MAINTAINED PC1 AND PC2 EXPRESSION IN THE ATT-20 VARIANT CELL-LINE 6T3 LACKING REGULATED SECRETION AND POMC - RESTORED POMC EXPRESSION AND REGULATED SECRETION AFTER CAMP TREATMENT

被引:23
作者
DAY, R
BENJANNET, S
MATSUUCHI, L
KELLY, RB
MARCINKIEWICZ, M
CHRETIEN, M
SEIDAH, NG
机构
[1] CLIN RES INST MONTREAL,JA DESEVE LABS MOLEC NEUROENDOCRINOL,MONTREAL,PQ H2W 1R7,CANADA
[2] UNIV BRITISH COLUMBIA,DEPT ZOOL,VANCOUVER,BC V6T 1Z4,CANADA
[3] UNIV CALIF SAN FRANCISCO,HORMONE RES INST,SAN FRANCISCO,CA 94143
关键词
D O I
10.1089/dna.1995.14.175
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Two variant cell lines were recently established from parent AtT-20 cells. Whereas HYA.15.10.T.2 have a reduced level of secretory granules, HYA.15.6.T.3 are completely devoid of both the regulated pathway of secretion and of dense-core secretory granules. AtT-20 cells normally express the processing enzymes PC1, PC2, furin, carboxypeptidase E, and peptidylglycine alpha-amidating monooxygenase, as well as proopiomelanocortin, chromogranin B, and 7B2. We measured the expression of these mRNAs in both variant cell lines. Although some differences in mRNA level were noted, HYA.15.10.T.2 and HYA.15.6.T.3 cell lines maintained their expression of the processing enzymes and of 7B2. Furthermore, PC1 and PC2 were shown to be functionally active in the HYA.15.6.T.3 cells. In contrast, proopiomelanocortin and chromogranin B mRNA levels were no longer detectable in HYA.15.6.T.3 cells. Interestingly, stimulation of the HYA.15.6.T.3 cells with cAMP restored proopiomelanocortin mRNA, beta-endorphin immunoreactivity, acid dense-core granules. Furthermore, at the ultrastructural level, beta-lipotropin immunoreactivity was detected in granules of cAMP-induced HYA.15.6.T.3 cells. Finally, depolarization of cAMP-induced HYA.15.6.T.3 cells with 56 mM potassium chloride resulted in a marked increase in the release of beta-endorphin immunoreactivity. These observations demonstrate that cAMP restores the regulated pathway of secretion in HYA.15.6.T.3 cells, which under untreated conditions do not demonstrate regulated release. These variant cell lines are unique models to understand better the relationship of the regulated pathway and the expression of the processing enzymes.
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页码:175 / 188
页数:14
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