Abstract
Abstract: :
Purpose: In zebrafish, gelsolin–1 (previously called C/L–gelsolin) is expressed highly in cornea (50% of water–soluble protein) and slightly in lens and early cleavage–stage embryos. Since this is an atypical expression pattern for vertebrate gelsolin, we searched for a second gelsolin gene. Methods: Gelsolin–2 was cloned by PCR based on EST data available from the GenBank database. Sequences for in situ hybridization probes were constructed in pBS–sk(–) for gelsolin–2 and pCS2 for gelsolin–1. Probes were labeled with the DIG kit from Roche. Multiple alignments were generated with clustalW from sequences obtained from GeneBank. Dendograms were created from the clustalW results and formatted with Tree View. Results: We cloned a second zebrafish gelsolin (gelsolin–2) which exhibits 56% identity to corneal–preferred zebrafish gelsolin–1 at the protein level. The gelsolin–2 protein has 71% identity to human gelsolin compared to 58% for gelsolin–1. As expected, gelsolin–2 exhibits six conserved gelsolin motifs. Phylogenetic analysis demonstrates that gelsolin–1 and gelsolin–2 diverged early in vertebrate evolution and that gelsolin–2 gave rise to the other vertebrate gelsolins. Gelsolin–2 mRNA is widely expressed at a low level in the embryo and adult; it does not accumulate in the cornea as gelsolin–1. Comparison of up to 10 kb of the upstream regions of these two genes indicated that only a few short stretches of sequence are comparable. In addition, the gene structures of the gelsolin–1 and gelsolin–2 differ, with 18 introns and 20 exons, and 16 introns and 17 exons for gelsolin–1 and gelsolin–2, respectively. Conclusions: The differences in gene structures and upstream putative regulatory sequences driving their diverse patterns of expression suggest that the two zebrafish gelsolin genes have undergone adaptive changes in regulation, consistent with functional differences for the proteins, following gene duplication.
Keywords: cornea: basic science • cornea: epithelium