Alvaro Rada-Iglesias - JRG VII

Developmental Genomics: Transcriptional regulation in development and disease

The ultimate goal of my laboratory is to decipher the genetic and epigenetic mechanisms controlling the deployment of gene expression programs during vertebrate embryogenesis. More specifically, by functionally and mechanistically characterizing enhancer regulatory landscapes, my laboratory aims at (i) uncovering transcriptional regulatory principles orchestrating mammalian embryogenesis and (ii) elucidating the genetic and epigenetic basis of human congenital diseases. 


The establishment of cell type-specific gene expression programs largely depends on a group of distal cis-regulatory elements broadly referred to as enhancers. By serving as binding platforms for transcription factors, enhancers can integrate signaling and epigenetic cues in order to exquisitely control the expression of their target genes. Furthermore, accumulating evidences suggest that changes in the regulatory activity or topology of enhancer landscapes due to genetic or structural variation, respectively, can represent a major cause of human disease.

Epigenetic state of enhancers as a major determinant of developmental competence

Developmental competence defines a cellular state in which progenitor cells are able to respond to inductive signals in order to progress towards a specific cellular lineage. Our recent findings using ESC-based differentiation models suggest that the competence of pluripotent cells to differentiate towards certain cellular lineages is determined by the poised state of a key set of developmental enhancers. Thus, we hypothesize that epigenetic state of enhancers is a major determinant of pluripotency and developmental competence.

We previously identified poised enhancers in human embryonic stem cells (ESC) as cis-regulatory sequences displaying a unique chromatin signature, which includes H3K27me3, a histone modification mediated by polycomb repressive complex 2 (PRC2). Most recently, we demonstrated that poised enhancers are necessary for the induction of major anterior neural genes during ESC differentiation. Interestingly, poised enhancers already established physical interactions with their target genes in ESC in a PRC2 dependent manner. Loss of PRC2 led to neither the activation of poised enhancers nor the induction of their target genes in ESC. In contrast, loss of PRC2 compromised the induction of major anterior neural genes representing poised enhancer targets. Hence, our work illuminates a novel function for PRC2, which we propose provides major anterior neural loci with a permissive regulatory topology that renders pluripotent cells competent for neural induction (Figure 1). Our future efforts will aim at elucidating two major questions: (i) Are poised enhancers functionally relevant in vivo?; (ii) Does PRC2 act as topological facilitator of anterior neural induction?

Structural and genetic disruption of enhancer regulatory landscape as a pervasive etiological mechanism in human congenital disease

Structural variants (SV) (i.e. deletions, duplications, inversions, translocations) can cause congenital abnormalities by disrupting the 3D organization of enhancer regulatory landscapes. However, elucidating the pathomechanism of human structural variation in vivo is complicated by the limited access to appropriate patient material and/or differences in gene dosage sensitivity between mice and humans. These limitations are well illustrated by Branchio-Oculo-Facial Syndrome (BOFS), a rare congenital disorder caused by heterozygous mutations within TFAP2A, a neural crest (NC) master regulator for which humans, but not mice, are haploinsufficient. We recently described a unique BOFS patient carrying a de novo heterozygous inversion in which one of the breakpoints is located downstream of TFAP2A, within a large Topologically Associating Domain (TAD) containing enhancers essential for the expression of this gene in human NC cells (hNCC). Using patient-specific hiPSC and various genomic approaches, we systematically evaluated the molecular consequences of the inversion and conclusively demonstrated that it disconnects the inverted TFAP2A allele from its cognate enhancers, leading to TFAP2A monoallelic expression in hNCC. In turn, this results in TFAP2A haploinsufficiency due to reduced TFAP2A binding to enhancers controlling the expression of genes involved in craniofacial morphogenesis and NCC migration. 

The medical relevance of enhancers is further supported by the fact that most of the genetic variants associated with complex human disorders overlap enhancers. Based on this general observation, we have recently developed an in silico toolkit to etiologically link human diseases and cell types and to identify potential disease-causative genetic variants overlapping relevant cis-regulatory elements. Using this in silico toolkit, we identified non-coding genetic variants overlapping enhancers active in hNCC and associated with orofacial clefts. Subsequently and focusing on a few orofacial cleft-risk loci, we used functional genomic technologies (4C-seq, ChIP-seq) in hNCC and frontonasal prominences isolated from chicken embryos to predict the genes that could be regulated by the enhancers of interest. Using this approach, we predicted that a hNCC enhancer close to the FAM49A gene controls the expression of a couple of distal (>2 Mb) genes (MYCN and DDX1). Subsequently, we used loss of function approaches in vitro (hNCC) and in vivo (chicken embryos) to demonstrate that MYCN and DDX1 play important roles during craniofacial development and, thus, are likely to be implicated in orofacial cleft etiology. 


Our recent work illustrates how a detailed and mechanistic understanding of enhancer function and enhancer regulatory landscapes can be used to uncover novel pathomechanisms implicated in human congenital disorders. In the future, this can lead to improved diagnosis, management and even treatment of some of these pathologies. 

Selected publications

Rada-Iglesias A, Bajpai R, Swigut T, Brugmann SA, Flynn RA, Wysocka J. (2011). A unique chromatin signature uncovers early developmental enhancers in humans. Nature 470(7333), 279-83. 

Respuela P, Nikolic P, Tan M, Frommolt P, Zhao Y, Wysocka J and Rada-Iglesias A. (2016). Foxd3 promotes exit from naïve pluripotency through enhancer decommissioning and inhibits germline specification. Cell Stem Cell. 18(1), 118-133. 

Cruz-Molina S, Respuela P, Tebartz C, Kolovos P, Nikolic M, Fueyo R, van Ijcken W.F.J, Grosveld F, Frommolt P, Bazzi H, Rada-Iglesias A. (2017). PRC2 facilitates the regulatory topology required for poised enhancer function during pluripotent stem cell differentiation. Cell Stem Cell 20(5), 689-705.

Nikolić M, Papantonis A, Rada-Iglesias A. (2017). GARLIC: A Bioinformatic Toolkit for Etiologically Connecting Diseases and Cell Type-Specific Regulatory Maps. Hum Mol Genet 26(4), 742-752. 

Laugsch M, Bartusel M, Alirzayeva H, Karaolidou A, Rehimi R, Crispatzu G, Nikolic M, Bleckwehl T, Kolovos P, van Ijcken W.F.J, Saric T, Kohler K, Frommolt P, Lachlan K, Baptista J, Rada-Iglesias A. (2018). Disruption of the TFAP2A regulatory domain causes Branchio-Oculo-Facial Syndrome (BOFS) and illuminates pathomechanisms for other human neurocristopathies. Under revision, Cell Stem Cell.

Dr. Alvaro Rada-Iglesias

Center for Molecular Medicine Cologne

Dr. Alvaro Rada-Iglesias

Principal Investigator - CMMC JRG VII

Publications - Alvaro Rada-Iglesias

Link to PubMed

Group Members

Rizwan Rehimi (PostDoc)
Magdalena Laugsch (PostDoc)
Rodrigo Osorno (PostDoc)
Giuliano Crispatzu (postdoc)
Michaela Bartusel (doctoral student)
Tomas Pachano (doctoral student)
Tore Bleckwehl (doctoral student)
Jan Appel (Lab Manager).

Figure 1

Current model describing poised enhancer function during the establishment of anterior neural identity.

Figure 2

Etiological mechanism in a unique BOFS patient in which an inversion leads the disconnection between TFAP2A and its cognate enhancers.