Astrocytes can support neuronal survival through a range of secreted signals that protect against neurotoxicity, oxidative stress, and apoptotic cascades. Thus, analyzing the effects of the astrocyte secretomemay provide valuable insight into these neuroprotective mechanisms. Previously, we characterized a potent neuroprotective activitymediated by retinal astrocyte conditionedmedia (ACM) on retinal and cortical neurons in metabolic stressmodels. However, the molecular mechanism underlying this complex activity in neuronal cells has remained unclear. Here, a chemical genetics screen of kinase inhibitors revealed phosphoinositide 3-kinase (PI3K) as a central player transducing ACM-mediated neuroprotection. To identify additional proteins contributing to the protective cascade, endogenous PI3Kwas immunoprecipitated fromneuronal cells exposed toACMor controlmedia, followed by MS/MS proteomic analyses.These data pointed toward a relatively small number of proteins that coimmunoprecipitated with PI3K, and surprisingly only five were regulated by theACMsignal. These hits included expected PI3K interactors, such as the plateletderived growth factor receptor A (PDGFRA), as well as novel RNA-binding protein interactors ZC3H14 (zinc fingerCCCH-Type containing 14) and THOC1 (THO complex protein 1). In particular, ZC3H14 has recently emerged as an important RNA-binding protein with multiple roles in posttranscriptional regulation. In validation studies, we show that PI3K recruitment of ZC3H14 is necessary for PDGF-induced neuroprotection and that this interaction is present in primary retinal ganglion cells. Thus, we identified a novel non cell autonomous neuroprotective signaling cascade mediated through PI3K that requires recruitment of ZC3H14 and may present a promising strategy to promote astrocyte-secreted prosurvival signals.

An endogenous PI3K interactome promoting astrocyte-mediated neuroprotection identifies a novel association with RNA-binding protein ZC3H14

Datti A.
Methodology
;
2021

Abstract

Astrocytes can support neuronal survival through a range of secreted signals that protect against neurotoxicity, oxidative stress, and apoptotic cascades. Thus, analyzing the effects of the astrocyte secretomemay provide valuable insight into these neuroprotective mechanisms. Previously, we characterized a potent neuroprotective activitymediated by retinal astrocyte conditionedmedia (ACM) on retinal and cortical neurons in metabolic stressmodels. However, the molecular mechanism underlying this complex activity in neuronal cells has remained unclear. Here, a chemical genetics screen of kinase inhibitors revealed phosphoinositide 3-kinase (PI3K) as a central player transducing ACM-mediated neuroprotection. To identify additional proteins contributing to the protective cascade, endogenous PI3Kwas immunoprecipitated fromneuronal cells exposed toACMor controlmedia, followed by MS/MS proteomic analyses.These data pointed toward a relatively small number of proteins that coimmunoprecipitated with PI3K, and surprisingly only five were regulated by theACMsignal. These hits included expected PI3K interactors, such as the plateletderived growth factor receptor A (PDGFRA), as well as novel RNA-binding protein interactors ZC3H14 (zinc fingerCCCH-Type containing 14) and THOC1 (THO complex protein 1). In particular, ZC3H14 has recently emerged as an important RNA-binding protein with multiple roles in posttranscriptional regulation. In validation studies, we show that PI3K recruitment of ZC3H14 is necessary for PDGF-induced neuroprotection and that this interaction is present in primary retinal ganglion cells. Thus, we identified a novel non cell autonomous neuroprotective signaling cascade mediated through PI3K that requires recruitment of ZC3H14 and may present a promising strategy to promote astrocyte-secreted prosurvival signals.
2021
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11391/1496470
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