Skin-type antifreeze protein expression in integumental cells of larval winter flounder

Abstract

Wholemount in situ hybridization using an antisense riboprobe complementary to a winter flounder Pleuronectes americanus skin-type antifreeze protein mRNA (WFp9) and immuno histochemistry using polyclonal antibodies to the corresponding protein detected cells expressing this gene in larval winter flounder integument. Immunohistochemistry revealed two distinct populations of cells. One population extended laterally along the length of the fish and was detectable using in situ hybridization. Staining in these cells declined following yolk-sac absorption suggesting that expression was only important here during early larval development. The polyclonal antibody for skin-type antifreeze protein also reacted with another population of cells scattered throughout the integument. These cells stained with alcian blue suggesting that they were integumental mucous cells. In situ hybridization using the above probe was not able to detect the corresponding transcript within the same cells. This suggests that another gene may be involved in the production of a similar protein in this case. These data suggest that two distinct populations of cells within the larval integument are involved in skin-type antifreeze protein expression and possibly involve the activity of at least two different genes.