Mlk3 Binding Protein Jnk Pathway In Cerebral Ischemia

Transient, severe global ischemia in rats and gerbils leads to selective and delayed neuronal death of pyramidal neurons in the hippocampal CA1, which is not detected until 2-4 d after induction of ischemia. Several lines of evidence have been proposed to suggest that apoptotic process underlies the molecular mechanism of this delayed neuronal cell death. Considerable evidence suggests that c-Jun NH2-terminal kinase (JNK) is an important kinase mediating the neuronal cell death in response to cerebral ischemia. Recently, an intracellular serine/threonine kinase, mixed-lineage kinase 3 (MLK3) has been identified as a novel upstream activator of the JNK pathway. MLK3 functions as a MAPK kinase kinase (MAPKKK) of the JNK stress pathway by directly phosphorylating and activating the JNK activators SEK1/MKK4 and MKK7. MLK3 has garnered attention as an important mediator of JNK-mediated neuronal apoptosis and ischemic injury. However, the molecular mechanisms that regulate MLK3 activity in cerebral ischemic-reperfusion has not been extensively delineated. The overall goal of the present study is to understand the molecular basis by which MLK3 and its signaling pathways are negatively regulated by targeting MLK3 interacting proteins in cerebral ischemia.1. POSH is an important mediator of JNK pathway activation and ischemic injury by compositing the POSH/MKK4/JNK signaling complex in vulnerable CA1 region following transient global ischemia.We investigated the expression and subcellular localization of the multidomain protein POSH (plenty of SH3s) by immunohistochemistry and Western blot analysis in rat hippocampal CA1 region following cerebral ischemia. Our results indicated that the cytosol immunoreactivity of POSH was strong in CA1-CA3 pyramidal cell but weak in DG granule cell of rat hippocampus both in sham control and after reperfusion. Coimmunoprecipitation experiments showed that the interactions of MLK3, MKK4 and phospho-JNKs with POSH were persistently enhanced during the early (30 min) and the later reperfusion period (from 1 d to 3 d) compared with sham...