Serotonin activates s6 kinase in a rapamycin-sensitive manner in Aplysia synaptosomes

The identification of tags that can specifically mark activated synapses is important for understanding how long-term synaptic changes can be restrict ed to specific synapses. The maintenance of synapse-specific facilitation i n Aplysia sensory to motor neuron cultures can be blocked by inhibitors of translation and by the drug rapamycin, which specifically blocks a signalin g pathway that regulates phosphorylation of translational regulators. One i mportant target of rapamycin is the phosphorylation and subsequent activati on of S6 kinase. To test whether S6 kinase is the target for the ability of rapamycin to block synapse-specific facilitation in Aplysia, we cloned Apl ysia S6 kinase, its substrate S6, and the S6 kinase kinase phosphoinositide -dependent kinase 1 (PDK-1). Serotonin, which induces synapse-specific faci litation, increased phosphorylation of Aplysia S6 kinase at threonine 399 i n a rapamycin-sensitive manner in Aplysia synaptosomes. The phosphorylation of threonine 399 by 5-HT was independent of phosphoinositide-3 kinase, dep endent on PKA and PKC, and occluded by the phosphatase inhibitor calyculin- A. 5-HT also increased S6 kinase activity and led to increased phosphorylat ion of S6 in synaptosomes. 5-HT increased levels of S6 in synaptosomes beca use of a rapamycin-sensitive increase in translation-stabilization of S6. A plysia PDK-1 bound to and phosphorylated Aplysia S6 kinase but only modulat ed phosphorylation of threonine 399 indirectly. These results suggest a mec hanism by which the levels of translation factors can be increased specific ally at activated synapses generating a longlasting synaptic tag.