Along-standingquestioninbiologyishowcellsrespondtomultiplesignalinginputswithaspecificresponse. ThisproposalinvestigatestheintersectionsofNotchandHhsignalingpathways,whicharewidelyrequired duringdevelopment,andbasicmechanismscongenitaleyedisease.Bothsignalingsystemsregulatecell proliferation,morphogenesis,cellpolarity,differentiation,apoptosisandstemcellmaintenance.However,the spatiotemporalcontextsforeachdiffersatthecellularandtissuelevelsduringopticvesiclemorphogenesis andgrowth.Thisproposalwilluseinvivocomplexconditional(cre-lox)mousegenetics,mousetransgenics, embryology,iPSC-derivedretinalorganoids,histology,immunohistochemistry,confocalmicroscopy,insitu hybridization,qPCRandbioinformaticstechnologiestoinvestigatebasic,mechanisticquestionsaboutthe initiationandprogressionofeyeformationfromtheembryonicbrain.Wewilladdressseveralimportant questions:1)WhatarethespatialandtemporalconstraintsforNotchversusHhincontrollinggrowth, morphogenesis,patterning,tissuepolarityandthetimingofdifferentiationintheopticvesicle,opticcup,RPE andopticstalk?3)WhataretheoptimalconditionsbywhichNotchandShhpathwayscontributetothe generationofretinalganglioncellandconephotoreceptorneuronsinretinalorganoidculture?1)Howdothe NotchandHhpathwaysregulatetheeyeexpressiondomainsofacommontargetgene,Hes1?
Thisresearchwillprovidefundamentalunderstandingofopticvesicleoutgrowthfromtheventralthalamusand morphogenesisintotheopticcup,RPEandstalk.Suchinformationultimatelyinformsthepathogenesisand/or treatmentofcongenitaldiseaseslikeanophthalmia,microphthalmia,coloboma,cyclopia,septo-opticdysplasia oropticnervehypoplasia.Thisstudywillcreatenewtransgenicandmutantmousemodelsandprovidenew insightsintoretinalstemcellbiology,twoovertgoalsoftheNationalEyeInstitute.Moreover,mechanistic informationabouttheNotchandHhsignalingpathwayswillbegained,withbroadimplicationsfor developmental,stemcellandcancerbiology.
