pThe SPOC1 cell, a novel goblet cell line derived from rat trachea, was tested for its ability to exhibit regulated mucin secretion in response to purinergic (Psub2/sub) agonists. High-molecular-mass glycoconjugates (HMMGs) purified by CsCl-density-gradient centrifugation had a buoyant density of 1.45 g/ml. The purified HMMG material exhibited a single major band with an apparent molecular mass of greater than 1000 kDa in SDS/ polyacrylamide gels stained with silver or blotted and stained with soya-bean agglutinin. [sup3/supH]HMMG was resistant to proteoglycan-degrading enzymes, but was susceptible to neuraminidase. The HMMG was approx. 91% carbohydrate by weight, and the glycosides were O-linked. The HMMG amino acid composition was enriched in Ser and Thr (sum 27%). Thus SPOC1-cell HMMG possess the characteristics of mucin. Mucin secretion by SPOC1 cells, grown on permeable supports and perfused luminally, was stimulated by ATP, UTP and adenosine 5′-[γ-thio]triphosphate (100 μM) 4–5-fold over a baseline of 4 ng/min. The three dose–effect relations were nearly identical (iK/isub0.5/sub ~4 μM). SPOC1 cells grown on plastic and rat tracheal epithelial primary cells responded similarly to ATP and/or UTP. SPOC1 cells failed to respond to other purinergic agonists, either luminally or serosally, and consequently seem to possess an apical membrane Psub2u/sub purinoceptor. SPOC1-cell total RNA was probed for Psub2u/sub purinoceptor mRNA. Using conserved primers for both reverse transcriptase and PCR, a single band of the predicted size was observed, which had a nucleotide base sequence identical with the rat Psub2u/sub purinoceptor mRNA. Thus SPOC1 cells secrete mucin under the control of a Psub2u/sub purinoceptor; they should prove useful in dissecting the associated cellular regulatory pathways./p
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