Focus on molecules: Pax-6, the eye master.

1. StructurePax-6 belongs to the family of paired box genes that containboth the hallmark paired box domain (PD) and a homeo box domain(HD), followed by a prolineeserineethreonine rich domain (PST)(Fig. 1A). The Pax-6 protein contains 422 aa and at least one tran-script variant, Pax-6-5a that contains a 14 aa insert in the PD (atamino acid position 47). The PD binds DNA in a bipartite fashion us-ing the N-terminal and C-terminal subdomains. The 5a insert abro-gates DNA binding by the N-terminal subdomain suggesting thatthe C-terminal subdomain dictates target speciﬁcity in this variant.Thestructureofthepairedboxdomainincomplexwitha26-bpop-timalDNAduplexhasbeendetermined(Xuetal.,1999)(Fig.1B).Thisstructure provides a detailed model of the interactions between Pax-6PD and DNA, and in particular how the N-, C-terminal subdomainsandlinkerregioncombinetoachieveDNAbindingspeciﬁcity.Specif-ically, both the N- and C-terminal subdomains fold into a helix-turn-helix motif, reminiscent of the homeo box domain fold (Xu et al.,1999).TheprimarysitesofDNAinteractionoccurbytheso-called‘‘rec-ognition’’helicesa3anda6(Fig.1B).Indeed,residue47ina3(andres-idues 42 and 44 to a lesser degree) dictates DNA speciﬁcity within thePax family. Interestingly, the linker between the N- and C-terminaldomains is also involved in DNA recognition and speciﬁcity. Finally,the structure also provides a framework for understanding the effect ofmutations known to be involved in disease (Fig. 1C, see below).2. FunctionPax-6 is a transcriptional factor involved in the development ofthe central nervous system and eye development. The correspondinggene in Drosophila is the eyeless, a mutation in the eyeless gene re-sults in animals with no eyes. However, mutations in Pax-6 cause noeye or small aye phenotype in mammals as well. Pax-6 has beenconsidered as the master gene for eye development. Indeed, initialstudies showed that ectopic expression of Pax-6 in Drosophila couldproduce ectopic eyes. Pax-6, however, seems to be involved ina feedback loop with another homeo box-containing gene, Six-3,in order to control development of the eye. Pax-6 also interactswith the homeo box-containing genes Pbx1 and HoxB1 and this in-teraction enhances its transcriptional activity. Except for being a mas-ter gene for eye development, Pax-6 plays signiﬁcant roles duringthe induction of the lens and retina differentiation. Inactivation ofPax-6 in the surface ectoderm after E9.5 resulted in arrest of lensdevelopment. Pax-6 affects differentiation of lens ﬁbers cells bycontrolling crystallin gene expression. Inactivation of Pax-6 affectsthe retinogenic potential of retinal progenitor cells (RPCs). RPCs be-come restricted to one cell fate that of amacrine interneurons. Also,regulation of Pax-6 expression controls the identity and differentia-tion potential of retinal pigment epithelial cells (for reviews seeGehring, 2002; Treisman, 2004).3. Disease involvementAswasmentionedabove,mutationsinPax-6leadtoeyelessorsmalleye phenotypes in mice. In humans, Pax-6 mutations are associatedwithaniridia(Fig. 1C).Othermutations inPax-6 have beenassociatedwithfovealhypoplasia,presenilecataract,aniridia-relatedkeratopathy,cranialandCNSmalformations.Mostofthemutationsappeartocauseloss of function (van Heyningen and Williamson, 2002).4. Future studiesWhilethefunctionofPax-6hasbeenstudiedquiteextensivelyanditsdevelopmentalroleisknown,itscooperationwithothergenesneedsfur-ther investigation. It is known than Pax-6 is part of a loop that involvesSix-3aswell.IdentiﬁcationofmoredownstreamtargetsforthispathwaywillhelpdelineatethemechanismsofthedifferentactionsofPax-6.AlsoresearchongeneswhosePax-6isthetargetmightshedlightonhowmas-ter genes are activated in undifferentiated cells. Recent work indicatesthatthePDandHDcanbindDNAinacooperativefashion.Furthermore,the PST domain seems to be important in transactivation and several