Biology subjects

Batista, C. R.

Publications and source records attributed to Batista, C. R..

3 recordsLinked to original sources

bioRxiv2024

Negative regulation of Activation-Induced Cytidine Deaminase gene transcription in developing B cells by a PU.1-interacting intronic region

Activation-induced cytidine deaminase (AID, encoded by Aicda) plays a key role in somatic hypermutation and class switch recombination in germinal center B cells. However, off-target effects of AID are implicated in human leukemia and lymphoma. A mouse model of precursor B cell acute lymphoblastic leukemia driven by deletion of the related transcription factors PU.1 and Spi-B revealed C->T transition mutations compatible with being induced by AID. Therefore, we hypothesized that PU.1 negatively regulates Aicda during B cell development. Aicda mRNA transcript levels were increased in leukemia cells and preleukemic bone marrow pre-B cells lacking PU.1 and/or Spi-B, relative to wild type cells. Using chromatin immunoprecipitation, PU.1 was found to interact with a negative regulatory region (R2-1) within the first intron of Aicda. CRISPR-Cas9-induced mutagenesis of R2-1 in cultured pre-B cells resulted in upregulation of Aicda in response to lipopolysaccharide stimulation. Mutation of the PU.1 interaction site and neighboring sequences resulted in reduced repressive ability of R2-1 in transient transfection analysis followed by luciferase assays. These results show that a PU.1-interacting intronic region negatively regulates Aicda transcription in developing B cells.

immunology↗

bioRxiv2022

Spi-C and PU.1 counterregulate Rag1 and Igκ transcription to effect immunoglobulin kappa recombination in small pre-B cells

B cell development requires the ordered rearrangement of Ig genes encoding H and L chain proteins that assemble into BCRs or Abs capable of recognizing specific Ags. Ig{kappa} rearrangement is promoted by chromatin accessibility and by relative abundance of RAG1/2 proteins. Expression of the E26-transformation-specific (ETS) transcription factor Spi-C is activated in response to double-stranded DNA breaks (DSBs) in small pre-B cells to negatively regulate pre-BCR signaling and Ig{kappa} rearrangement. However, it is not clear if Spi-C regulates Ig{kappa} rearrangement through transcription or by controlling RAG expression. In this study, we investigated the mechanism of Spi-C negative regulation of Ig{kappa} light chain rearrangement. Using an inducible expression system in a pre-B cell line, we found that Spi-C negatively regulated Ig{kappa} rearrangement, Ig{kappa} transcript levels, and Rag1 transcript levels. We found that Ig{kappa} and Rag1 transcript levels were increased in small pre-B cells from Spic-/- mice. In contrast, Ig{kappa} and Rag1 transcript levels were activated by PU.1 and were decreased in small pre-B cells from PU.1-deficient mice. Using chromatin immunoprecipitation analysis, we identified an interaction site for PU.1 and Spi-C located in the Rag1 promoter region. These results suggest that Spi-C and PU.1 counterregulate Ig{kappa} transcription and Rag1 transcription to effect Ig{kappa} recombination in small pre-B cells.

immunology↗

bioRxiv2018

Deletion of genes encoding PU.1 and Spi-B leads to B cell acute lymphoblastic leukemia associated with driver mutations in Janus Kinases

Precursor B-cell acute lymphoblastic leukemia (B-ALL) is associated with recurrent mutations that occur in cancer-initiating cells. There is a need to understand how spontaneous driver mutations influence clonal evolution in leukemia. The ETS-transcription factors PU.1 and Spi-B (encoded by Spi1 and Spib) execute a critical role in B cell development and serve as complementary tumour suppressors by opposing the proliferative events mediated by IL-7R signaling. Here, we used a mouse model to conditionally delete Spi1 and Spib genes in developing B cells. These mice developed B-ALL with a median time to euthanasia of 18 weeks. We performed RNA and whole-exome sequencing (WES) on leukemias isolated from Mb1-Cre{Delta}PB mice and identified single-nucleotide variants (SNVs) in Jak1, Jak3 and Ikzf3 genes, resulting in amino acid changes and in the gain of early stop-codons. JAK3 mutations resulted in amino acid substitutions located in the pseudo-kinase (R653H, V670A) and in the kinase (T844M) domains. Introduction of these mutations into wild-type pro-B cells conferred survival and proliferation advantages. We conclude that mutations in Janus kinases represent secondary drivers of leukemogenesis in the absence of Spi-B and PU.1 transcription factors. This mouse model represents an useful tool to study clonal evolution and tumour heterogeneity in B-ALL.

cancer biology↗