The expression of transforming growth factor alpha receptor protein and its activation in chicken ovarian granulosa cells of maturing follicles

Epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alp ha) are structurally related growth factors that exert their biological act ions by binding to the same cell-surface receptor, EGF receptor. However, i n chicken cells, human EGF binds with approximately 100-fold lower affinity than human TGF-alpha in a previous study, we localized EGF/TGF-alpha recep tor immunohistochemically in the granulosa and theca of the developing foll icles of laying hens. We have also shown that TGF-alpha binds to cell-surfa ce receptors of the granulosa cells. The present study characterizes the na ture of the EGF/TGF-alpha receptor. Immunoprecipitation of receptor protein s from cultured granulosa cells with an anti-EGF receptor antibody (12E) sh ows the expression of a 170-kDa receptor protein. The expression of the rec eptor protein decreases with follicular enlargement between the F3 and F1. Incubation of the cells with [I-125]TGF-alpha followed by cross-linking wit h bis(sulphosuccinimidyl)suberate showed that TGF-alpha binds a similar (17 0 kDa) receptor protein immunoprecipitated with the 12E anti-EGF receptor a ntibody. The binding of TGF-alpha to granulosa cells caused receptor protei n oligomerization, yielding the monomeric (170 kDa) and dimeric (340 kDa) p rotein forms. Oligomerization seemed to favour the formation of the dimeric rather than the monomeric form. Culturing granulosa cells with luteinizing hormone or follicle-stimulating hormone increased the expression of both m onomer and dimer forms of the receptor proteins compared with the control. Western blotting analysis with anti-phosphotyrosine antibody revealed that the lysates of TGF-alpha-stimulated cells express phosphotyrosine-containin g receptor proteins of 170 kDa and 340 kDa. The results show that chicken g ranulosa cells express the 170-kDa EGF/TGF-alpha receptor protein, which di merizes on binding to TGF-alpha, suggesting that the receptor protein may b e involved in the signal transduction of TGF-alpha actions in the chicken g ranulosa cells. (C) 1998 Chapman & Hall.