Detection of multiple PRL- and GH-like proteins in human pituitary by Western blot analysis.
Surgically removed normal and tumorous pituitary tissues from a prolactinoma patient were analyzed by Western blot techniques for PRL and GH variants. Criteria for identification were the Rf of the bands within the gel, immunologic crossreactivity to specific antisera, and structural verification by tyrosine peptide-mapping of individual bands from the gel. The authors found the tumor-tissue to be characterized by the presence of a PRL band greater in concentration than in the normal tissue and the virtual absence of a GH band. Immunoblotting of the electrophoretically resolved proteins form both types of tissues revealed several new bands crossreactive with human PRL and GH antibodies. Some of the new bands were of Mr greater than the monomeric PRL and GH and others were of lower Mr. Relative of two of the low mobility Mr PRL-immunoreactive bands designated as 16K and 8K corresponded to the Rf of the two fragments of cleaved PRL, which suggested that cleaved PRL occurred naturally in the human pituitary gland. The most conspicuous of the new PRL-immunoreactive bands, a 25,000 Mr protein migrating slightly behind PRL, displayed strong crossreactivity to hPRL antibodies and was present in greater concentration in the prolactinoma tissue than in the normal tissue. These properties suggested that it was related to hPRL and perhaps represented its glycosylated variant. However, its tyrosine peptide map did not resemble that of hPRL. Thus, it is not clear whether it represented G-hPRL or a new pituitary protein that cross-reacts with hPRL antibodies. In addition, two other bands of low Mr, designated as unknown 1 and unknown 2, reacted with hPRL antibodies. Immunostaining with hGH antibodies revealed the 20K-hGH variant, the F1 fragment of cleaved hGH, and a pair of new bands immediately behind GH that could represent glycosylated hGH--possibly a product of Seeburg's variant hGH gene. Both PRL and GH antibodies elicited numerous bands of high Mr by the technique employed, far more than ever observed by Sephadex chromatography. The nature of the high Mr bands remains unknown. Further characterization of these new PRL- and GH-immunoreactive proteins might help in the understanding of the multiple physiologic functions of PRL and GH in man.
Sinha
Y N
YN
Ott
K
K
Vanderlaan
W P
WP
D000236:Adenoma / Q000032:analysis* / Q000473:pathology; D000328:Adult; D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D013006:Growth Hormone / Q000032:analysis*; D006801:Humans; D007158:Immunologic Techniques; D008970:Molecular Weight; D010449:Peptide Mapping; D010903:Pituitary Gland, Anterior / Q000032:analysis* / Q000473:pathology; D010911:Pituitary Neoplasms / Q000032:analysis* / Q000473:pathology; D011388:Prolactin / Q000032:analysis*
D011388:Prolactin; D013006:Growth Hormone