Our Current Research

The Chromatin Biology Laboratory was established by Dr Vaquero in 2008, with the major aim to understand the mechanisms of stress response and their impact in cancer and aging. Specifically, the group focus their efforts in defining the contribution of sirtuins to this response in the maintenance of genome stability, epigenetics and metabolic homeostasis. To fulfil this main objective, the work of the group covers a range of research from basic aspects of sirtuin biology to the study of their contribution to the development of human pathologies, such as cancer and aging.

Since its foundation, the lab has described several novel Sirtuins-dependent mechanisms in genome stability protection and epigenetics (Mol Cell 2011, Genes &Dev 2013, Epigenetics 2017, Cell Rep 2017, Nat comm 2018, Sci Adv 2020, Aging 2021), has developed fruitful collaborations in this context (Dev. Cell 2013; EMBOJ 2016; J Hepatol 2017;Sci Reports 2017; Cell Death dis 2018; NAR 2017,2019,2020; PNAS 2017,2020) and has participated in the main discussions of Sirtuins field (Genes&Dev 2009, Science 2010, Cancer Cell 2012, Oncogene 2014, FEBS J 2015, Proteomics 2017; Mol Cell Oncol 2018; Mol. Reprod. Dev 2020).

Sirtuins are NAD+
Fig1. Sirtuins are NAD+- dependent enzymes that play a key role in the response to different types of stress including Metabolic, oxidative and genotoxic stress. These stress conditions are a threat in both Prokaryotes and Eukaryotes which have led to the development of a complex set of mechanisms to efficiently adapt to these conditions. In Eukaryotes the response to these conditions takes place at both genomic level and metabolic level. Sirtuins coordinate the response at both levels.

The main strands of the group’s work are:

Sirtuin enzymatic duality

Although the majority of sirtuins are NAD+ - dependent deacetylases, some family members such as SIRT6, also harbor a second enzymatic activity, a mono-ADP-ribosyltransferase (mADPRT) activity. This functional duality is intriguing and is one focus of the group’s work. Studies from our group identified SIRT7 as a dual Sirtuin, and found that, contrary to what was previously assumed, the ADPRT activity in both SIRT6 and SIRT7 lies in a second active site located in a different protein domain away from the primary deacetylation site (Simonet el al., 2020). The active site is based in a ELHGN motif and is conserved through evolution in the whole SIRT6/SIRT7 lineage. In this context, residues E and N of the motif are key players in the mADPRT activity (labeled in blue in the video)(Fig2).

The mADPRT activity
Fig2. The mADPRT activity of dual Sirtuins SIRT6 and SIRT7 is catalyzed by a second active site in protein domain physically separated from the primary deacetylation site (in red). In the cartoon the structure of the SIR2 conserved domain of SIRT7 is shown. Top left, the mADPRT domain forms a cavity containing the conserved motif ELHGN. Top right, E and N are key in the mADPRT catalytic activity. SIRT7 N189 mutated to Ala abrogated specifically the mADPRT activity but not the deacetylase activity (HDAC) tested in in vitro assays with a H3K18 acetylated peptide. Mutation of H187Y had the opposite effect (From Simonet et al., Sci Adv. 2020). Bottom, video of the perspective around the SIRT7 catalytic domain ending in the mADPRT domain. Residues E and N are exposed in the cavity and marked in blue.

The work of the group aims to define the nature of this duality through:

  • Defining the functional interplay between both activities
  • Molecular and functional characterization of the poorly understood mADPRT activity
  • Identification of novel targets of these activities
  • The contribution of both enzymatic activities to the role of SIRT6/SIRT7 in cancer
Sirtuin-dependent mechanisms of genomic stability

The protection of genome integrity is a major priority for the cells. It involves a wide range of factors, mechanisms and pathways that are intimately coupled with the basic regulatory circuitry that controls cell life and survival. Among the most relevant pathways are DNA repair and DNA damage signaling, cell cycle and the regulation of the dynamics of global chromatin structure.

Our group aims to define the safeguard mechanism of the response to these stress response conditions through the following aspects:

  • Maintenance of constitutive Heterochromatin (CH) structure
  • DNA damage Signaling and repair
  • Cell cycle checkpoint control
  • Epigenetic regulation of Meiosis and Gametogenesis

In the first case, our studies center in Pericentric Heterochromatin (PCH) where we have done important discoveries to understand both the specific contribution of the heterochromatin structural proteins HP1 isoforms HP1a,b,g in PCH structure, and the involvement of Sirtuins in PCH integrity. In this sense our studies showed that HP1a and g are functionally associated to Suv39h1, the main H3K9me3 histone methyltransferase and a keystone of genome stability (Raurell-Vila et al., Epigenetics 2017). Interestingly, while HP1a has a specific role in the maintenance of PCH epigenetic and structural compartmentalization, HP1b is functional associated to H4K20me3 (Bosch-Presegue et al., Cell Reports 2017) (Fig3).

The mADPRT activity
Fig3. HP1 isoforms play distinctive roles in pericentric heterochromatin (PCH). Top, HP1α defines, together with CTCF, a compartmentalized structure in PCH foci that restricts spreading epigenetic marks H4K20me3 and H3K27me3. Bottom, loss of HP1α impairs CTCF localization and promotes spreading of all three heterochromatin-associated factors in PCH (From Bosch-Presegué et al., Cell Rep. 2017).

Regarding the role of Sirtuins in this context, we previously defined a functional interplay between Suv39h1 and SIRT1 in PCH regions (Vaquero et al., Nature 2007). Our studies discovered a new mechanism of control of PCH integrity upon stress, by the control of Suv39h1 stability and turnover in PCH regions (Bosch-Presegue et al 2010). This seems to be an important adaptative mechanism that ensures a proper epigenetic control of PCH structure (Fig4).

The mADPRT activity
Fig4. SIRT1 binds to the histone methyltransferase SUV39H1 and regulates its stability in pericentric constitutive heterochromatin (PCH). Upon stress, SIRT1 upregulation promotes the stabilization of SUV39H1 and a subsequent increased in turnover of SUV39H1 in PCH, which promotes genome stability (From Bosch-Presegué et al., Mol. Cell 2011).

In the case of cell cycle regulation, our studies identified a role for SIRT2 in the control of G2/M checkpoint through regulation of the H4K16ac (Vaquero et al., Genes&Dev 2006, Serrano et al., Genes&Dev 2013), an epigenetic mark directly linked to genome integrity as, in contrast to the rest of acetylation marks, it directly regulates the folding of the chromatin fiber. SIRT2 not only is responsible for the removal of H4K16ac before the start of mitosis, but also regulates the deposition of its antagonist mark, H4K20me1, through a functional interplay with the main H4K20me1 HMT (Serrano et al., Genes&Dev 2013). This interplay alters the global levels of H4K20me1 and its associated marks H4K20me2,3, which has key consequences not only in mitosis progression but also in DNA replication and DNA repair (Fig5).

The mADPRT activity
Fig5. SIRT2 regulates the key epigenetic mark H4K20me1 in G2/M transition through deacetylation of H4K16ac and a functional interplay with the H4K20me1-specific histone methyltransferase PR-Set7. Top, metaphase chromosomes of mouse embryonic fibroblasts derived from mice Wt or Sirt2-/- show hyperacetylation of H4K16 and hypomethylation of H4K20me1 in Sirt2-/- cells. Bottom left, DNA FISH of centromeric or telomeric regions show a wide range of chromosomal abnormalities in Sirt2-/- chromosomes. Duplication (red) or depletion (green) of the regions are indicated. Bottom right, Model proposed for the interplay between SIRT2 and PR-Set7. In G2/M SIRT2 deacetylates H4K16ac, allowing H4K20me1 methylation by PR-Set7, which allows proper compaction of chromosomes and completion of mitosis. This has an impact in the following cell cycle at the level of DNA replication and DNA repair given the key role of H4K20me2 and H4K20me3 in these mechanisms (From Serrano et al., Genes Dev 2013).
Role of Sirtuins in B-cell differentiation and Leukemia

Sirtuins play an important role in the hematopoietic system, although is poorly understood. Thus, Sirtuins are involved in the maintenance of hematopoietic stem cells, cell differentiation and immune response, and have been associated with the development of some types of leukemia.

Our current work aims to understand Sirtuin contribution to hematopoiesis and leukemia through the development of two main subjects:

  • The role of Sirtuins in B-cell differentiation
  • Their implication in Leukemia

Our evidence suggest that some Sirtuins have a central role in specific phases of B-cell differentiation, probably though modulation of stress-dependent mechanisms. By a multidisciplinary approach from in vitro molecular studies to in vivo mouse models we aim to define the contribution of these Sirtuins to the differentiation process, the impact of sirtuin-dependent stress response in the maintenance of genome stability in this process, and its impact in immune response in vivo.

We are also particularly interested in understanding the functional implication of sirtuins in cancer, and in particular in the context of hematopoietic pathologies like leukemia and lymphomas. Our main efforts are currently focused in two types of Leukemia, pediatric B-ALL and in AML. The development of our lines of work should provide key evidence to understand the molecular basis of these pathologies, and to identify novel candidates that can be potentially used as markers of prognosis or novel therapeutical targets.

Sirtuins, CR and aging

The implication of Sirtuins in aging seems to have three major coordinates, the control of stress response, the maintenance of genome stability and the regulation of metabolism. In this sense, the beneficial effects of diet interventions such as a calorie restriction(CR) have been shown to alleviate aging phenotype and increase lifespan. Given their key role in metabolic homeostasis Mammalian Sirtuins are involved in the response to these stress conditions, although whether they participate in lifespan is an issue still under debate. In this sense, loss of several of these Sirtuins have a direct impact in aging. For instance, the knockdown of SIRT6 and SIRT7 mice induce a hyperaccelerated aging phenotype (Kawahara et al., Cell 2006, Vazquez et al., EMBO J 2016) which involves a wide range of defects that range from development to genome stability and metabolism.

Our work found that SIRT7 plays an important role in the response to glucose starvation and CR through a functional interplay with H2A variant macroH2A through a mechanism that involves at different levels both enzymatic activities of SIRT7 (Simonet et al., Sci Adv 2020). The regulatory axis SIRT7/macroH2A regulates a set of genes in response to glucose starvation and CR involved in a wide range of functions, from growth factors, second messenger signaling-related factors, epigenetic regulators, and transcription factors, among others. Importantly, reflecting the close link between CR and aging, we demonstrated the SIRT7-dependent regulation of these genes upon aging. Our current studies aim to understand the involvement of Sirtuin function (SIRT6/SIRT7) in the beneficial effects of nutrient restriction on aging development. We also aim to define the specific contribution of mAPDRT activity of SIRT7 to this response.

Development of new methodologies to measure Sirtuin activity

The importance of Sirtuins in Biomedical studies is reflected by the considerable number of publications focused in Sirtuins in a wide range of functional contexts, from basic science to clinical studies in a wide range of human pathologies such as cancer or aging. For this purpose, pharmaceutical companies have engaged in two major approaches in Sirtuin-related lines of work, including 1) The discovery of novel drugs to modulate the activity of specific Sirtuin family members; and 2) The development of methods to monitor Sirtuin enzymatic activity (deacetylase activity) in samples derived from cells or tissues.

One of the major handicaps in the study of Sirtuins is the lack of a reliable detection method that can provide a full, direct and specific measurement of the activity. To this date, the commercially available kits of sirtuin activity are mostly designed to monitor deacetylation and are based on the use of a specific acetylated peptide fused to a fluorophore that can switch the emission wavelength upon deacetylation. In our group we are currently developing a new method, currently under optimization, to assess both Sirtuin deacetylation and/or mADPRTion.

Josep Carreras Leukaemia Research Institute Can Ruti Campus, Ctra de Can Ruti, Camí de les Escoles s/n
08916 Badalona, Barcelona

Office (+34)-935572800 ext 4240
Lab (+34)-935572800 ext 4241

©2024 - Chromatin Biology Lab