Colour patterns are involved in many aspects of the lives of animals ranging from mate recognition and predator avoidance to thermoregulation. As pigmentation patterns are major functional end products in animals, their development must be tightly regulated. The biosynthesis of the most common pigment in animals, melanin, is well known. However, the ways by which pigmentation patterns are formed and evolved are poorly understood. Because pigmentation patterns are rapidly evolving traits in the Drosophila genus, they offer an ideal framework to study how modifications in gene regulation lead to phenotypic evolution. This proposal aims to investigate a key locus for melanin pattern evolution in Drosophilids, the yellow gene and its cis-regulatory elements. The wings of Drosophila biarmipes males display a melanin spot, which is important for sexual behaviour and selection. Yellow protein expression in the pupal wing correlates to this spot. In Drosophila melanogaster, Yellow protein is absent in the corresponding area and no spot is present. This is due to functional differences in the upstream regulatory region of yellow. Intriguingly, the transcription factors are present for making the spot in D. melanogaster. I propose to localize functional differences in cis-regulatory elements of the yellow gene of different Drosophila species and to identify the transcription factors that recognize sites that have changed.

The phenomenon that all members of a plant species exhibit resistance to the pathogens that infect other plant species is known as ‘non-host’ resistance. In Arabidopsis the mutation LRA5+-1 reduces non-host resistance to some bacterial and fungal phytopathogens. While there is no macroscopic phenotype associated with the mutation, transcriptomic data show that LRA5+-1 plants lack basal expression of defence transcripts, indicating that a central node in defence signalling is affected. LRA5+-1 has been mapped to a ~100 kb interval on chromosome II. Complementation and sequencing studies are in progress, and the cloning of LRA5+-1 is expected shortly. I will use biochemical approaches to elucidate the role of LRA5+-1 in non-host resistance. LRA5+-1 is a dominant gain-of-function mutation. Therefore both the mutant and the wild-type proteins will be analysed. To this end I will generate transgenic lines of Arabidopsis expressing epitope-tagged versions of LRA5+-1 and LRA5 (LRA5s). This will allow me to study the dynamics of the proteins in response to plant defence signalling molecules and pathogen challenge, and to identify proteins associated with LRA5s by coimmunoprecipitation. The molecular and biochemical approaches chosen to study LRAs are not only likely to unravel some of the mechanisms underlying non-host resistance they will also furnish me with firsthand knowledge of new techniques that will be of great value to me when I come to lead a research group of my own one day.

The proposed research aims at the design of hybrid structures composed of functional molecular building blocks and nanometer scale systems. Combining my previous background in molecular model systems and the host lab expertise in nanoscience will allow me to expand my scientific skills and scope. We suggest developing a versatile and robust architecture for studying fundamental biological and chemical processes. Understanding the fundamental processes and phenomena utilized by nature for efficiently carrying out catalysis, assembly, locomotion, and other vital mechanisms requires tools capable of differentiating the ensemble behavior and the underlying individual molecular properties. Thus, we will progress towards an experimental platform where the ‘sensing’ (i.e. observable response), and the studied motif can be tailored in a modular manner. Such capabilities are expected to be attainable by the combination of recent advances in nanodevices, chemical modification of surfaces, and biosynthesis in Prof. Lieber’s group and implementation of my previous research experience. The development of molecular functionalized nanowire hybrids that integrate the molecular-level complexity with an efficient transudation and enhancement of the information produced as a result of specific molecular interactions and response may provide a highly desired bridge between the molecular level physical environment and a well-resolved readout signal. Recent progress achieved at Prof. Lieber’s group demonstrated that the selective detection at single molecule level (that is, stochastic sensing) is possible with nanowire or nanotube field- effect transistors.

Antiandrogen (antagonist) therapy is the standard therapy for prostate cancer. But many men eventually fail and die of a relapse of the hormone-refractory, androgen-independent prostate cancer (AIPC). Understanding the mechanisms of AIPC is the key towards developing new therapies. Androgen receptor (AR) regulates gene expression in the presence of androgens (agonists) or antogonists, recruiting coactivator or corepressor complexes, respectively, to promoters. The host lab found that N-CoR corepressors are required for the inhibitory action of antagonists. The antagonism actions of currently used antagonists depend on N-CoR, or they function as agonists in the case of AIPC. So to identify N-CoR-independent compounds/ligands is the key to understand and prevent AIPC. I will identify antagonists in N-CoR-deficient cells, and use them to elucidate mechanisms of alternative coregulator assembly. The N-CoR-deficient prostate cancer cells will be employed in a screen of compounds libraries. Compounds will be scored in the high throughput, reporter-based screening as antagonists in those cells. Positives will be evaluated for their ability to repress AR target genes in AIPC cells, to recruit alternative corepressors and/or to release coactivators on promoters of AR target genes. The ability of compounds will be ultimately tested using cell culture and in vivo xenograft models of AIPC. This fellowship will provide me a thorough training in endocrinology/biochemistry, which is a perfect complement to my PhD study in genetics. Also it is a unique opportunity to conduct independent research at a top-ranking lab and will serve as a transition of my career to an independent scientist.

Glycosyltransferases, the enzymes that are responsible for the synthesis of the vast majority of oligosaccharides in the biosphere, are an attractive alternative to chemical approaches for the synthesis of complex oligosaccharides. My proposed research is to use directed evolution methodologies in order to engineer glycosyltransferases for the enzymatic synthesis of novel oligosaccharides. More specifically, I plan to evolve bacterial glycosyltransferases, to utilize readily synthesized and inexpensive glycosyl fluorides as donor sugars for transfer to a range of different glycosyl acceptors. In addition I propose to change the selectivity of the glycosyltransferases to accommodate different sugar acceptors and open the possibility of enzymatically synthesizing novel oligosaccharides. The engineered enzymes will shed light on the mechanisms of glycosyltransferases and the oligosaccharide products may be of value in the search for new therapeutic agents. The fellowship experience will give me the opportunity to conduct research in the large and important field of carbohydrate processing enzymes, which will be different from my previous research experience where I never dealt with carbohydrates or glycosyltransferases. I will be exposed, during the fellowship period, to a variety of ideas and techniques that are unique to the glycobiology world and this will enhance my professional growth, eventually as an independent investigator conducting research in this important field.

The overall structural organization of the nervous system is mostly bilaterally symmetric. However, deviations from this left/right symmetry are observed in for example different patterns of gene expression in otherwise symmetrical structures. In C. elegans, an interesting example of asymmetric gene expression is the chemosensory neurons ASER and ASEL. These neurons asymmetrically express 3 putative receptors, which correlates with different chemosensory activities. Several transcription factors have been shown by my host laboratory to work in a regulatory cascade that leads to the asymmetric expression of the receptors, however, these transcription factors have already an asymmetric activity. The aim of my project is to identify the initial signalling pathway that leads to the asymmetries observed in the L/R ASE neurons. To determine the molecular nature of the pathway I am presently undertaking a genetic screen. At this point I have already retrieved 16 asymmetry mutant. These mutants will be characterized and their epistatic relationship determined to examine their role in the process of asymmetry generation. I expect that the identification of neural asymmetry-inducing mechanisms will provide major insights into brain patterning. My proposed work will allow me to learn C. elegans genetics as well as how to address neuronal development issues. This will be a complete new field and approach for me and will complement my training in cell biology and biochemistry that I had during my PhD. In the future as an independent researcher I would like to continue investigating brain patterning complementing the genetic studies with cell biology and biochemistry techniques.

1) PROJECT: This project aims to study the conformational changes undergone by the cytosolic chaperonin CCT upon nucleotide binding and to measure the forces behind these changes. These forces may play an important role in forcing the substrate molecules to fold inside the chaperonin. A unique aspect of the studies proposed here involves the characterization of this system using methods of single molecule analysis. These methods have several advantages over those that utilise statistical ensembles of molecules because, unlike the latter, single molecule experiments make it possible to follow the molecular trajectory of the reaction and the signals are not confused by the simultaneous presence of multiple intermediates of the reaction. Moreover, single molecule manipulation methods make it possible to recognise and directly measure the forces generated during specific steps in a reaction, and even apply external force to alter the rate of these force-generating steps. 2) HOW IT DIFFERS FROM PREVIOUS RESEARCH EXPERIENCE: My previous research has been developed in the frame of applied physics, theoretical and focused on fundamental optical problems related with wave propagation, near field optics and optical forces. 3) PROFESSIONAL GROWTH: This project offers me the opportunity to work in a biological problem and extend my previous background to the derived applications in biology. It will also involve interpreting and handling with real assays, complementing my theoretical point of view with that of the experiments. In particular, I will train on novel biophysical techniques in single-molecule analysis, namely, optical tweezers and single-molecule fluorescence microscopy.

A living organism is a highly evolved system made up from a large number of interwoven molecular networks. These networks primarily involve proteins which enable and control every chemical process that takes place in the cell. I propose to study the behaviour of proteins in molecular simulations. The big challenge of such simulations is the so called time-scale gap. The folding time of a protein is in the range from milliseconds to seconds, whereas the longest molecular simulation of a protein is in the range of microseconds. I propose to use experimental data as a constrain in molecular simulations to overcome this time-scale gap. The ultimate goal of my research plan is to gain insight of the failures of cellular control and regulation processes that can lead to neurodegenerative diseases. This research will involve (i) the study of the folding and misfolding of proteins, (ii) the interaction of a misfolded protein with a cell membrane, and (iii) the binding of a protein with the receptor of a cell nucleus. In order to achieve these goals I will have to move into a new research area. I am a computational physicist with an extensive experience on crystal nucleation of colloidal suspensions. My proposed research involves computer simulations of complex biological systems. To carry out this research it is essential to get a deep understanding of the biological systems as well as to understand the experimental opportunities needed as input for the simulations. The support of this fellowship would give me the opportunity to become a biophysicist with a deep understanding of both physics and biology.