Recombinant limb buds constructed of an ectodermal jacket containing d
issociated anterior wing bud mesoderm cells form limb-like structures
lacking anteroposterior polarity. Polarity can be restored by implanti
ng a small piece of polarizing region into the recombinant limb. We us
ed recombinant limbs to study expression of Msx1 and Msx2 and 5' Hoxd
genes in limb development. Detectable mesodermal expression of Msx1 an
d Msx2 was lost during preparation of the recombinants. When expressio
n was restored both genes were expressed in a symmetrical fashion in t
he apical mesoderm whether or not the recombinant was polarized. The n
ormal anterior pattern of Msx1 and Msx2 was never reestablished. The r
oles Msx1 and Msx2 play in limb patterning are at present not clear. I
n nonpolarized recombinant limbs the 5' Hoxd genes were sequentially a
nd uniformly expressed in the subridge mesoderm across the anteroposte
rior axis. They exhibited nested domains of expression showing exclusi
vely distal restriction. In polarized recombinants the 5' Hoxd genes w
ere expressed progressively in an asymmetric, nested fashion adjacent
to the implanted polarizing region; there was also a distal mesodermal
region further from the implant that did not express the most 5' Hoxd
genes. Both of these observations are similar to the normal limb. Our
results demonstrate that the polarizing region is not necessary for t
he activation or maintenance of 5' Hoxd gene expression but that its p
resence is required in order for the normal pattern to be established.
We propose that the expression of the Hoxd genes is an intrinsic prop
erty of rapidly growing limb cells and that their expression is relate
d to the limb bud proximodistal outgrowth. Our data also support the i
nfluence of the polarizing region upon Hoxd gene expression in two way
s. First there is a short-range action responsible for the normal asym
metric expression and second a long-range effect suppressing the expre
ssion of 5'-Hoxd genes by the anterior cells. (C) 1994 Academic Press,
Inc.