Abstract
T-cell receptor gene beta (TCRβ) gene rearrangement represents a complex, tightly regulated molecular mechanism involving excision, deletion and recombination of DNA during T-cell development. RUNX1, a well-known transcription factor for T-cell differentiation, has recently been described to act in addition as a recombinase cofactor for TCRδ gene rearrangements. In this work we employed a RUNX1 knock-out mouse model and demonstrate by deep TCRβ sequencing, immunostaining and chromatin immunoprecipitation that RUNX1 binds to the initiation site of TCRβ rearrangement and its homozygous inactivation induces severe structural changes of the rearranged TCRβ gene, whereas heterozygous inactivation has almost no impact. To compare the mouse model results to the situation in Acute Lymphoblastic Leukemia (ALL) we analyzed TCRβ gene rearrangements in T-ALL samples harboring heterozygous Runx1 mutations. Comparable to the Runx1+/− mouse model, heterozygous Runx1 mutations in T-ALL patients displayed no detectable impact on TCRβ rearrangements. Furthermore, we reanalyzed published sequence data from recurrent deletion borders of ALL patients carrying an ETV6-RUNX1 translocation. RUNX1 motifs were significantly overrepresented at the deletion ends arguing for a role of RUNX1 in the deletion mechanism. Collectively, our data imply a role of RUNX1 as recombinase cofactor for both physiological and aberrant deletions.
Highlights
T-cell receptor gene beta (TCRβ) gene rearrangement represents a complex, tightly regulated molecular mechanism involving excision, deletion and recombination of DNA during T-cell development
Whereas wildtype and Runx1+/− thymi presented with a distinct cortex and medullary zone, loss of cortex and medulla structures was observed in Runx1−/− mice (Fig. 2d)
We investigated the frequency of RUNX1 motifs overlapping with either single heptamer sites or heptamers associated with recombination signal sequences (RSS) modules at the borders of recurrent ETV6-RUNX1 Acute Lymphoblastic Leukemia (ALL) deletions in comparison to corresponding motifs from the whole genome
Summary
T-cell receptor gene beta (TCRβ) gene rearrangement represents a complex, tightly regulated molecular mechanism involving excision, deletion and recombination of DNA during T-cell development. Thereby functional IG or TCR receptors are assembled from preexisting sets of Variable (V), Joining (J) and — in case of TCRβ, TCRδ and IGH — from additional Diversity (D) gene segments[16] These segments are flanked by recombination signal sequences (RSS) composed of conserved heptamer and nonamer sequences separated by a spacer of 12 or 23 base pairs (Fig. 1). The RUNX1 DNA-binding core motif TGTGGNNN overlaps with the RSS heptamer and nonamer motifs which are crucial for the recombination of IG and TCR gene segments (Fig. 1) This raises the possibility that RUNX1 might act as a recombinase cofactor for both physiological and non-physiological deletions
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