B.Th. Walther

Dept. of Molecular Biology, Univ. of Bergen, N-5020 Bergen, NORWAY

Female sexual maturation proceeds by pivotal hepatic biosynthetic processes under the control of estrogens. In fish, oogenesis involves both zonagenesis and vitellogenesis, forming the bulk of the eggshell (zona) and yolk, respectievely (Walther 1993). Estradiol induces mRNAs for zona radiata proteins (zrp; Oppen-Berntsen et al. 1990) and for vitellogenin (vtg), by interacting with hepatic estrogen receptor, followed by protein synthesis.
Such inductions also occur in primary cultures of hepatocytes from juvenile Atlantic salmon, which secrete zrp and vtg into the culture medium (Oppen-Berntsen et al. 1992). In vivo, zrp and vtg are transported in the blood for specific uptake by the ovaries (Oppen-Berntsen et al. 1994). Recent evidence shows that zonagenesis occurs in response to lower levels of estradiol than are required to initiate vitellogenesis in juvenile Atlantic salmon (Celius & Walther 1998). Zonagenesis may be initiated already at levels approximating those sufficient for induction of hepatic estrogen receptor (ER). Furthermore, at high levels of estradiol, vitellogenesis appears to be somewhat delayed compared to zonagenesis. The finding of zonagenesis induced at putatively constitutive ER-levels, points to initiation of puberty by non-classical steroid signal pathways (Celius & Walther, unpubl.).
In analogy to Sumpter & Jobling (1995), we used in vitro induction of zonagenesis to assess the estrogenic potential by xenobiotics and mycotoxins (Celius et al. 1999). Zonagenesis provides an alternate and supplementary assay for rapid assessment of xenoestrogenicity, as shown in vivo for xenobiotics such as nonylphenol (Arukwe et al. 1997). Nonylphenol, lindane, bisphenol A, and DDT, as well as zearalenone, and both isomers of zearalenol, all induced zonagenesis in vitro, albeit with various potencies. The relative activities of the zearalenol isomers compared to their in vivo activity. Results from this assay may rationalize how various xenobiotics, despite their highly disparate molecular structures (e.g. lindane & DDT), may all exhibit the biological property of estrogenicity.
Finally, the reproductive relevance of zonagenesis is discussed in the context of the evolution of vertebrate gestation, and of sexual reproduction (Walther 1993; 1998). Zonagenesis may protect vertebrate gestation in fish, reptiles and mammals, but zrp-deposition reflects the evolution of the female reproductive tract (Walther 1999). If zonagenesis exists in all vertebrates, it may be a common biomarker for xenoestrogenic action in vertebrates. Since zonagenesis was thought to be more limited than vitellogenesis, this conclusion is surprising.

References
Arukwe, A. et al. (1997) Environ. Health Perspectives 105: 2 - 5.
Celius, T. & Walther, BT (1998) J. Endocrinology 158: 258 - 266.
Celius, T. et al. (1999) Environ. Health Perspectives 107: 63 - 68.
Oppen-Berntsen, DO et al. (1990) Devel. Biology 137: 258 - 265.
Oppen-Berntsen, DO et al. (1992) J. Endocrinology 135: 293 - 302.
Oppen-Berntsen, DO et al. (1994) J. Experimental Zoology 268: 59 - 70.
Sumpter, JP & Jobling, S. (1995) Environ. Health Perspectives 103 (suppl.): 173 - 178.
Walther, BT (1993) Physiol. Biochem. of Fish Larvae, pp.2-21. U. of Bergen.
Walther, BT (1998) Proc. EMBO Reproduction Workshop. p.14, U. of Bergen.
Walther. BT (1999) Sarsia (in press).

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