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

Aristea S. Galanopoulou, N. G. Seidah, Y. C. Patel

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

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.

Original languageEnglish (US)
Pages (from-to)111-118
Number of pages8
JournalBiochemical Journal
Volume311
Issue number1
StatePublished - 1995
Externally publishedYes

Fingerprint

Somatostatin
Rats
Proprotein Convertase 2
Processing
prosomatostatin
Radioimmunoassay
Colforsin
Epidermal Growth Factor
Conversion efficiency
Estradiol
Hormones
Insulin
Secretory Vesicles
Cell Extracts
Transfection
Enzymes
High Pressure Liquid Chromatography

ASJC Scopus subject areas

  • Biochemistry

Cite this

Heterologous processing of rat prosomatostatin to somatostatin-14 by PC2 : Requirement for secretory cell but not the secretion granule. / Galanopoulou, Aristea S.; Seidah, N. G.; Patel, Y. C.

In: Biochemical Journal, Vol. 311, No. 1, 1995, p. 111-118.

Research output: Contribution to journalArticle

@article{1d3e2a16915a48a185d96626f0a15f01,
title = "Heterologous processing of rat prosomatostatin to somatostatin-14 by PC2: Requirement for secretory cell but not the secretion granule",
abstract = "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.",
author = "Galanopoulou, {Aristea S.} and Seidah, {N. G.} and Patel, {Y. C.}",
year = "1995",
language = "English (US)",
volume = "311",
pages = "111--118",
journal = "Biochemical Journal",
issn = "0264-6021",
publisher = "Portland Press Ltd.",
number = "1",

}

TY - JOUR

T1 - Heterologous processing of rat prosomatostatin to somatostatin-14 by PC2

T2 - Requirement for secretory cell but not the secretion granule

AU - Galanopoulou, Aristea S.

AU - Seidah, N. G.

AU - Patel, Y. C.

PY - 1995

Y1 - 1995

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=0029102824&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029102824&partnerID=8YFLogxK

M3 - Article

C2 - 7575441

AN - SCOPUS:0029102824

VL - 311

SP - 111

EP - 118

JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - 1

ER -