Ein neuer TGF-beta-induzierter Signalweg: ALK1 aktiviert den neuroprotektiven Transkriptionsfaktor NF-kappaB

Transforming growth factor-betas (TGF-betas) are pleiotropic cytokines involved in development and maintenance of the nervous system and other tissues in various physiologic and pathophysiologic conditions. Previous studies had revealed that application of TGF-betas resulted in a substantial pro-survival capacity of these cytokines in several neural lesion paradigms. TGF-beta1 exerted a significant protection from hypoxic, metabolic, excitotoxic, prooxidative or apoptotic stimuli. Notably, neurons treated with TGF-beta2 exhibited rather apoptotic features and decreased the activity of the neuroprotective transcription factor NF-kappaB. Furthermore, in cells outside the central nervous system, TGF-beta1 contributes to pro-apopotitic and pro-fibrotic processes. Over-expression of this cytokine in the liver leads to fibrosis and a prominent demise of cells. The major aim of the present study was to find explanations for this apparent discrepancy. The characterization of the molecular basis of the neuroprotective capacity and the elaboration of neuronal TGF-beta signal transduction pathways was used to shed light on these context dependent effects. TGF-beta-initiated signal transduction pathways are controlled by so called type II receptors which associate with type I receptors (activin-like kinases - ALKs) which initiate phosphorylation of intracellular effectors, the Smad proteins. BMPs (bone morphogenetic proteins, TGF-beta superfamily members) initiate phosphorylation of Smad1/5 proteins, whereas TGF-beta themselves transduce signals via ALK5 (TbetaR-I)-initiated phosphorylation of Smad2/3 proteins. Induction of the common TGF-beta signal transduction pathway via Smad2 and Smad3 in neuronal cells has been delineated in the course of the present study. Recently, an alternative type I receptor has been described in endothelial cells, the ALK1 receptor. ALK1 initiates phosphorylation of the Smad1/5 proteins as it previously had been shown for BMP-initiated signaling. Interestingly, the present study revealed for the first time neuronal expression of the ALK1 receptor. A prominent expression in neurons was noted which even was increased in animal models of ischemic injury; whereas glial cells did not exhibit significant expression of ALK1. In addition, incubation of hippocampal neurons with TGF-beta1 lead to a substantial increase in Smad1 phosphorylation and this behaviour could be mimicked by transfection of constitutively-active (ca) mutant forms of ALK1. The transcription factor NF-kappaB plays a fundamental role in neuroprotection. Treatment with TGF-beta1 enhanced NF-kappaB activity in gelshift and reporter-gene analyses. Homodimers, consisting of the p65 NF-kappaB subunit were identified as the effectors of this response. Of note, these effects could be mimicked by transfection of caALK1. Ectopic expression of caALK5, however, failed to result in NF-kappaB activation. In this line, treatment of hippocampal neurons with TGF-beta2, which had been shown to have a greater intrinsic activity on the ALK1 receptor than TGF-beta1, did not yield an increased transcriptional activity of NF-kappaB. Conclusion: The data presented suggest that TGF-beta1 initiates two receptor-mediated pathways in primary neurons. Differences in the expression levels of the receptors may account for the differential outcome of TGF-beta treatment in different tissues and under different cellular contexts. Ultimately, it has been delineated that TGF-beta1 initiates the ALK1 pathway in neurons resulting in subsequent NF-kappaB survival signaling.