Hedgehog(Hh)proteinsarewidelydistributedthroughouttheanimalkingdomandplayimportantregulatoryrolesinvariousdevelopmentprocesses.ThethreeknownmammalianHhproteins,Sonic(Shh),Desert(Dhh)andIndian(Ihh)arestructurallyrelatedandshareahighdegreeofaminoacidsequenceidentity(e.g.ShhandIhhare93%identical).Although,HhproteinshaveuniqueexpressionpatternsanddistinctBIOLOGicalroleswithintheirrespectiveregionsofsecretion,theysignalthroughthesamePatched(Ptc)/Smoothened(Smo)receptorcomplex,activatethesameintracellularsignalingpathway,andcansubstituteforeachotherinexperimentalsystems.ThematurebiologicallyactiveformofHhmoleculesisproducedbyautocatalyticcleavageoftheirprecursorproteinsandcorrespondstoapproximatelytheN-terminalhalfoftheprecursormolecule.RecombinanthumanShhisa20.6kDaproteinconsistingof174aminoacidresidues,whichcorrespondtotheactiveN-terminalportionoftheShhprecursor.
Figure:invitrorelativecellproliferationassayinrenalcellcarcinomacelllinesinRenCa(PanelA)andACNHcells(PanelB)treatedwithdimethylsulfoxide.OnlyRenCacellsshowedthatCyblockedcellproliferationenhancedbyr-Shhtreatment.ACNHcellsdidnotshowanyinhibitingeffectofCyoncellproliferation.CellproliferationofVwassetas1.0.Thetreatmentswereperformedfor48hours.Eachpointrepresentstriplicateaverages±standarddeviationorON.AsterisksshowsignificantcellproliferationscomparedwithV.V,vehicle-treatedcontrol;S,recombinantsonichedgehog(r-Shh)protein(1µg/mL);Cy,cyclopamine(5µM).KoreanJUrol.2013Aug;54(8):547-554.http://dx.doi.org/10.4111/kju.2013.54.8.547
ADNAsequenceencodingaminoacidresiduesCys25-Gly198ofmouseShh(Echelard,Y.etal.,1993,Cell75:1417-1430)wasfusedtoa6Xhistidinetagatthecarboxy-terminus.ThefusionproteinwasexpressedinE.coli.
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