PAK-dependent STAT5 serine phosphorylation is required for BCR-ABL-induced leukemogenesis

Abstract

The transcription factor STAT5 (signal transducer and activator of transcription 5) is frequently activated in hematological malignancies and represents an essential signaling node downstream of the BCR-ABL oncogene. STAT5 can be phosphorylated at three positions, on a tyrosine and on the two serines S725 and S779. We have investigated the importance of STAT5 serine phosphorylation for BCR-ABL-induced leukemogenesis. In cultured bone marrow cells, expression of a STAT5 mutant lacking the S725 and S779 phosphorylation sites (STAT5SASA) prohibits transformation and induces apoptosis. Accordingly, STAT5SASA BCR-ABL+ cells display a strongly reduced leukemic potential in vivo, predominantly caused by loss of S779 phosphorylation that prevents the nuclear translocation of STAT5. Three distinct lines of evidence indicate that S779 is phosphorylated by group I p21-activated kinase (PAK). We show further that PAK-dependent serine phosphorylation of STAT5 is unaffected by BCR-ABL tyrosine kinase inhibitor treatment. Interfering with STAT5 phosphorylation could thus be a novel therapeutic approach to target BCR-ABL-induced malignancies.

Emerging therapeutic targets in myeloproliferative neoplasms and peripheral T-cell leukemia and lymphomas

“…RAS-RAF signaling can be triggered by normal cytokine, growth factor, or hyperactive JAK action and it constitutes a core cancer pathway [7]. RAS-RAF is upstream of MAPK-ERK signaling and it can further activate RHO and RAC GTPase proteins, which were reported to be essential for nuclear shuttling of STAT5A [42]. Furthermore, oncogenic RAS requires mitochondrial functions of STAT3, illustrating that the RAS-RAF pathway is interconnected with aberrant JAK-STAT, cytokine, or growth factor signaling.…”

Expansion of BCR/ABL1+ cells requires PAK2 but not PAK1

“…Whole‐cell lysates were harvested as described previously (Berger et al , 2014). Proteins were separated on a 7% sodium dodecyl sulphate polyacrylamide gel and transferred to nitrocellulose membranes.…”

“…Single‐cell suspensions were analysed by a BD FACS Canto II flow cytometer equipped with 488, 633, and 405 nm lasers using FACS Diva software (Becton Dickinson, Franklin Lakes, NJ, USA) as described before (Berger et al , 2014). Propidium iodide (PI) and apoptosis stainings were performed as described previously (Berger et al , 2014) and according to manufacturer's instructions (Annexin V Apoptosis Detection Kit eFluor ® 450, 88‐8006; eBioscience, San Diego, CA, USA).…”

“…Propidium iodide (PI) and apoptosis stainings were performed as described previously (Berger et al , 2014) and according to manufacturer's instructions (Annexin V Apoptosis Detection Kit eFluor ® 450, 88‐8006; eBioscience, San Diego, CA, USA). eGFP + or mCherry + KU812 cells were high‐purity FACS sorted on a FACS Aria III equipped with a 488 nm laser at 4°C (Becton Dickinson).…”

“…The gain of 3q29 is associated with poor prognosis (Beà et al , 2004; Bea et al , 2005; Salaverria et al , 2007). In BCR/ABL1 + leukaemia, PAK1 and PAK2 are upstream regulators of the transcription factor STAT5 – a key node for initiation and maintenance of disease (Hoelbl et al , 2006, 2010; Friedbichler et al , 2010; Berger et al , 2014). …”