Genes Regulated by NPM-ALK Fusion Kinase Play a Key Role in the Activation of AP-1 Transcription Factors.

Background: About half of nodal anaplastic large cell lymphomas (ALCL) express the nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) fusion protein, which is the product of a t(2;5)(p23;q35) chromosomal translocation. Expression of this protein has been shown to result in neoplastic change. Combining suppression subtractive hybridization (SSH) and cDNA microarray analysis we aimed at elucidating the consequences of NPM-ALK expression.

Methods: SSH cDNA libraries were constructed using mRNA from human embryonic kidney (293) cells transfected with active or kinase-dead NPM-ALK constructs, as well as pools of NPM-ALK positive and negative ALCL cell lines. The resulting cDNA clones and genes relevant for cancer pathogenesis were spotted, generating specific cDNA microarrays comprising 4992 genes. mRNA expression patterns were analyzed in individual cell lines. Real time quantitative RT-PCR of 20 selected genes validated the mRNA expression data of the microarrays.

Results: Expression of a set of 102 genes distinguishes NPM-ALK-negative (FE-PD, MAC-2A) from NPM-ALK-positive ALCL cell lines (SU-DHL-1, JB-6, SUP-M2, SR-786, DEL and Karpas 299). The majority are involved in regulation of cell cycle and apoptosis. 38 of these genes also discriminate 293 cells with respect to their NPM-ALK expression. Interestingly, AP-1 target genes, such as GM-CSFRA, GM-CSFRB, ARF5, FAS, FASL, and BCL3, are increased in NPM-ALK expressing cell lines. Electrophoretic mobility shift assay (EMSA) verifies NPM-ALK dependent AP-1 DNA binding activity.

Conclusion: This study reveals genes specifically regulated by NPM-ALK lymphoma kinase. Further, we demonstrate that AP-1 activation is a critical target of NPM-ALK signalling.