REGULATION OF THE TYROSINASE GENE EXPRESSION IN THE ASCIDIAN EMBRYO

Tadpole larvae of the Japanese ascidian, Halocynthia roretzi, develop two sensory pigment cells in the brain, named otolith and ocellus. These melanin-containing cells specifically express tyrosinase enzyme-activity and play an important role for the detection of gravity and light. So far, genes encoding tyrosinase, a key enzyme in melanin biosynthesis, have been isolated from various vertebrate species. To carry out a phylogenetic study on the regulation of tyrosinase gene expression, we have cloned a putative tyrosinase gene from Halocynthia roretzi. The 5' flanking sequences of vertebrates have several conserved sequences. But their function as cis-elements is not uncertain. The 5' upstream region of the ascidian gene contains some of the similar sequences. To determine the requirements of the sequences for expression of the ascidian tyrosinase gene, the 5' flanking sequence was fused to a lacZ reporter gene and used for microinjection into fertilized ascidian eggs. A construct containing 1.8kb upstream of the gene expressed in four precursor cells of the pigment cells at the tailbud stage. Deletions of 300bp or more from the 5' end of the construct resulted in failure in to express the reporter gene. Analysis are now under way to map the responsible elements.