Figure 1.

Expression and [³⁵SO₄] sulfate labeling of PRPg2 alone and along with basic PRP in AtT-20 cells. (A) Western blot of PRPg2 secreted from cells expressing PRPg2 alone or coexpressing PRPg2 and basic PRP (bPRP). The blot was probed with a mixture of antibodies recognizing PRPg2 and basic PRP followed by biotin-conjugated goat anti-rabbit IgG and ¹²⁵I-labeled strepavidin. The PRPg2 core (∗) is easily visible, whereas the modified proteoglycan is more diffuse and not easily visible at this exposure. Two forms of basic PRP are detected: the major one (33,000) is glycosylated and the minor one (31,000) is nonglycosylated. The level of expression of PRPg2 is 10% of that of endogenous ACTH except for the clone in lane 2, for which the level of expression of PRPg2 is 6% of that of ACTH. The level of expression of basic PRP is 10, 20, and 115% of that of ACTH. (B) PRPg2 from clones shown in A was labeled with [³⁵S]sulfate for 3 h, and equal amounts of product were immunoprecipitated and analyzed by SDS-PAGE and fluorography. [³⁵S]sulfate incorporation into PRPg2 was quantitated by phosphorimager analysis and the amount relative to that observed when PRPg2 is expressed alone is indicated beneath each lane.