Transcription Factor Phosphorylation by pp90 rsk2 IDENTIFICATION OF Fos KINASE AND NGFI-B KINASE I AS pp90 rsk2

Abstract The in vitro phosphorylation of transcription factors by growth factor-activated protein kinases has resulted in the discovery of a number of activities whose identities and relationships to one another are unclear. Fos kinase is a growth factor-stimulated serine/threonine protein kinase that phosphorylates c-Fos at serine 362 within the carboxyl-terminal regulatory domain. Fos kinase activation is dependent on p21ras and mitogen-activated protein kinase/ERK kinase kinase (MEK) activity and is independent of phosphatidylinositol 3-kinase activity. We have purified Fos kinase by affinity chromatography using the Sepharose-linked protein kinase inhibitor, bisindolylmaleimide (BIM). Fos kinase has an apparent molecular mass of 88 kDa, and mass spectrophotometric analysis of the isolated protein showed that it produced tryptic fragments identical to those predicted for pp90rsk2. Fos kinase isolated from nerve growth factor-stimulated PC12 cells is indistinguishable from NGFI-B kinase I, based on their chromatographic behavior, substrate specificities, and relative sensitivity to BIM. Furthermore, we have distinguished Fos kinase from calcium/cAMP response element-binding protein (CREB) kinase. Therefore, Fos kinase and NGFI-B kinase I and pp90rsk2 represent the same protein kinase species. Moreover, we report that pp90rsk2 exists within nerve growth factor-stimulated PC12 cells as two chromatographically and immunologically distinct species. Finally, we demonstrate that CREB kinase is distinct from pp90rsk2.