Environmental
endocrine disruptors (EEDs) that affect
androgen or
estrogen activity may disrupt gene regulation during phallus development to cause
hypospadias or a masculinized clitoris. We treated developing male tammar wallabies with
estrogen and females with
androgen from day 20-40 postpartum (pp) during the
androgen imprinting window of sensitivity.
Estrogen inhibited phallus elongation but had no effect on urethral closure and did not significantly depress testicular
androgen synthesis.
Androgen treatment in females did not promote phallus elongation but initiated urethral closure. Phalluses were collected for transcriptome sequencing at day 50 pp when they first become sexually dimorphic to examine changes in two signaling pathways, sonic hedgehog (SHH) and wingless-type MMTV integration site family (WNT)/β-
catenin. SHH
mRNA and β-
catenin were predominantly expressed in the urethral epithelium in the tammar phallus, as in eutherian mammals.
Estrogen treatment and
castration of males induced an upregulation of SHH, while
androgen treatment downregulated SHH. These effects appear to be direct since we detected putative
estrogen receptor α (ERα) and
androgen receptor (AR) binding sites near SHH. WNT5A, like SHH, was downregulated by
androgen, while WNT4 was upregulated in female phalluses after
androgen treatment. After
estrogen treatment, WIF1 and WNT7A were both downregulated in male phalluses. After
castration, WNT9A was upregulated. These results suggest that SHH and WNT pathways are regulated by both
estrogen and
androgen to direct the proliferation and elongation of the phallus during differentiation. Their response to exogenous
hormones makes these genes potential targets of EEDs in the etiology of abnormal phallus development including
hypospadias.