Membrane-bound progesterone receptors coupled to G proteins in the fungus Rhizopus nigricans

Abstract

Steroid binding sites with high affinity for progesterone (K d = 40 þ 14 nM determined by binding, and K d = 71 þ 22 nM determined by displacement studies) and lower affinity for 21-hydroxyprogesterone and for testosterone, but no affinity for estradiol-17L, onapristone and K-naphthoflavone were detected in the enriched plasma membrane fraction of the fungus Rhizopus nigricans. The amount of steroid binding sites is in accordance with the value of B max = 744 þ 151 fmol (mg protein)31 . In the membrane fraction, progesterone induced about 30% activation of G proteins over basal level, as determined by GTPase activity (EC 50 = 32 þ 8 nM) and by the guanosine 5P-O-(3-thiotriphosphate) (GTPQS) binding rate (EC 50 = 61 þ 21 nM). The affinity of receptors for progesterone was substantially decreased in the presence of GTPQS and of cholera toxin. Our results suggest the existence of progesterone receptors in the membrane of Rhizopus nigricans and their coupling to G proteins. ß

Identification and partial characterization of Rhizopus nigricans Gβ proteins and their expression in the presence of progesterone

“…We examined progesterone effects in R. nigricans at the plasma membrane level, where progesterone receptors have been shown to be coupled to G proteins [19], and the signal further transmitted to cAMP [9]. Since the ␣ subunits of the stimulatory (Gs) and inhibitory (Gi) types of G proteins do not appear to be involved in the response of R. nigricans to progesterone [9], we anticipated that the G protein ␤␥ subunits play an important role in regulating the signalling by progesterone.…”

Specific interactions of steroids, arylhydrocarbons and flavonoids with progesterone receptors from the cytosol of the fungus Rhizopus nigricans

“…The mechanism of enzyme induction by progesterone has not yet been elucidated, although some similarities to steroid action in higher eucaryotic organisms were observed. In our preliminary reports, we detected progesterone binding molecules in the R. nigricans plasma membrane fraction [11] and in the cytosol [12]. No biological role of membrane receptors in progesterone signalling has been detected; we were not able either to confirm or to exclude the participation of cytosolic receptors in the induction of progesterone hydroxylases [12].…”

“…nigricans responds to xenobiotic progesterone from the surroundings by inducing progesterone-hydroxylating enzymes containing cytochrome P450 [4,10]. During our investigation of progesterone signalling in the fungus two different routes were proposed, one initiated by progesterone binding to receptors at the fungal plasma membrane [11] and the other involving intracellular progesterone receptors [12]. Our previous studies were focused on the kinetic characteristics of progesterone receptors, resulting in identification of a single type of membrane receptor with a K d of ≈50 nM [11] and two types of cytosolic receptors with K d of ≈5.0 nM and ≈30.0 nM [12].…”

Possible involvement of G-proteins and cAMP in the induction of progesterone hydroxylating enzyme system in the vascular wilt fungus Fusarium oxysporum

“…In R. nigricans, evidence has been obtained for direct binding of progesterone to a G-proteincoupled membrane receptor [8]. If a similar mechanism is operating in F. oxysporum, the G␤ subunit Fgb1 could be part of the receptor signaling complex.…”

“…Specific G-protein-coupled progesterone receptors have been detected in plasma membrane fractions [8] as well as in the cytosol [9]. Heterotrimeric G-protein ␤ subunits and cyclic adenosine monophosphate (cAMP) have also been shown to play a key role in the mechanism of enzyme induction by progesterone [6].…”