Chromatin, Metabolism and Cell Fate

ICO - German Trias i Pujol

Josep Carreras Leukaemia Research Institute
Edifici IMPPC
Can Ruti CampusCtra de Can Ruti
Camí de les Escoles s/n
08916 Badalona, Barcelona, Spain

Office 2-17 Lab 2-15 (second floor)

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Summary

Epigenetic information is written in chromatin. But how exactly do epigenetic mechanisms operate on the molecular level? How do chromatin alterations contribute to cell fate transitions? How does the environment influence these processes? And how does the metabolic state of a cell impact on its chromatin structure and its epigenetic memory?

These are questions we address in the lab. Studying stem cells and cancer we focus on molecular aspects of epigenetic regulation and on the question whether we can translate this knowledge into diagnostic and therapeutic tools for the management of diseases such as leukemia and myelodysplastic syndrome.

Research

In our scientific approach we combine biochemical techniques, genetic manipulation of cell cultures and ultrasequencing of enriched chromatin fractions to address mechanistic and functional aspects of epigenetics. As model system for cell differentiation we are using embryonic stem cells, myoblasts and hematopoietic cells. Key findings are validated in vivo. For the study of cancer we combine established cell lines, primary cultures and other patient samples.

Ongoing projects in the lab fall in one or several of three main themes:

  • The link between metabolism and epigenetic regulation.
  • The regulation and molecular function of histone variants.
  • Chromatin as drug target in myelodysplastic syndrome (MDS) and leukemia.

These are two examples of ongoing projects.

Chromatin modifiers as drug targets in MDS

Functional studies have been severely limited by the inefficient ex vivo growth of primary MDS cultures. In order to circumvent this problem we have characterized MDS patient-derived AML cell lines and identified lines that retain MDS characteristics such as a limited differentiation capacity. Our current efforts aim at the development of methods that will allow us to screen both FDA-approved drugs as well as novel compounds as first step of the drug development process. Furthermore genetic engineering allows us to test drug-gene interactions.

As several chromatin modifiers were found mutated in MDS patients we use genetic engineering in MDS-AML cell cultures to test the functional relevance of known and novel chromatin modifiers and their alterations. In collaboration with Katharina Götze (TU Munich) we will validate promising results in co-cultures of primary cells isolated from MDS patient bone marrow.

The regulation and function of the macroH2A histone variants

Histones form the protein core of the nucleosome, which is the modular building block of chromatin structure. MacroH2A is the only histone with a tripartite structure consisting of a N-terminal histone-fold, an intrinsically unstructured linker domain and a C-terminal macro domain. The incorporation of a macroH2A protein into the nucleosome places the macro domain in close proximity to the symmetrical axis and can be considered as the most extensive chromatin modification at the building block level.

From a large set of loss-of-function studies we could conclude that macroH2A is required for the establishment and maintenance of differentiated epigenomes. Want to know more? Please have a look at our recent reviews on macroH2A (Creppe, 2012; Posavec, 2013; Cantariño, 2013). In our current work we study context-dependent functions of macroH2A and the regulation of the locus-specific incorporation of macroH2A.

People

Selected publications

Buschbeck M, Hake SB

Variants of core histones and their roles in cell fate decisions, development and cancer.

Nat. Rev. Mol. Cell Biol. 1 Feb 2017, . Epub 1 Feb 2017
Histone variants endow chromatin with unique properties and show a specific genomic distribution that is regulated by specific deposition and removal machineries. These variants - in particular, H2A.Z, macroH2A and H3.3 - have important roles in early embryonic development, and they regulate the lineage commitment of stem cells, as well as the converse process of somatic cell reprogramming to pluripotency. Recent progress has also shed light on how mutations, transcriptional deregulation and changes in the deposition machineries of histone variants affect the process of tumorigenesis. These alterations promote or even drive cancer development through mechanisms that involve changes in epigenetic plasticity, genomic stability and senescence, and by activating and sustaining cancer-promoting gene expression programmes.
Més informació
Palau A, Mallo M, Palomo L, Rodríguez-Hernández I, Diesch J, Campos D, Granada I, Juncà J, Drexler HG, Solé F, Buschbeck M

Immunophenotypic, cytogenetic, and mutational characterization of cell lines derived from myelodysplastic syndrome patients after progression to acute myeloid leukemia.

Genes Chromosomes Cancer Mar 2017, 56 (3) 243-252. Epub 21 Nov 2016
Leukemia cell lines have been widely used in the hematology field to unravel mechanistic insights and to test new therapeutic strategies. Myelodysplastic syndromes (MDS) comprise a heterogeneous group of diseases that are characterized by ineffective hematopoiesis and frequent progress to acute myeloid leukemia (AML). A few cell lines have been established from MDS patients after progression to AML but their characterization is incomplete. Here we provide a detailed description of the immunophenotypic profile of the MDS-derived cell lines SKK-1, SKM-1, F-36P; and MOLM-13. Specifically, we analyzed a comprehensive panel of markers that are currently applied in the diagnostic routine for myeloid disorders. To provide high-resolution genetic data comprising copy number alterations and losses of heterozygosity we performed whole genome single nucleotide polymorphism-based arrays and included the cell line OHN-GM that harbors the frequent chromosome arm 5q deletion. Furthermore, we assessed the mutational status of 83 disease-relevant genes. Our results provide a resource to the MDS and AML field that allows researchers to choose the best-matching cell line for their functional studies. © 2016 Wiley Periodicals, Inc.
Més informació
Diesch J, Zwick A, Garz AK, Palau A, Buschbeck M, Götze KS

A clinical-molecular update on azanucleoside-based therapy for the treatment of hematologic cancers.

Clin Epigenetics 2016, 8 71. Epub 21 Jun 2016
The azanucleosides azacitidine and decitabine are currently used for the treatment of acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) in patients not only eligible for intensive chemotherapy but are also being explored in other hematologic and solid cancers. Based on their capacity to interfere with the DNA methylation machinery, these drugs are also referred to as hypomethylating agents (HMAs). As DNA methylation contributes to epigenetic regulation, azanucleosides are further considered to be among the first true "epigenetic drugs" that have reached clinical application. However, intriguing new evidence suggests that DNA hypomethylation is not the only mechanism of action for these drugs. This review summarizes the experience from more than 10 years of clinical practice with azanucleosides and discusses their molecular actions, including several not related to DNA methylation. A particular focus is placed on possible causes of primary and acquired resistances to azanucleoside treatment. We highlight current limitations for the success and durability of azanucleoside-based therapy and illustrate that a better understanding of the molecular determinants of drug response holds great potential to overcome resistance.
Més informació
Cantarino N, Musulen E, Valero V, Peinado MA, Perucho M, Moreno V, Forcales SV, Douet J, Buschbeck M

Downregulation of the Deiminase PADI2 is an Early Event in Colorectal Carcinogenesis and Indicates Poor Prognosis.

Mol. Cancer Res. 8 Jun 2016, . Epub 8 Jun 2016
Peptidyl arginine deiminases (PADIs) are a family of enzymes that catalyze the poorly understood post-translational modification converting arginine residues into citrullines. In this study, the role of PADIs in the pathogenesis of colorectal cancer was investigated. Specifically, RNA expression was analyzed and its association with survival in a cohort of 98 colorectal cancer patient specimens with matched adjacent mucosa and 50 controls from donors without cancer. Key results were validated in an independent collection of tumors with matched adjacent mucosa and by mining of a publicly available expression data set. Protein expression was analyzed by immunoblotting for cell lines or immunohistochemistry (IHC) for patient specimens that further included 24 cases of adenocarcinoma with adjacent dysplasia and 11 cases of active ulcerative colitis. The data indicate that PADI2 is the dominantly expressed PADI enzyme in colon mucosa and is up-regulated during differentiation. PADI2 expression is low or absent in colorectal cancer. Frequently this occurs already at the stage of low-grade dysplasia. Mucosal PADI2 expression is also low in ulcerative colitis. The expression level of PADI2 in tumor and adjacent mucosa correlates with differential survival: low levels associate with poor prognosis.
Més informació
Creppe C, Palau A, Malinverni R, Valero V, Buschbeck M

A Cbx8-Containing Polycomb Complex Facilitates the Transition to Gene Activation during ES Cell Differentiation.

PLoS Genet. Des 2014, 10 (12) e1004851. Epub 11 Des 2014
Polycomb proteins play an essential role in maintaining the repression of developmental genes in self-renewing embryonic stem cells. The exact mechanism allowing the derepression of polycomb target genes during cell differentiation remains unclear. Our project aimed to identify Cbx8 binding sites in differentiating mouse embryonic stem cells. Therefore, we used a genome-wide chromatin immunoprecipitation of endogenous Cbx8 coupled to direct massive parallel sequencing (ChIP-Seq). Our analysis identified 171 high confidence peaks. By crossing our data with previously published microarray analysis, we show that several differentiation genes transiently recruit Cbx8 during their early activation. Depletion of Cbx8 partially impairs the transcriptional activation of these genes. Both interaction analysis, as well as chromatin immunoprecipitation experiments support the idea that activating Cbx8 acts in the context of an intact PRC1 complex. Prolonged gene activation results in eviction of PRC1 despite persisting H3K27me3 and H2A ubiquitination. The composition of PRC1 is highly modular and changes when embryonic stem cells commit to differentiation. We further demonstrate that the exchange of Cbx7 for Cbx8 is required for the effective activation of differentiation genes. Taken together, our results establish a function for a Cbx8-containing complex in facilitating the transition from a Polycomb-repressed chromatin state to an active state. As this affects several key regulatory differentiation genes this mechanism is likely to contribute to the robust execution of differentiation programs.
Més informació
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Current projects

LAS VARIANTES DE HISTONAS MACROH2A VINCULAN LA ARQUITECTURA DEL GENOMA AL METABOLISMO

Responsable:Marcus Buschbeck
Codi:BFU2015-66559-P
Financiadors:
Data d'inici:01/01/2016
Data de finalització:31/12/2018

Dissecting the Role of Polycomb Complexes in the Pathogenesis of Myelodysplastic Syndromes (MDS) and the Evolution to Acute Myeloid Leukemia (DJCLS R 14/16)

Responsable:Marcus Buschbeck
Codi:DJCLS R 14/16
Financiadors:
Data d'inici:01/08/2015
Data de finalització:31/07/2017

How does the histone variant macroH2A regulate muscle metabolism in health and disease?

Responsable:Marcus Buschbeck
Codi:18738
Financiadors:
Data d'inici:16/02/2015
Data de finalització:26/10/2017

Chromatin-metabolism interactions as targets for healthy living (ChroMe)

Responsable:Marcus Buschbeck
Codi:ChroMe
Financiadors:
Data d'inici:01/03/2016
Data de finalització:29/02/2020

Previous projects

Drug repositioning as a fast and cost effective approach to personalized therapies. A pilot study on myelodysplastic syndrome and acute myeloid leukaemia

Responsable:Marcus Buschbeck
Codi:AECC_IMPPC_MB_2015
Financiadors:
Data d'inici:28/02/2014
Data de finalització:31/03/2016

Plan Nacional - Epigenetic Regulators of Stem Cell Function

Responsable:Marcus Buschbeck
Codi:SAF2012-39749
Financiadors:
Data d'inici:01/01/2013
Data de finalització:31/12/2015