ERC FUNDED PROJECTS

Understanding how proteins respond to mutations is of paramount importance to biology and disease. While protein stability and misfolding have been instrumental in rationalizing the impact of mutations, we recently discovered that an alternative route is also frequent, where mutations at the surface of symmetric proteins trigger novel self-interactions that lead to infinite self-assembly. This mechanism can be involved in disease, as in sickle-cell anemia, but may also serve in adaptation. Importantly, it differs fundamentally from aggregation, because misfolding does not drive it. Thus, we term it “agglomeration”. The ease with which agglomeration can occur, even by single point mutations, shifts the paradigm of how quickly new protein assemblies can emerge, both in health and disease. This prompts us to determine the basic principles of protein agglomeration and explore its implications in cell physiology and human disease. We propose an interdisciplinary research program bridging atomic and cellular scales to explore agglomeration in three aims: (i) Map the landscape of protein agglomeration in response to mutation in endogenous yeast proteins; (ii) Characterize how yeast physiology impacts agglomeration by changes in gene expression or cell state, and, conversely, how protein agglomerates impact yeast fitness. (iii) Analyze agglomeration in relation to human disease via two approaches. First, by predicting single nucleotide polymorphisms that trigger agglomeration, prioritizing them using knowledge from Aims 1 & 2, and characterizing them experimentally. Second, by providing a proof-of-concept that agglomeration can be exploited in drug design, whereby drugs induce its formation, like mutations can do. Overall, through this research, we aim to establish agglomeration as a paradigm for protein assembly, with implications for our understanding of evolution, physiology, and disease.

2 574 819 €

Start date: 2019-04-01, End date: 2024-03-31

When triggered by nutrient limitation, the Gram-positive bacterium Bacillus subtilis and its relatives enter a pathway of cellular differentiation culminating in the formation of a dormant cell type called a spore, the most resilient cell type known. Bacterial spores can survive for long periods of time and are able to endure extremes of heat, radiation and chemical assault. Remarkably, dormant spores can rapidly convert back to actively growing cells by a process called germination. Consequently, spore forming bacteria, including dangerous pathogens, (such as C. botulinum and B. anthracis) are highly resistant to antibacterial treatments and difficult to eradicate. Despite significant advances in our understanding of the process of spore formation, little is known about the nature of the mature spore. It is unrevealed how dormancy is maintained within the spore and how it is ceased, as the organization and the dynamics of the spore macromolecules remain obscure. The unusual biochemical and biophysical characteristics of the dormant spore make it a challenging biological system to investigate using conventional methods, and thus set the need to develop innovative approaches to study spore biology. We propose to explore the nature of spores by using B. subtilis as a primary experimental system. We intend to: (1) define the architecture of the spore chromosome, (2) track the complexity and fate of mRNA and protein molecules during sporulation, dormancy and germination, (3) revisit the basic notion of the spore dormancy (is it metabolically inert?), (4) compare the characteristics of bacilli spores from diverse ecophysiological groups, (5) investigate the features of spores belonging to distant bacterial genera, (6) generate an integrative database that categorizes the molecular features of spores. Our study will provide original insights and introduce novel concepts to the field of spore biology and may help devise innovative ways to combat spore forming pathogens.

1 630 000 €

Start date: 2008-10-01, End date: 2013-09-30

The metabolism of cancer cells is altered to meet cellular requirements for growth, providing novel means to selectively target tumorigenesis. While extensively studied, our current view of cancer cellular metabolism is fundamentally limited by lack of information on variability in metabolic activity between distinct subcellular compartments and cells. We propose to develop a spatio-temporal fluxomics approach for quantifying metabolic fluxes in the cytoplasm vs. mitochondria as well as their cell-cycle dynamics, combining mass-spectrometry based isotope tracing with cell synchronization, rapid cellular fractionation, and computational metabolic network modelling. Spatio-temporal fluxomics will be used to revisit and challenge our current understanding of central metabolism and its induced adaptation to oncogenic events – an important endeavour considering that mitochondrial bioenergetics and biosynthesis are required for tumorigenesis and accumulating evidences for metabolic alterations throughout the cell-cycle. Our preliminary results show intriguing oscillations between oxidative and reductive TCA cycle flux throughout the cell-cycle. We will explore the extent to which cells adapt their metabolism to fulfil the changing energetic and anabolic demands throughout the cell-cycle, how metabolic oscillations are regulated, and their benefit to cells in terms of thermodynamic efficiency. Spatial flux analysis will be instrumental for investigating glutaminolysis - a ‘hallmark’ metabolic adaptation in cancer involving shuttling of metabolic intermediates and cofactors between mitochondria and cytoplasm. On a clinical front, our spatio-temporal fluxomics analysis will enable to disentangle oncogene-induced flux alterations, having an important tumorigenic role, from artefacts originating from population averaging. A comprehensive view of how cells adapt their metabolism due to oncogenic mutations will reveal novel targets for anti-cancer drugs.

1 481 250 €

Start date: 2017-02-01, End date: 2022-01-31

The generation of animals by nuclear transfer demonstrated that the epigenetic state of somatic cells could be reset to an embryonic state, capable of directing the development of a new organism. The nuclear cloning technology is of interest for transplantation medicine, but any application is hampered by the inefficiency and ethical problems. A breakthrough solving these issues has been the in vitro derivation of reprogrammed Induced Pluripotent Stem “iPS” cells by the ectopic expression of defined transcription factors in somatic cells. iPS cells recapitulate all defining features of embryo-derived pluripotent stem cells, including the ability to differentiate into all somatic cell types. Further, recent publications have demonstrated the ability to directly trans-differentiate somatic cell types by ectopic expression of lineage specification factors. Thus, it is becoming increasingly clear that an ultimate goal in the stem cell field is to enable scientists to have the power to safely manipulate somatic cells by “reprogramming” their behavior at will. However, to frame this challenge, we must understand the basic mechanisms underlying the generation of reprogrammed cells in parallel to designing strategies for their medical application and their use in human disease specific research. In this ERC Starting Grant proposal, I describe comprehensive lines of experimentation that I plan to conduct in my new lab scheduled to open in April 2011 at the Weizmann Institute of Science. We will utilize exacting transgenic mammalian models and high throughput sequencing and genomic screening tools for in depth characterization of the molecular “rules” of rewiring the epigenome of somatic and pluripotent cell states. The proposed research endeavors will not only contribute to the development of safer strategies for cell reprogramming, but will also help decipher how diverse gene expression programs lead to cellular specification during normal development.

1 960 000 €

Start date: 2011-11-01, End date: 2016-10-31