ABSTRACT The LIM class homeobox gene Xlim-1 is expressed in Xenopus embryos in the lineages leading to (i) the notochord, (ii) the pronephros, and (iii) certain cells of the central nervous system (CNS). In its first expression phase, Xlim-1 mRNA arises in the Spemann organizer region, accumulates in prechordal mesoderm and notochord during gastrulation, and decays in these tissues during neurula stages except that it persists in the posterior tip of the notochord. In the second phase, expression in lateral mesoderm begins at late gastrula, and converges to the pronephros at tailbud stages. Expression in a central location of the neural plate also initiates at late gastrula, expands anteriorly and posteriorly, and becomes established in the lateral regions of the spinal cord and hindbrain at tailbud stages. Thus Xlim-1 expression precedes morphogenesis, suggesting that it may be involved in cell specification in these lineages. Enhancement of Xlim-1 expression by retinoic acid (RA) was first detectable in the dorsal mesoderm at initial gastrula. During gastrulation and early neurulation, RA strongly enhanced Xlim-1 expression in all three lineages and also expanded its expressing domains; this overexpression correlated well with RA phenotypes such as enlarged pronephros and hindbrain-like structure. Exogastrulation reduced Xlim-1 expression in the lateral mesoderm and ectoderm but not in the notochord, suggesting that the second phase of Xlim-1 expression requires mesoderm/ectoderm interactions. RA treatment of exogastrulae did not revert this reduction.