The progesterone receptor‐like properies of Xenopus laevis oocyte melanosomes are probably due to eumelanin

Abstract

The binding of progesterone to t h e melanosomes of t h e Xenopus laeuis oocyte was examined. Based upon the similarities between the eumelanin complex, which is prepared by stripping melanosomes of their proteins, and synthetic melanin in regard to their association with progesterone and other steroids, the data strongly suggest that eumelanin rather than a protein is responsible for the receptor-like properties of the melanosomes.It is generally assumed that steroid-responsive cells contain a receptor that interacts with the steroid to form a n active hormone-receptor complex. This complex is thought to initiate the various biochemical and cellular responses characteristic of the target cell. Since describing the mechanism of action of steroid hormones partially depends upon evidence to validate or refute this concept, there has been an intensive search for receptors for several years. The most common approach has been to isolate cellular fractions and look for macromolecules that have receptor-like binding properties, namely, specificity for the hormone with both saturability and an affinity within the range of physiological hormone concentrations (Wira et al., '71). Putative steroid receptors, all of which are proteins, have, indeed, been demonstrated in many cells and tissues known to respond, respectively, to estrogens, progestins, androgens, glucocorticosteroids, mineralocorticosteroids, or vitamin D (Mainwaring, '75; Baulieu et al., '75).Although none of these hormone-protein complexes have been shown directly to mediate the hormonal effect, these findings have encouraged investigators to use the binding assay to examine other steroid responsive cells for putative receptors. The amphibian oocyte is a case in point.The Stage 6Xenopus laevis oocyte (Dumont, '72) undergoes germinal vesicle breakdown and matures to metaphase I1 after exposure to progesterone (Smith, '75; Wasserman and Smith, '78). Any progesterone receptor is supposedly located at or near the surface of the cell since it has been shown that externally ( 1 ) that the induction of oocyte maturation proceeds by different mechanisms in albino versus pigmented oocytes; ( 2 ) that a putative receptor, perhaps analogous to proteins naturally associated with melanosomes in the pigmented oocyte, such as tyrosinase, are present in the albino oocyte but in another compartment (Wyllie and DeRobertis, '76) ; (3) that a putative receptor is artifactually associated with some component of melanosomes; and (4) that the receptor-like properties of melanosomes are due to the pigment (eumelanin) itself rather than to an associated protein.In order to approach these, and perhaps other explanations, we began studies designed to determine whether proteins or eumelanin ' Supported by NIH Grant HD-04229. 375

A marked animal–vegetal polarity in the localization of Na+,K+‐ATPase activity and its down‐regulation following progesterone‐induced maturation

“…We have shown the Na + ,K + ‐ATPase is predominantly localized over the animal hemisphere. Similarly, the following are also mainly expressed in the animal hemisphere: GABA receptor (Chapell et al, 1998), muscarinic acetylcholine receptors, thyrotropin releasing hormone receptors (Kusano et al, 1982; Oron et al, 1988), progesterone receptors (Coffman et al, 1979; Smith, 1989), reduced folate carrier (hRFC) (Subramanian et al, 2001), endogenous G‐proteins and other signaling components (Oron et al, 1988), Ca 2+ ‐sensitive Cl‐channels (Robinson, 1979), U‐cadherin (Muller et al, 1992), and β1‐integrin (Muller et al, 1993). Kir (2.1) potassium channels (Wild and Paysan, 2002), apical amiloride‐sensitive epithelial Na + channels (Mastroberardino et al, 1998), and nicotinic acetylcholine receptors (Barnard et al, 1992), on the other hand, are targeted to the vegetal hemisphere.…”

High-Affinity Binding of Progesterone to the Plasma Membrane of Xenopus Oocytes: Characteristics of Binding and Hormonal and Developmental Control1

“…In membrane samples prepared from pigmented frogs, the melanin contamination could not be totally eliminated. Because progesterone can bind to melanin [14], Scatchard analyses of the samples prepared from albino frog ovaries were performed to determine whether receptor binding activity could also be found in melanin-free preparations. Similar results were obtained in these preparations (Table 1), indicating that melanin did not interfere with the progesterone binding activity in the regular samples.…”

“…This is relevant to note, since it has been reported that progesterone can bind to melanin [14]. It is also important to mention that intracellular (cytosolic and organelle) binding sites for progesterone in the amphibian oocyte have been previously ruled out [8,18].…”

“…However, oocytes of the albino mutant of X. laevis, which completely lack melanosomes as well as premelanosomes, can be induced to mature by progesterone [13]. Also, the melanosome component that binds progesterone is eumelanin rather than a protein [14]. Thus, the melanosome "receptor" seems to play no physiological role in oocyte maturation [13].…”