Heterologous processing of rat prosomatostatin to somatostatin-14 by PC2: Requirement for secretory cell but not the secretion granule

The role of PC2 in prosomatostatin (PSS) processing was investigated in GH₃/GH₄C₁ pituitary cells. These cells are sparsely granulated, express different amounts of PC2 and no PC1. We describe heterologous processing of rat PSS (rPSS) co-expressed with PC2 in stably transfected cells, correlate PC2 protein levels under different conditions of transfection with efficiency of PSS processing to somatostatin-14 (SS-14), determine the effect of modulating cell granularity on enzyme expression and PSS processing, and compare the relative potency of PC2 with that of PC1. PSS and cleavage products were monitored by HPLC and radioimmunoassay of SS-like immunoreactivity (SSLI). Radioimmunoassay analysis of N-terminal PC2-like immunoreactivity (PC2 LI) in GH₄C₁:rPSS, GH₄C₁:rPSS + PC2 and GH₃:rPSS transfectants showed a gradient of PC2 protein of 1:2.6:3.4 in cell extracts and 1:4.7:9 in secretion media from these cells respectively. The concentration of PC2 protein correlated with SS-14 conversion efficiency was 36 ± 3% in GH₄C₁:rPSS cells, 56 ± 7% in GH₄C₁:rPSS-PC2 cells and 100%, in GH₃:rPSS cells. Treatment of GH₄C₁:rPSS + PC2 cells with epidermal growth factor, insulin, and β-estradiol to induce granules, significantly increased basal and forskolin-stimulated co-release of SS LI and PC2 LI, but had no influence on SS-14 processing efficiency. Hormone treatment led to a small increase in the ratio of mature PC2 (68 kDa) to proPC2 (75 kDa) forms. PC1 stably transfected in GH₄C₁ cells produced significantly greater SS-14 conversion (62% in cells, 66% in media) compared with PC2 transfectants (53% in cells, 47% in media). These results provide the first proof that PC2 can effect dibasic processing of mammalian PSS, and, along with PC1, qualifies as an authentic SS-14 convertase. The activity of PC2 requires the milieu of the secretory cell but not the secretory granule.