inclusion of a discards ban (‘landing obligation’), in the reform of the EU CFP, these issues have gained a tremendous attention from the economic, scientific, political and social point of view. Demersal trawl fisheries off the southern coast of Portugal capture an extraordinary diversity of species and generate considerable amounts of bycatch and discards. Bycatch includes commercially valuable target-species and bycatch species with low or no commercial value, but the great majority consists of unmarketable species, that are discarded. Bony fishes are dominant in bycatch and discards and the most discarded are of low or no commercial value. The reasons for discarding are fundamentally economic in nature (lack of commercial value) for bycatch species, and legal and administrative (legal minimum landing size) for commercially important species. The study of the reproductive biology of Galeus melastomus, discarded by crustacean trawls, suggests that a minimum landing size should be established for this species, and explains the importance of such a study in the assessment and management of fisheries. The discovery of a new species of the ray Neoraja iberica n. sp. contributes to the knowledge of the local marine biodiversity in Portuguese waters and of the global marine biodiversity. The three cases of abnormal hermaphroditism recorded in Etmopterus spinax, are the first cases known to date of hermaphroditism in this species. There is a need to find solutions to the problem of bycatch and discards of trawl fisheries in the Algarve coast. A combination of technical, regulatory and economic measures to minimize bycatch and reduce discards, before implementing a ‘landing obligation’, is thought to be the best approach to apply in the southern Portuguese multispecies trawl fisheries.