Meeting Abstract
Our research group recently cloned and characterized a putative membrane androgen receptor from teleost ovarian tissue that is homologous to the zinc transporter protein ZIP9 (Slc39a9). Since the discovery of its androgen receptor activity, ZIP9 has been found to mediate androgen actions in a number of cell culture models from various tissues. However, ZIP9 has not been examined in an in vivo model so the precise physiological functions of this receptor remain unclear. A ZIP9-mutant strain of zebrafish was developed using a CRISPR-Cas9 system to examine the role of the protein in teleost reproduction. Mutant females have reduced fecundity and spawn significantly fewer eggs than wild-type fish. ZIP9-mutant females also produce a high proportion of eggs that do not undergo chorion elevation, a characteristic of normal egg activation. Eggs that show this phenotype have low fertilization rates and produce larvae that exhibit a high incidence of pericardial/yolk sac edema and reduced growth compared to larvae hatched from wild-type eggs. Zinc detection using fluorescent probes indicated that in wild-type eggs, zinc is localized to intracellular vesicles prior to activation, but once activation occurs the number of zinc containing vesicles decreases and a rise in extracellular zinc is detected. This suggests that zinc is released during activation in fish eggs similar to observed in mammalian eggs. ZIP9-mutant eggs that show the abnormal activation phenotype also show abnormal zinc vesicle morphology in that the vesicles are significantly smaller than those of wild-type eggs. Thus, the potential disruption of zinc regulation during egg activation and/or maturation in ZIP9-mutant fish may account for the abnormal activation phenotype and the reduction in viable offspring produced by mutant fish.
