The mes-metencephalic boundary (isthmus) has been suggested to act as an or
ganizer in the development of the optic tectum. Pax-5 was cloned as a candi
date for regulator of the organizing center. Isthmus-specific expression of
Pax-5 and analogy with the genetic cascade in Drosophila suggest that Pax-
5 may be at a higher hierarchical position in the gene regulation cascade o
f tectum development. To examine this possibility, a gain-of-function exper
iment on Pax-5 was carried out. in ovo electroporation on E2 chick brain wi
th the eucaryotic expression vector that encodes chick Pax-5 cDNA was used.
Not only was a considerable amount of Pax-5 expressed ectopically in the t
ransfected brain, but irregular bulging of the neuroepithelium was induced
in the diencephalon and mesencephalon. At Pax-5 misexpressing sites, uptake
of BrdU was increased. Histological examination of E7 transfected brain re
vealed that Pax-5 caused transdifferentiation of diencephalon into the tect
um-like structure. In the bulges of the E7 mesencephalon, differentiation o
f laminar structure was repressed when compared to the normal side, in tran
sfected embryos, En-2, Wnt-1 and Fgf8 were up-regulated ectopically, and Ot
x2 was down-regulated in the diencephalon to mesencephalon. Moreover, Ephri
n-A2, which is expressed specifically in the tectum with a gradient highest
at the caudal end, is suggested to be involved in pathfinding of the retin
al fibers, and was induced in the bulges. When the mouse Fgf8 expression ve
ctor was electroporated, Pax-5 and chick Fgf8 were also induced ectopically
. These results suggest that Pax-5, together with Fgf8, hold a higher posit
ion in the genetic hierarchy of the isthmus organizing center and regulate
its activity.