Life Sciences Research for Lifelong Health

Claudia Ribeiro de Almeida

Research Summary

We study the molecular mechanisms underlying diversification of antibody genes by B cells, which are a key part of the immune system. Antibodies are encoded in the genome by immunoglobulin heavy (IgH) and light (IgL) chain loci, which go through processes of DNA deletion-recombination and DNA mutation at different stages of B cell development. These changes ensure that B cells produce a tailored and highly specific antibody response against a wide range of pathogens. Our aim is to understand the basic principles of immunoglobulin gene diversification to gain insight into how B cells can effectively fight infections, how these responses change through our lives and the role this plays in age-related immune dysfunction.


Our research focuses on characterising the role of RNA and RNA binding proteins (RBPs) in controlling V(D)J recombination and class switch recombination (CSR) at immunoglobulin loci. We are interested in the study of DEAD/DEAH-box RNA helicases, a large family of RBPs that utilize ATP hydrolysis to remodel RNA structure or RNA-protein interactions. One example is the DEAD-box RNA helicase 1 (DDX1), which is required for the mechanism of IgH CSR through modulation of RNA secondary structure.

We use a number of cellular and molecular biology tools together with mouse genetics, ex vivo cellular systems and in vitro biochemical assays, to investigate the function of RBPs in B cells undergoing immunoglobulin gene rearrangements. We are also engaged in identifying novel RBPs implicated in these processes using genomic and proteomic approaches.

Latest Publications

RNA Helicase DDX1 Converts RNA G-Quadruplex Structures into R-Loops to Promote IgH Class Switch Recombination.
Ribeiro de Almeida C, Dhir S, Dhir A, Moghaddam AE, Sattentau Q, Meinhart A, Proudfoot NJ

Class switch recombination (CSR) at the immunoglobulin heavy-chain (IgH) locus is associated with the formation of R-loop structures over switch (S) regions. While these often occur co-transcriptionally between nascent RNA and template DNA, we now show that they also form as part of a post-transcriptional mechanism targeting AID to IgH S-regions. This depends on the RNA helicase DDX1 that is also required for CSR in vivo. DDX1 binds to G-quadruplex (G4) structures present in intronic switch transcripts and converts them into S-region R-loops. This in turn targets the cytidine deaminase enzyme AID to S-regions so promoting CSR. Notably R-loop levels over S-regions are diminished by chemical stabilization of G4 RNA or by the expression of a DDX1 ATPase-deficient mutant that acts as a dominant-negative protein to reduce CSR efficiency. In effect, we provide evidence for how S-region transcripts interconvert between G4 and R-loop structures to promote CSR in the IgH locus.

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Molecular cell, 70, 1097-4164, 650-662.e8, 2018

PMID: 29731414

Dynamic Control of Long-Range Genomic Interactions at the Immunoglobulin κ Light-Chain Locus.
Ribeiro de Almeida C, Hendriks RW, Stadhouders R

The Igκ locus, which is spread over 3Mb of genomic DNA and contains >100 variable (V) genes, serves as an important model system to study long-range chromatin interactions. Here, we will discuss how in developing B cells in the bone marrow the accessibility of individual Vκ segments is controlled by many lineage-specific and ubiquitously expressed transcription factors that act on various cis-regulatory elements, including promoters, enhancers, and insulators. This dynamic control furthermore involves changes in subnuclear localization, histone modification, DNA demethylation, and three-dimensional locus compaction. In pro-B cells, the Igκ locus adopts a poised conformation as full contraction has been achieved and many key transcription factors already occupy the locus. Subsequently, the combined activation of pre-B cell antigen receptor signaling pathways and attenuation of IL-7R signaling in small resting pre-B cells dramatically modifies the transcription factor landscape, supporting the induction of monoallelic Igκ gene rearrangements. Hereby, the intronic and 3' Igκ enhancer elements coordinately focus their activities in the Vκ region toward frequently used Vκ genes. Recent work has drawn attention to the intriguing role of the CTCF-associated regulatory elements Cer and Sis, which are located in the Vκ-Jκ intervening region and control Igκ locus contraction and Vκ repertoire diversity. This involves CTCF-mediated locus insulation, restricting enhancer activity to the Vκ region and suppressing the preferential recombination to proximal Vκ genes. A picture emerges in which the dynamic control of long-range genomic interactions ensures correct timing of Igκ locus recombination and provides appropriate opportunities for individual Vκ gene segments to engage in Vκ-Jκ rearrangement.

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Advances in immunology, 128, 1557-8445, 183-271, 2015

PMID: 26477368

Pre-B cell receptor signaling induces immunoglobulin κ locus accessibility by functional redistribution of enhancer-mediated chromatin interactions.
Stadhouders R, de Bruijn MJ, Rother MB, Yuvaraj S, Ribeiro de Almeida C, Kolovos P, Van Zelm MC, van Ijcken W, Grosveld F, Soler E, Hendriks RW

During B cell development, the precursor B cell receptor (pre-BCR) checkpoint is thought to increase immunoglobulin κ light chain (Igκ) locus accessibility to the V(D)J recombinase. Accordingly, pre-B cells lacking the pre-BCR signaling molecules Btk or Slp65 showed reduced germline V(κ) transcription. To investigate whether pre-BCR signaling modulates V(κ) accessibility through enhancer-mediated Igκ locus topology, we performed chromosome conformation capture and sequencing analyses. These revealed that already in pro-B cells the κ enhancers robustly interact with the ∼3.2 Mb V(κ) region and its flanking sequences. Analyses in wild-type, Btk, and Slp65 single- and double-deficient pre-B cells demonstrated that pre-BCR signaling reduces interactions of both enhancers with Igκ locus flanking sequences and increases interactions of the 3'κ enhancer with V(κ) genes. Remarkably, pre-BCR signaling does not significantly affect interactions between the intronic enhancer and V(κ) genes, which are already robust in pro-B cells. Both enhancers interact most frequently with highly used V(κ) genes, which are often marked by transcription factor E2a. We conclude that the κ enhancers interact with the V(κ) region already in pro-B cells and that pre-BCR signaling induces accessibility through a functional redistribution of long-range chromatin interactions within the V(κ) region, whereby the two enhancers play distinct roles.

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PLoS biology, 12, 1545-7885, e1001791, 2014

PMID: 24558349

 

Group Members

Latest Publications

RNA Helicase DDX1 Converts RNA G-Quadruplex Structures into R-Loops to Promote IgH Class Switch Recombination.

Ribeiro de Almeida C, Dhir S, Dhir A

Molecular cell
70 1097-4164:650-662.e8 (2018)

PMID: 29731414

Dynamic Control of Long-Range Genomic Interactions at the Immunoglobulin κ Light-Chain Locus.

Ribeiro de Almeida C, Hendriks RW, Stadhouders R

Advances in immunology
128 1557-8445:183-271 (2015)

PMID: 26477368

The DNA-binding factor Ctcf critically controls gene expression in macrophages.

Nikolic T, Movita D, Lambers ME

Cellular & molecular immunology
11 2042-0226:58-70 (2014)

PMID: 24013844

Allelic exclusion of the immunoglobulin heavy chain locus is independent of its nuclear localization in mature B cells.

Holwerda SJ, van de Werken HJ, Ribeiro de Almeida C

Nucleic acids research
41 1362-4962:6905-16 (2013)

PMID: 23748562

DNA-binding factor CTCF and long-range gene interactions in V(D)J recombination and oncogene activation.

Ribeiro de Almeida C, Stadhouders R, Thongjuea S

Blood
119 1528-0020:6209-18 (2012)

PMID: 22538856

Gene expression profiling in mice with enforced Gata3 expression reveals putative targets of Gata3 in double positive thymocytes.

van Hamburg JP, de Bruijn MJ, Ribeiro de Almeida C

Molecular immunology
46 1872-9142:3251-60 (2009)

PMID: 19729201

Critical role for the transcription regulator CCCTC-binding factor in the control of Th2 cytokine expression.

Ribeiro de Almeida C, Heath H, Krpic S

Journal of immunology (Baltimore, Md. : 1950)
182 1550-6606:999-1010 (2009)

PMID: 19124743

CTCF regulates cell cycle progression of alphabeta T cells in the thymus.

Heath H, Ribeiro de Almeida C, Sleutels F

The EMBO journal
27 1460-2075:2839-50 (2008)

PMID: 18923423

Enforced expression of GATA3 allows differentiation of IL-17-producing cells, but constrains Th17-mediated pathology.

van Hamburg JP, de Bruijn MJ, Ribeiro de Almeida C

European journal of immunology
38 0014-2980:2573-86 (2008)

PMID: 18792410