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1: Zoolog Sci. 2005 Sep;22(9):1045-50.

Molecular cloning and expression in gonad of Rana rugosa WT1 and Fgf9.

Yamamura Y, Aoyama S, Oshima Y, Kato T, Osawa N, Nakamura M.

Department of Biology, Faculty of Education and Integrated Arts and Sciences,
Waseda University, Tokyo, Japan.

Sry (sex-determining region on the Y chromosome) is required for testicular
differentiation in mammals. In addition to Sry, other genes such as WT1, Fgf9,
Dax1, Dmrt1 and Sox9 are widely accepted to be involved in the sex determination 
in vertebrates. However, the roles of these genes during sex determination still 
remain unclear in amphibians. This study was undertaken to examine the expression
of WT1 and Fgf9 in the developing gonad of amphibians. We first isolated the WT1 
cDNA from the frog Rana rugosa. Like WT1 in mice, R. rugosa WT1 showed 2
isoforms; i.e., one had an additional 3 amino acids, KTS, included between the
third and fourth zinc fingers. However, 17 amino acids in exon 5 of mammalian WT1
could not be found in R. rugosa WT1, which is also the case in turtle and
chicken. The mRNA of both isoforms (+KTS, -KTS) was detected in the lung, kidney 
and testis, but not in the ovary and muscle of adult frogs. The 2 isoforms were
expressed first in the embryos at stage 23. Thereafter, the expressions remained 
constant in the gonad attached to mesonephros of both sexes during sex
determination. We next isolated the R. rugosa Fgf9 cDNA encoding 208 amino acids.
The amino acid sequence of Fgf9 had similarity greater than 92% with chicken,
mouse and human Fgf9s, suggesting that Fgf9 is highly conserved among vertebrate 
classes. Fgf9 was expressed in the ovary of an adult frog strongly, but in the
lung weakly. In contrast, the Fgf9 mRNA was hardly detected in the kidney, testis
and muscle. Moreover, Fgf9 did not show a sexually dimorphic expression pattern
during sex determination in R. rugosa. The results, taken together, suggest that 
both WT1 and Fgf9 are expressed in the indifferent gonad prior to sex
determination without any difference in the expression between males and females.
Thus, it seems unlikely that they are a key factor to initiate the divergence
leading to testicular or ovarian differentiation in R. rugosa.

PMID: 16219985 [PubMed - indexed for MEDLINE]

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