Identification, cellular and subcellular distribution of 21 and 72 kDa proteins (tuberalins?) secreted by specific cells of the pars tuberalis

Abstract

The cell types of the pars tuberalis (PT) are the follicular cells, the pars distalis cells and the so-called PT-specific cells. The latter are distinct endocrine cells displaying melatonin receptors. Although the nature of the secretory product(s) of the PT-specific cells has not yet been clarified, the function of these cells has started to be unfolded. For practical reasons, previous authors have designated the, as yet, unidentified PT hormone(s) as tuberalin(s). PT-specific cells synthesise the common alpha subunit of the pars distalis glycoprotein hormones, and it has been suggested that tuberalin would correspond to the beta chain of a specific glycoprotein secreted by these cells. The aims of the present investigation were to identify the compounds secreted by the specific cells of bovine PT, and to establish their cellular and subcellular distribution. For this purpose, proteins secreted into the culture medium of PT explants were separated by electrophoresis and used to raise antibodies. Two of these proteins, with an apparent molecular mass of 21 and 72, generated antibodies (Ab-21 and Ab-72) that differentially immunoreacted with PT-specific cells. These two antibodies were used for immunoblotting of conditioned medium and of PT explants, and for light and electron microscopy immunocytochemistry. In immunoblots, Ab-21 reacted with compounds of 21, 22, 47 and 52 kDa, whereas Ab-72 revealed a compound of 72 kDa only. Ab-72 immunoreactive material corresponded to a protein, here designated as tuberalin I, secreted by a small population of PT-specific cells (type 2 cells), and stored in 140 nm secretory granules. Immunoreactive tuberalin I was missing from bovine pars distalis and from rat PT. The predominant population of PT-specific cells (type 3 cells) secreted and stored, within 280 nm secretory granules, an Ab-21 immunoreactive protein, here designated as tuberalin II. All cells of rat PT immunoreacted with Ab-21. In the cells of bovine and rat PT, immunoreactive tuberalin II was mostly confined to a paranuclear spot; this spot also bound wheat germ agglutinin and reacted with an antibody against the alpha chain of glycoprotein pars distalis hormones. It is suggested that tuberalin II would correspond to the beta chain of a specific glycoprotein secreted by type 3 PT-specific cells. In bovine PT, the cells displaying immunoreactive tuberalins I and II did not react with any of the antibodies against pars distalis hormones.