FOXO1 forkhead domain mutants in B-cell lymphoma lack transcriptional activity

Somatic point mutations of the FOXO1 transcription factor were reported in non-Hodgkin lymphoma including diffuse large B-cell lymphoma, follicular lymphoma and Burkitt lymphoma. These alterations were associated with a poor prognosis and resistance to therapy. Nearly all amino acid substitutions are localized in two major clusters, affecting either the N-terminal region (Nt mutations) or the forkhead DNA-binding domain (DBD mutations). While recent studies have focused on Nt mutations, we characterized FOXO1 DBD mutants. We analyzed their transcriptional activity, DNA binding, phosphorylation and protein-protein interaction. The majority of DBD mutants showed a decrease in activity and DNA binding, while preserving AKT phosphorylation and interaction with the cytoplasmic ATG7 protein. In addition, we investigated the importance of conserved residues of the α-helix 3 of the DBD. Amino acids I213, R214, H215 and L217 appeared to be crucial for FOXO1 activity. Our data underlined the key role of multiple amino-acid residues of the forkhead domain in FOXO1 transcriptional activity and revealed a new type of FOXO1 loss-of-function mutations in B-cell lymphoma.