Abstract
Purpose: :
Homeodomain transcription factors Six3 and Six6 (with identical amino acid sequence in their homoedomains) exhibit temporospatial expression in mouse retinal development, with the expression of Six3 prior to that of Six6. Importantly, Six3 and Six6 are co-expressed in the immature and mature retinal progenitor cells. Conditional inactivation of Six3 at E10.5 retinas in Six6-null mice demonstrated that functional interactions between Six3 and Six6 are required for retinal development. We are testing the hypothesis that the maintenance of retinal progenitors and multi-lineage cell differentiation require the joint functions of homeobox genes Six3 and Six6 in mice.
Methods: :
CAGG-CreER or Pax6 α-Cre mice were used to conditionally delete floxed Six3 at E10.5 retinal cells in Six6 nulls.
Results: :
Individual inactivation of Six6 or Six3 in E10.5 retinal cells did not cause any major defect in the identity of retinal progenitors and terminal cell differentiation. In contrast, simultaneous inactivation of both Six3 and Six6 lead to the following severe defects in the maintenance of retinal progenitors and multi-lineage cell differentiation in E14.5 Six3;Six6 null retinas. 1) The expression of Sox2 (an essential factor that controls the competency of retinal progenitor cells) and its downstream target Notch1 was severely reduced or ablated. 2) The expression of Math5, Brn3b and Islet 1 (specific markers for ganglion cell differentiation) was absent or severely downregulated. In addition, the expression of Otx2 and Crx (specific markers for early photoreceptor cell differentiation) was absent or severely reduced. In contrast, the expression of NeuroD (an important factor that is involved in amacrine cell differentiation) was largely unaffected. 3) The expression of Otx1 (a specific marker for ciliary margin) and Axin2 (an endogenous readout of Wnt/β-catenin signaling that promotes ciliary margin cell fate) was significantly elevated. 4) Pax6 expression (an important transcription factor that is highly expressed in wildtype ciliary margin and moderately expressed in retinal progenitor cells (RPCs) and is required for RPC multipotency) was increased to the level of Pax6 expression in wildtype ciliary margin. This elevated Pax6 expression in Six3;Six6 null retinas likely reflects Pax6 expression in ciliary margin. 5) The expression of Rax (an important transcription factor that is required for optic vesicle formation and Otx2 expression) was largely unaffected.
Conclusions: :
Maintenance of retinal progenitors and multi-lineage cell differentiation require the joint functions of homeobox genes Six3 and Six6 in mice.
Keywords: development • differentiation • retina