Calnuc does not affect constitutive secretion of GAG chains or basal secretion of gACTH and its precursors. (A) Overexpressed calnuc does not affect constitutive secretion of glycosaminoglycan (GAG) chains from either parental AtT-20 cells (NT) or those stably transfected with calnuc-GFP over the time period indicated (15 min to 2 h). ~85% of the GAG chains were secreted from both cell types at 2 h. Results (mean ± SD) represent the average of values obtained in 3 separate experiments. Cells were pretreated with 0.5 mM xyloside and subsequently pulse-labeled with [35S]sulfate for 5 min. Secreted GAG chains were precipitated with cetylpyridinium chloride. Samples collected by vacuum filtration were counted by liquid scintillation. (B) Comparing to parental cells, in those overexpressing calnuc, no significant difference of gACTH and POMC + intermediate is observed both extracellularly (Lanes 1 and 2) and intracellularly (Lnaes 3 and 4). However, relatively increased secreted (Lane 2) and intracellular (Lane 4) mature ACTH are shown when calnuc is overexpressed. Med: medium; Cell: intracellular.