Interactome Analysis Identifies Novel Targets of Phosphoinositide 3-kinase (PI3K) that Mediate Astrocyte Neuroprotection

In a homeostatic state, astrocytes can support neuronal survival and function through a range of secreted signals that protect against neurotoxicity, oxidative stress, and apoptotic cascades. Thus, the analysis of astrocyte conditioned media (ACM) may provide valuable insight into the nature of these protective mechanisms, and how they might be promoted. Previously, we characterized a potent neuroprotective activity mediated by ACM in neurons and the retina in metabolic stress models. However, the molecular entity and mechanism underlying this activity remained unclear. Here, a chemical genetics screen revealed phosphoinositide 3-kinase (PI3K) as a central player transducing ACM-mediated neuroprotection. To identify additional proteins contributing to the protective activity, endogenous neuronal PI3K was immunoprecipitated from astrocytes exposed to ACM or control media, and MS/MS analyses were undertaken. MS data analyses pointed toward only five additional proteins that co-immunoprecipitated with PI3K and were regulated by the ACM signal. These hits included expected PI3K interactors, such as the platelet-derived growth factor receptor A (PDGFRA), and novel interactors, such as the zinc finger CCCH-type containing 14 (ZC3H14). ZC3H14 has recently emerged as an important RNA binding protein that modifies poly-adenosine tail lengths on nascent mRNA transcripts. In downstream validation studies we show that Platelet Derived Growth Factor-BB (PDGF-BB) strongly activates PI3K signaling to protect neuronal cells. Finally, PDGF-BB treatment induced recruitment of ZC3H14 to PI3K, and inhibiting this interaction eliminated ACM-mediated neuroprotection. Thus, we identified a novel ACM- and PDGF-induced neuroprotective signaling cascade mediated through PI3K that involves recruitment of ZC3H14. Enhancing this pathway may present a promising strategy to promote astrocyte-secreted neuroprotective signals.

Department dc:contributor.department

Laboratory Medicine and Pathobiology

Year dc:date.issued

2019

Author dc:creator

• Alqawlaq, Samih Ahmad

Advisor dc:contributor.advisor

• Sivak, Jeremy

• Astrocyte
• Astrocyte Conditioned Media
• Glaucoma
• Neuroprotection
• PI3K Interactome
• ZC3H14

Handle dc:identifier.uri

http://hdl.handle.net/1807/97042

OAI identifier oai:identifier

oai:utoronto.scholaris.ca:1807/97042