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All retinoic acid receptors (RAR alpha, beta, gamma) have two isoforms, whose function is unknown. We now show that at least for RAR gamma, the isoforms are differentially distributed in the embryo. RAR gamma 1 and RAR gamma 2 are detected in the head region, whereas RAR gamma 2 is the sole isoform expressed in the tail region. Specifically, it is expressed in the chordoneural hinge, a region of the tailbud that has organizing properties. Treatment with high doses of retinoic acid (RA) reduces expression in this region. The results are discussed in terms of the known teratogenic effects of RA in the tail region.
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8199051
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Fig. 1 (Ieft). RARgamma is expressed in anterior and posterior domains. Whole-mount in situ hybridization was performed using a common RARgamma probe that detects both RARgamma isoforms. (A) Expression is first detected in the dorsal lip of stage 11 midgastrula embryos. During neurulation
(B), stage 19, the endomesoderm of the head and anterior neural tissue are labeled in the anterior domain. The three germ layers are stained in
the posterior domain. At late tailbud stages (C), stage 32, expression is limited to the branchial arches and the tip of the tail and posterior spinal
cord. The embryo in (C) contains pigment cells seen as brown spots. DL, dorsal lip: ANT, anterior domain; POS, posterior domain; BA,
branchial arches; TB, tailbud; bar length, 200 um.
Fig. 2 (right). RAR gamma isoforms are differentially expressed. Specific probes were hybridized to late neurula/early tailbud embryos. (A) Diagram
indicating position and size of isoform-specific probes, which were derived from clones isolated by Blumberg et al. (1992). (B) RARgammaI expression
is seen solely in the anterior expression domain. (C) RARgamma2 is expressed both in the headendomesoderm and in the posterior prospective
tailbud area. (D) A control sense probe shows no staining after an equal time of incubation. Artifactual precipitates of proteinaceous material
can be seen in the gut cavity (see D), Bar, 200 umm.
Fig. 3. RARgamma isoforms display differential distribution at tailbud stages. (A,B), RARgammal expression in head and tail respectively'. Note weak
expression in the branchial arches (A) and the absence of expression in the tailbud (B). (C.D), RARgamma2 expression in head and tail respectively.
Expression can be seen in branchial arches as well as in the tailbud region. The distribution of RARgamma2 in the tail is indistinguishable from that
obtained using the full length RARgamma common probe. Bar, 200 um
Fig. 4 RARgamma is expressed in the chordoneural hinge. (A) Diagram indicating the anatomy of the tailbud region, (B-D) Adjacent sagittal sections
of a stage 32 tailbudembryo hybridized to the full length RARgamma probe. Staining can be clearly seen to lie anterior to the neurenteric canal,
encompassing the chordoneural hinge and extending anteriorly along the floor plate of the neural tube. The diagram shown in panel (A) is
derived from the examination of serial sections. EN, endoderm: FP, floor plate of the spinal cord: NEC, neurenteric canal: PAG, postanal gut;
PW, posterior wall; SO, somite. Bar. 200 um
Fig. 5. RA decreases RARgamma tailbud expression at teratogenic doses. Whole mount in situ hybridizations with a full-length RARgamma probe showing.
(A) untreated embryos cultured in 0.1 � Barth's medium, (B-D) embryos treated from stage 13 onwards with 10-8, 10-7 , 10-6M all-trans RA, respectively. Chordoneural hinge expression is decreased in (D). Arrowheads indicate the position of the malformed postanal gut. Bar, 200 um.
