A novel kinase, AATYK induces and promotes neuronal differentiation in a human neuroblastoma (SH-SY5Y) cell line

Apoptosis Associated Tyrosine Kinase (AATYK), a novel protein recently isol ated from differentiating 32D mouse myeloid cells, contains a putative tyro sine kinase domain and several binding motifs for src homology 2 (SH-2) and src homology 3 (SH-3) domain containing proteins. We observed that AATYK i s expressed in different regions of the brain. Although it might play a rol e in normal nervous system development by modulating apoptosis, little is k nown regarding its function in the brain or its intracellular localization and kinase activity. Recognizing its homology with Insulin-like growth fact or-I (IGF-I) receptor (IGF-IR) and the critical role of IGF-I in neuronal s urvival, we hypothesized that AATYK plays an important role in neuronal dif ferentiation/apoptosis. To test this hypothesis, we transfected the human a drenergic neuroblastoma (NB):SH-SY5Y cells with AATYK cDNA under a tetracyc line-repressible promoter and established stable cell lines that readily ex press AATYK on removal of tetracycline. AATYK immunoprecipitated from these cell lysates is an active kinase. Indirect immunofluorescent staining of t he clones revealed AATYK to be localized in the cytoplasm. By itself, AATYK overexpression fur short duration (2-3 days) did nor induce differentiatio n in the stable SH-SY5Y clones. On the other hand, overexpression for longe r periods (7-8 days) per se, significantly (P<0.05-0.001) increased the per cent of differentiated cells as well as the neurite length. AATYK-induced d ifferentiation was in the same range as the differentiation induced by agen ts like all-trans retinoic acid (RA), 12-O-Tetradecanoyl phorbol 13-acetate (TPA) and IGF-I. In addition, AATYK significantly promoted the neuronal di fferentiation induced by these agents. Our results demonstrate for the firs t time that AATYK is an active, non-receptor, cytosolic kinase which induce s neuronal differentiation and also promotes differentiation induced by oth er agents in the SH-SY5Y cells. (C) 2000 Elsevier Science B.V. All rights r eserved.