PHYSIOLOGICAL AND EVOLUTIONARY IMIPLICATIONS OF THE PATTERN OF EXPRESSION OF OXYGEN-BINDING HEMOPROTEINS IN ANTARCTIC NOTOTHENIOID FISHES

PHYSIOLOGICAL AND EVOLUTIONARY IMIPLICATIONS OF THE PATTERN OF EXPRESSION OF OXYGEN-BINDING HEMOPROTEINS

IN ANTARCTIC NOTOTHENIOID FISHES

By Kimberly Borley

Thesis Advisor:  Dr. Bruce Sidell

A Lay Abstract of the Dissertation Presented

in Partial Fulfillment of the Requirements for the

Degree of Doctor of Philosophy

(in Biochemistry and Molecular Biology)

May, 2010

Antarctic icefish do not express the oxygen-binding protein hemoglobin (Hb).  The icefish cardiovascular system is characterized by more blood vessels, larger hearts, and a larger blood volume compared to Hb-expressing Antarctic fishes.  In addition to delivering oxygen to tissues, Hb is known to degrade nitric oxide (NO), a small biological signaling molecule.  I tested the hypothesis that the absence of Hb in icefishes results in an increase in NO, which stimulates angiogenesis, the process responsible for new blood vessel growth.  I found icefish plasma contains a higher concentration of NO breakdown products compared to Hb-expressing fishes indicating that NO is more abundant in icefishes.  The expression of several genes involved in angiogenesis was not found to differ between adult fish that express Hb and those that do not, indicating that, while NO levels are higher in icefish, angiogenesis is not active in the adult animal.  To further investigate whether loss of Hb directly can increase NO and stimulate angiogenesis, Hb-expressing fishwere treated with phenylhydrazine HCl, a chemical that ruptures red blood cells.  After 10 days of treatment with phenylhydrazine, anemic fish were found to have higher levels of NO and an increased expression of angiogenesis genes compared to control animals, suggesting a causative relationship between loss of Hb and induction of angiogenesis, which may be active during early development stages of icefishes. 

            Several species of icefish have also lost the expression of myoglobin (Mb), an oxygen-binding protein expressed in the heart of most icefish species.  Previous studies indicate Mb expression was lost on four separate occasions during the evolution of icefishes.  The mutations resulting in the loss of Mb have been elucidated for all but one species, D. hunteri.  I sequenced the Mb gene from D. hunteri and identified a mutation that was previously proposed as the mechanism responsible for the loss of Mb in C. aceratus.  Sequencing of Mb from all 16 species of icefish demonstrated the same mutation is also present in many species that express Mb suggesting another mechanism may be responsible for the loss of Mb expression in C. aceratus and D. hunteri.