Functional studies on BolA and related genes: increasing the understanding of a protein with pleiotropic effects

Dissertation presented to obtain a Doctoral degree in Biology by Instituto de Tecnologia Química e Biológica

Cilia motility studies in zebrafish embryos

A thesis submitted in fulfilment of the requirements for the degree of Masters in Molecular Genetics and Biomedicine

The role of Rac1-modulated gene transcription in tumorigenesis

Dissertation presented to obtain the Ph.D degree in Biology

Phylogeny and phenotypic characterization of pathogenic cryptococcus species and closely related saprobic taxa in the tremellales

Eukaryotic Cell, Vol.8, Nº3

Identification of mating type genes in the bipolar basidiomycetous yeast rhodosporidium toruloides: first insight into the MAT Locus structure of the sporidiobolales

Eukaryotic Cell, Vol.7, Nº6

The genetic basis of morphological change in convergent evolution of natural populations: identifying candidate genes behind convergent evolution in blind cavefish astyanax mexicanus

Dissertation presented to obtain the Ph.D. degree in Biology at the Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa.

Epidemiological characterization, antimicrobial resistance and virulence mechanisms of human and animal streptococci

Dissertação para obtenção do Grau de Doutor em Biologia

New mechanisms that regulate the expression of genes implicated in the process of ketogenesis

Dissertação para obtenção do Grau de Mestre em Biotecnologia

Understanding the relationship between central metabolism and virulence in the human pathogen Streptococcus pneumoniae

Dissertation presented to obtain the Ph.D. degree in Biochemistry

Regulation of autoimmune neuroinflammation by stress-responsive genes

Dissertation presented to obtain the Ph.D degree in Biology

Non-clinical isolates bring new findings on enterococcal virulence

Dissertation presented to obtain the PhD degree in Biology

Signal transduction pathways involving the hypertension-related WNK1 and WNK4 protein kinases

Dissertação apresentada para obtenção do Grau de Doutor em Biologia, na especialidade de Genética Molecular, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Inhibition of SNF1-related protein kinase1 by trehalose 6-phosphate and other metabolites and the interrelation with plant gorwth

All life forms need to monitor carbon and energy availability to survive and this is especially true for plants which must integrate unavoidable environmental conditions with metabolism for cellular homeostasis maintenance. Sugars, in the heart of metabolism, are now recognized as crucial signaling molecules that translate those conditions. One such signal is trehalose 6- phosphate (T6P), a phosphorylated dimer of glucose molecules which levels correlate well with those of sucrose (Suc). Central integrators of stress and energy regulation include the conserved plant Snf1-related kinase1 (SnRK1) which respond to low cellular energy levels by up-regulating energy conserving and catabolic metabolism and down-regulating energy consuming processes. In 2009 T6P was shown to inhibit SnRK1. The in vitro inhibition of SnRK1 by T6P was confirmed in vivo through the observation that genes normally induced by SnRK1 were repressed by T6P and vice-versa, promoting growth processes. These observations provided a model for the regulation of growth by sugar.(...)

Evolutionary dynamics of Chlamydia trachomatis genome and identification of molecular patterns of hypothetical protein coding genes

The obligate intracellular bacterium Chlamydia trachomatis is a human pathogen of major public health significance. Strains can be classified into 15 main serovars (A to L3) that preferentially cause ocular infections (A-C), genital infections (D-K) or lymphogranuloma venereum (LGV) (L1-L3), but the molecular basis behind their distinct tropism, ecological success and pathogenicity is not welldefined. Most chlamydial research demands culture in eukaryotic cell lines, but it is not known if stains become laboratory adapted. By essentially using genomics and transcriptomics, we aimed to investigate the evolutionary patterns underlying the adaptation of C. trachomatis to the different human tissues, given emphasis to the identification of molecular patterns of genes encoding hypothetical proteins, and to understand the adaptive process behind the C. trachomatis in vivo to in vitro transition. Our results highlight a positive selection-driven evolution of C. trachomatis towards nichespecific adaptation, essentially targeting host-interacting proteins, namely effectors and inclusion membrane proteins, where some of them also displayed niche-specific expression patterns. We also identified potential "ocular-specific" pseudogenes, and pointed out the major gene targets of adaptive mutations associated with LGV infections. We further observed that the in vivo-derived genetic makeup of C. trachomatis is not significantly compromised by its long-term laboratory propagation. In opposition, its introduction in vitro has the potential to affect the phenotype, likely yielding virulence attenuation. In fact, we observed a "genital-specific" rampant inactivation of the virulence gene CT135, which may impact the interpretation of data derived from studies requiring culture. Globally, the findings presented in this Ph.D. thesis contribute for the understanding of C.trachomatis adaptive evolution and provides new insights into the biological role of C. trachomatishypothetical proteins. They also launch research questions for future functional studies aiming toclarify the determinants of tissue tropism, virulence or pathogenic dissimilarities among C. trachomatisstrains.

Gold nanoprobes for assessing expression of critical genes in the infection pathway of MRSA

Staphylococcus aureus is an important opportunistic pathogen that can cause a wide variety of diseases from mild to life-threatening conditions. S. aureus can colonize many parts of the human body but the anterior nares are the primary ecological niche. Its clinical importance is due to its ability to resist almost all classes of antibiotics available together with its large number of virulence factores. MRSA (Methicillin-Resistant S. aureus) strains are particularly important in the hospital settings, being the major cause of nosocomial infections worldwide. MRSA resistance to β-lactam antibiotics involves the acquisition of the exogenous mecA gene, part of the SCCmec cassette. Fast and reliable diagnostic techniques are needed to reduce the mortality and morbidity associated with MRSA infections, through the early identification of MRSA strains. The current identification techniques are time-consuming as they usually involves culturing steps, taking up to five days to determine the antibiotic resistance profile. Several amplification-based techniques have been developed to accelerate the diagnosis. The aim of this project was to develop an even faster methodology that bypasses the DNA amplification step. Gold-nanoprobes were developed and used to detect the presence of mecA gene in S. aureus genome, associated with resistance traits, for colorimetric assays based on non-crosslinking method. Our results showed that the mecA and mecA_V2 gold-nanoprobes were sensitive enough to discriminate the presence of mecA gene in PCR products and genomic DNA (gDNA) samples for target concentrations of 10 ng/μL and 20 ng/μL, respectively. As our main objective was to avoid the amplification step, we concluded that the best strategy for the early identification of MRSA infection relies on colorimetric assays based on non-crosslinking method with gDNA samples that can be extracted directly from blood samples.