Pj. Oshaughnessy et L. Murphy, CYTOCHROME-P-450 17-ALPHA-HYDROXYLASE PROTEIN AND MESSENGER-RNA IN THE TESTIS OF THE TESTICULAR FEMINIZED (TFM) MOUSE, Journal of molecular endocrinology, 11(1), 1993, pp. 77-82
The testicular feminized (Tfm) mouse lacks functional androgen recepto
rs and develops with a female external phenotype and internal testes.
The testes of these animals contain normal, or close to normal, number
s of Leydig cells but secrete very low amounts of androgen due to a la
ck of 17alpha-hydroxylase activity. To determine whether this loss of
activity is due to a lack of enzyme synthesis or a change in catalytic
activity we have examined 17alpha-hydroxylase cytochrome P-450 (P-450
(17alpha)) protein and mRNA levels in the testes of Tfm mice. Levels o
f P-450(17alpha) protein were measured by immunoblotting, while mRNA w
as measured following reverse transcription (RT) and amplification by
the polymerase chain reaction (PCR). Conditions for RT-PCR were determ
ined which allowed semiquantification of P-450(17alpha) mRNA relative
to beta-actin mRNA. In extracts of Tfm testes P-450(17alpha) protein w
as undetectable using antiserum against porcine P-450(17alpha). In con
trast, a protein of around 54 kDa was clearly detectable in extracts o
f control cryptorchid testes. Using RT-PCR, P-450(17alpha) mRNA was de
tectable in both control and Tfm testes but, expressed in terms of bet
a-actin mRNA, levels of P-450(17alpha) mRNA in control testes were 40-
fold higher than those in Tfm testes. If the total amount of RNA extra
cted from each testis is taken into account then P-450(17alpha) mRNA l
evels per testis were up to 400-fold higher in control testes. These r
esults show that the reduced level of 17alpha-hydroxylase activity in
Tfm testes is related to reduced protein synthesis. Previous results h
ave shown that androgens reduce P-450(17alpha) mRNA levels in cultured
Leydig cells. Results from this study suggest, however, that androgen
s are required to induce normal levels of P-450(17alpha) mRNA in Leydi
g cells.