We aim to understand the molecular basis of gene regulation specificity in Drosophila using genetics, genomics, biochemistry and live-imaging tools.
Research
In multicellular organisms, genes are turned ON and OFF in robust spatial and temporal patterns. Regulatory elements that activate or silence gene transcription are inherently promiscuous and can act over long genomic distances. A basic question is how regulatory elements find their target promoters and restrict their activity to their targets.
On the one hand, we study how regulatory crosstalk is prevented. Insulators were discovered in Drosophila and subsequently in various organisms from yeast to humans, as DNA elements that block communication between a regulatory element and a gene promoter when placed in between. In flies and humans, insulators exert their activity by recruiting insulator proteins. We study the biological relevance and molecular mechanisms of insulator proteins.
On the other hand, we study how genes find their appropriate regulatory elements across surprisingly long genomic distances and skip over non-target genes. We address the biological relevance of these interactions by disrupting them, and we study how they form.
Our studies in flies reveal new evolutionary perspectives into the relevance of 3D genome folding for correctly wiring genes to their regulatory elements.
Publications
Click on red arrow for access to datasets and code, click on green arrow for paper summary.
Protocol for detecting genomic insulators in Drosophila using insulator-seq, a massively parallel reporter assay
Anastasiia Tonelli*, Pascal Cousin*, Maria Cristina Gambetta**
Enhancer-promoter interactions become more instructive in the transition from cell-fate specification to tissue differentiation
Tim Pollex, Adam Rabinowitz, Maria Cristina Gambetta, Raquel Marco-Ferreres, Rebecca R Viales, Aleksander Jankowski, Christoph Schaub, Eileen E M Furlong
Chromosome-level organization of the regulatory genome in the Drosophlia nervous system
Giriram Mohana*, Julien Dorier*, Xiao Li*, Marion Mouginot, Rebecca C. Smith, Héléna Malek, Marion Leleu, Daniel Rodriguez, Jenisha Khadka, Patrycja Rosa, Pascal Cousin, Christian Iseli, Simon Restrepo, Nicolas Guex, Brian D. McCabe, Aleksander Jankowski**, Michael S. Levine**, Maria Cristina Gambetta**
Whereas CTCF and other DNA-binding proteins only bind to a subset of fly contact domain boundaries, Cp190 is present at a large fraction of boundaries.
Flies use Cp190 as an adaptor protein recruited by DNA-bound proteins such as CTCF and others to form boundaries.
DNA-bound CTCF cannot form a robust boundary in the absence of Cp190 (despite CTCF’s conserved ability to directly interact with cohesin).
Overall, Cp190 is required to form most promoter-distal boundaries but is dispensable to form promoter-proximal boundaries.
Cp190 is thus currently the major player in fly boundary formation.
Cp190 assembles into diverse multisubunit complexes that share similar enhancer-blocking activity in a quantitative insulator reporter assay.
Cp190 is essential for early development and prevents regulatory cross-talk between specific gene loci that pattern the fly embryo.
Cp190 is, in contrast, dispensable for long-range enhancer-promoter communication at tested loci.
This work demonstrates that diverse mechanisms evolved to partition genomes into independent physical and regulatory domains.
This work also reveals that the enhancer-blocking and enhancer-enabling functions typically ascribed to fly insulators are separable.
Datasets
NGS
Raw and processed data (Hi-C, NG Capture-C, and ChIP-seq) GEO: GSE180376
3D genomics across the tree of life reveals condensin II as a determinant of architecture type
C. Hoencamp, O. Dudchenko, A.M.O. Elbatsh, S. Brahmachari, J.A. Raaijmakers, T. van Schaik, A.S. Cacciatore, V.G. Contessoto, R.G.H.P. van Heesbeen, B. van den Broek, A.N. Mhaskar, H. Teunissen, B.G. St Hilaire, D. Weisz, A.D. Omer, M. Pham, Z. Colaric, Z.Z. Yang, S.S.P. Rao, N. Mitra, C. Lui, W.J. Yao, R. Khan, L.L. Moroz, A. Kohn, J. St Leger, A. Mena, K. Holcroft, Maria Cristina Gambetta, F.B.A. Lim, E. Farley, N. Stein, A. Haddad, D. Chauss, A.S. Mutlu, M.C. Wang, N.D. Young, E. Hildebrandt, H.H. Cheng, C.J. Knight, T.L.U. Burnham, K.A. Hovel, A.J. Beel, P.J. Mattei, R.D. Kornberg, W.C. Warren, G. Cary, J.L. Gomez-Skarmeta, V. Hinman, K. Lindblad-Toh, F. Di Palma, K. Maeshima, A.S. Multani, Sen Pathak, L. Nel-Themaat, R.R. Behringer, P. Kaur, R.H. Medema, B. van Steensel, E. de Wit, J.N. Onuchic, M. Di Pierro, E.L. Aiden & B.D. Rowland
In flies and mammals, dedicated DNA elements (insulators) recruit protein factors (insulator binding proteins, or IBPs) to shield promoters from regulatory elements.
In mammals, a single IBP called CCCTC-binding factor (CTCF) is known, whereas genetic and biochemical analyses in Drosophila identified a larger repertoire of IBPs.
Insulators are thought to fold chromosomes into conformations that affect regulatory element-promoter communication.
Comparing and contrasting observations in flies and mammals reveal that these species have different requirements for insulation, but that insulation is a conserved and critical gene regulation strategy.
The Insulator Protein CTCF Is Required for Correct Hox Gene Expression, but Not for Embryonic Development in Drosophila
In stark contrast to mammalian cells that die without CTCF, flies lacking zygotic CTCF survive to adults with homeotic defects.
The lack of major embryonic defects was assumed to be due to the maternal supply of CTCF protein, that could rescue the early development of CTCF mutants.
Here, we generated animals that completely lack both maternal and zygotic CTCF and found that, contrary to expectation, these mutants progress through embryogenesis and larval life. They develop to pharate adults, which fail to eclose from their pupal case.
These mutants show exacerbated homeotic defects compared to zygotic mutants, misexpressing the HOX gene Abdominal-B outside of its normal expression domain early in development.
This indicates that loss of Drosophila CTCF is not accompanied by widespread effects on gene expression, which may be due to redundant functions with other insulator proteins.
Rather, CTCF is required for correct HOX gene expression patterns and for the viability of adult Drosophila.
A critical perspective of the diverse roles of O-GlcNAc transferase in chromatin
O-linked β-N-Acetylglucosamine (O-GlcNAc) is a posttranslational modification that is catalyzed by O-GlcNAc transferase (Ogt) and found on a plethora of nuclear and cytosolic proteins in animals and plants.
Studies in different model organisms revealed that while O-GlcNAc is required for selected processes in Caenorhabditis elegans and Drosophila, it has evolved to become required for cell viability in mice.
Nevertheless, a principal cellular process that engages O-GlcNAcylation in all of these species is the regulation of gene transcription.
Here, we revisit several of the primary experimental observations that led to current models of how O-GlcNAcylation affects gene expression.
O-GlcNAcylation prevents aggregation of the Polycomb group repressor polyhomeotic
The glycosyltransferase Ogt adds O-linked N-Acetylglucosamine (O-GlcNAc) moieties to nuclear and cytosolic proteins.
Drosophila embryos lacking Ogt protein arrest development with a remarkably specific Polycomb phenotype, arising from the failure to repress Polycomb target genes.
The Polycomb protein Polyhomeotic (Ph), an Ogt substrate, forms large aggregates in the absence of O-GlcNAcylation both in vivo and in vitro.
O-GlcNAcylation of a serine/threonine (S/T) stretch in Ph is critical to prevent nonproductive aggregation of both Drosophila and human Ph via their C-terminal sterile alpha motif (SAM) domains in vitro.
Full Ph repressor activity in vivo requires both the SAM domain and O-GlcNAcylation of the S/T stretch.
Ph mutants lacking the S/T stretch reproduce the phenotype of ogt mutants, suggesting that the S/T stretch in Ph is the key Ogt substrate in Drosophila.
We propose that O-GlcNAcylation is needed for Ph to form functional, ordered assemblies via its SAM domain.
Structural basis for targeting the chromatin repressor Sfmbt to Polycomb response elements
Claudio Alfieri*, Maria Cristina Gambetta*, Raquel Matos, Sebastian Glatt, Peter Sehr, Sven Fraterman, Matthias Wilm, Jürg Müller** & Christoph W. Müller**
Polycomb group (PcG) protein complexes repress developmental regulator genes by modifying their chromatin.
Here, we report the crystal structure of the core of the Drosophila PcG protein complex Pleiohomeotic (Pho)repressive complex (PhoRC), which contains the Polycomb response element (PRE)-binding protein Pho and Sfmbt.
The spacer region of Pho, separated from the DNA-binding domain by a long flexible linker, forms a tight complex with the four malignant brain tumor (4MBT) domain of Sfmbt.
The highly conserved spacer region of the human Pho ortholog YY1 binds three of the four human 4MBT domain proteins in an analogous manner but with lower affinity.
Structure-guided mutations that disrupt the interaction between Pho and Sfmbt abolish formation of a ternary Sfmbt:Pho:DNA complex in vitro and repression of developmental regulator genes in Drosophila.
PRE tethering of Sfmbt by Pho is therefore essential for Polycomb repression in Drosophila.
The role of the histone H2A ubiquitinase Sce in Polycomb repression
Luis Gutiérrez, Katarzyna Oktaba, Johanna C. Scheuermann, Maria Cristina Gambetta, Nga Ly-Hartig & Jürg Müller
Development (Cambridge, England) 2011 · PMID / PMC / DOI
Essential role of the glycosyltransferase sxc/Ogt in polycomb repression
Maria Cristina Gambetta, Katarzyna Oktaba & Jürg Müller
Polycomb group proteins are conserved transcriptional repressors that control animal and plant development.
Here, we found that the Drosophila Polycomb group gene super sex combs (sxc) encodes Ogt, the highly conserved glycosyltransferase that catalyzes the addition of Nacetylglucosamine (GlcNAc) to proteins in animals and plants.
Genome-wide profiling in Drosophila revealed that GlcNAc-modified proteins are highly enriched at Polycomb response elements.
Among different Polycomb group proteins, Polyhomeotic is glycosylated by Sxc/Ogt in vivo.
sxc/Ogt–null mutants lacked O-linked GlcNAcylation and failed to maintain Polycomb transcriptional repression even though Polycomb group protein complexes were bound at their target sites.
Polycomb repression appears to be a critical function of Sxc/Ogt in Drosophila and may be mediated by the glycosylation of Polyhomeotic.
Prof. Maria Cristina Gambetta
Center for Integrative Genomics (CIG)
University of Lausanne
Unil-Sorge district
Genopode building
CH-1015 Lausanne
Switzerland
Are you interested in investigating basic mechanisms of gene regulation specificity?
We are looking for talented and enthusiastic team members at any level with a background in molecular biology, genetics, genomics or imaging. You are eager to engage in scientific discussions, capable of working in a team as well as independently.
Our lab is hosted at the Center for Integrative Genomics (CIG) at the University of Lausanne (UNIL), a vibrant, well-funded institute with a focus on functional genomics and equipped with modern core facilities. We are embedded in the broader Lausanne research environment that includes two universities (UNIL, EPFL), the Swiss Institute of Bioinformatics, Ludwig Center for Cancer Research, University Hospital, and a cluster of biotech companies flourishing in the larger lake Geneva area. We tightly network with other gene regulation labs at UNIL and EPFL, and collaborate with the on-site Bioinformatics Competence Center. There are regular possibilities to present and participate in local or international conferences and workshops. Hard and soft skill, and career development courses are offered on campus. We offer a nice working place in a multicultural, diversified and dynamic academic environment. PhD students will enroll in the UNIL Doctoral Program in Quantitative Biology.
Spontaneous applications are welcome anytime for Master’s/PhD/postdoc fellowships, while fully-funded positions are advertised below.
! We are currently hiring postdoctoral fellows !
To apply or request more information, email mariacristina.gambetta [at] unil.ch. Provide your CV, a brief description of your research experience, and why you think your research interests complement ours.
We encourage applications from candidates who will contribute to our team’s diversity.
