Three novel LIM domain homeobox cDNAs encoding proteins structurally related to the Isl-1 protein were isolated from a chinook salmon pituitary cDNA library. Southern blot analysis of genomic DNA indicate that they are derived from three distinct genes, designated as isl-2a, isl-2b, and isl-3 genes. Nucleotide sequence analysis of reverse transcriptase-polymerase chain reaction amplified products reveal that the isl gene family contains two members (a and b) each of both isl-1 and isl-2 genes, and one member of isl-3 gene in the two tetraploid salmonid species, chinook salmon and rainbow trout, and only one member each of isl-1, isl-2, and isl-3 genes in the diploid zebrafish. The expression of the three isl genes in the rainbow trout were studied by reverse transcriptase-polymerase chain reaction analysis of embryonic and adult RNAs, and by in situ hybridization analysis of 8-week-old hatchlings. The transcripts of all three genes could be detected as early as 4 weeks postfertilization (the eye stage) and increased dramatically in 5-week-old embryos. In the adult, the three isl mRNAs appear to be differentially distributed in various tissues. The level of isl-1 mRNA is generally higher than those of isl-2 and isl-3 mRNAs. In situ hybridization analysis indicates that the transcripts of all three genes are localized in subsets of neurons in the brain and spinal cord. In the retina, isl-1 mRNA could be found in both the ganglion and inner nuclear layers while isl-2 and isl-3 mRNAs could only be detected in the ganglion layer. High level of isl-1 mRNA could also be found in mid-gut and interrenal organ where endocrine cells are densely populated. Based on these observations, we speculate that the three structurally related isl genes may play similar roles in cell determination and differentiation in the developing nervous system.

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