Secretion of a putative metalloproteinase inhibitor by ovine granulosa cells and luteal tissue.
Ovine granulosa cells respond, through transcriptional mechanisms, to preovulatory concentrations of gonadotrophins by secreting an Mr 30,000 polypeptide. To determine the stages of the oestrous cycle at which this polypeptide is secreted, corpora lutea were collected on Days 3, 7, 10, 13 and 16 (Day 0 = oestrus; n = 4 per group), cut into 1-mm slices, and incubated for 6-7 h with [35S]methionine. Radiolabelled polypeptides of intra- and extra-cellular origin were separated by polyacrylamide gel electrophoresis and quantitated by densitometry. The Mr 30,000 polypeptide was secreted at all stages of the luteal phase tested (Days 3-16), and represented approximately 24% of the total labelled polypeptide present in the medium; polypeptides of approximate Mr 14,000, 25,000 and 46,000 accounted for most of the other secreted proteins. Neither pituitary hormones (LH, FSH, prolactin) nor cholera toxin (chosen to activate adenylate cyclase) affected the rate of production of Mr 30,000 polypeptide, indicating that, once secretion has been initiated in the granulosa cells, it is not readily modulated by hormonal intervention after luteinization. Incubation of luteinized granulosa cells with tunicamycin (inhibits N-linked glycosylation reactions) showed that the secreted polypeptide consists of a heavily glycosylated amino acid backbone of approximately Mr 20,000. Western blot analysis established further that the polypeptide was not an inhibin subunit. However, NH2-terminal amino acid sequencing of the first 25 amino acids revealed a 68% sequence identity between the secreted polypeptide (Mr 30,000) and a human tissue inhibitor of metalloproteinases.
Smith
M F
MF
Department of Animal Sciences, University of Missouri, Columbia 65211.
Moor
R M
RM
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D003338:Corpus Luteum / Q000187:drug effects / Q000378:metabolism*; D000242:Cyclic AMP / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004971:Estrus / Q000502:physiology; D005260:Female; D006065:Gonadotropins, Pituitary / Q000494:pharmacology; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D007265:Inhibins / Q000032:analysis; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors*; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D012756:Sheep / Q000502:physiology*; D014415:Tunicamycin / Q000494:pharmacology
D006065:Gonadotropins, Pituitary; D010455:Peptides; D014415:Tunicamycin; D007265:Inhibins; D000242:Cyclic AMP; D008666:Metalloendopeptidases