We have established a system, the CA77 rat medullary thyroid carcinoma cell line, for studying the products of somatostatin (SS) gene expression. Based on the amino acid sequence of proSS, we developed a RIA for the amino terminus of proSS (proSS-NTP) and demonstrated in acidic cell extracts two major proSS-NTP-containing species of 8000 and 4000 daltons. Studies were then performed on species secreted into culture medium. Serial dilutions of culture medium showed tracer displacement curves parallel to serial dilutions of synthetic proSS-NTP standard. Analysis by gel filtration chromatography of 24-h culture medium showed the major proSS-NTPcontaining species to have an estimated mol wt of 8000 daltons. No 4000-dalton species was observed. The acute effects of calcium and glucagon, known secretagogues of SS, on secretion of immunoreactive (i) proSS-NTP were investigated in 3-h experiments. Basal (0.5 mM calcium) secretory rates (mean ± SE) of iproSS-NTP and iSS were 1.29 ± 0.36 and 7.38 ± 1.51 ng/mg acid-extractable protein, respectively. High calcium (3 mM) stimulated iproSS-NTP and iSS secretion 302 ± 100% and 363 ± 105%, respectively. High calcium plus 10−6 M glucagon also stimulated secretion of iproSSNTP and iSS in a coordinate fashion. Analyses by gel filtration chromatography of 3-h culture medium revealed that high calcium markedly increased the 8000-dalton proSS-NTP-containing species. No 4000-dalton species was observed. The absence of 4000-dalton proSS-NTP species in 24-h culture medium, the lack of degradation of 4000-dalton proSS-NTP (recovered from CA77 cell extracts) added to tissue culture medium, and the selective secretion of the 8000-dalton proSSNTP species under both basal and stimulated conditions coordinate with the secretion of SS indicate that the 4000-dalton proSS-NTP-containing species is not secreted.
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