Expression of thromboxane A2 receptor gene and thromboxane A2 synthase in bovine corpora lutea.
Studies were undertaken to investigate the expression of thromboxane (TXA2) receptor gene, from mRNA to functional receptor protein in terms of ligand binding, along with the cellular and subcellular distribution of the enzyme that catalyzes the formation of the ligand for the receptors. Bovine corpora lutea contained a single TXA2 receptor mRNA transcript of 2.8 kb. All the cell types in bovine corpora lutea contained immunoreactive TXA2 synthase, TXB2, TXA2 receptor transcripts, and receptor protein that bound the TXA2 antagonist 9,11-dimethylmethano-11,12-methano-16 (3-iodo-4-hydroxyphenyl)-13-14-dihydro-13-aza-15 alpha beta-omega-tetranor TXA2. The large luteal cells (20-35 microns) contained more receptor transcripts, receptor protein, and immunoreactive TXA2 synthase than did the small luteal cells (12-19 microns), luteal blood vessels, and nonluteal cells (7-12 microns). After correction for the cellular area differences, small luteal cells were seen to contain more receptor protein than did large luteal cells and nonluteal cells. All the cells showed an increase of TXA2 receptors and catalytically active TXA2 synthase from mid-luteal phase to early pregnancy, suggesting the possibility that TXA2 could be a luteotropic eicosanoid. Bovine lung homogenates (a positive control), bovine luteal plasma membranes-mitochondria-lysosomes fraction, rough-smooth endoplasmic reticulum-Golgi fraction, and highly purified nuclei contained 65-kDa immunoreactive protein, presumably representing TXA2 synthase. In addition, the luteal fractions, but not bovine lung, contained other small and large molecular-size immunoreactive proteins. Immunogold electron microscopy showed that immunoreactive TXA2 synthase was present primarily in plasma membranes, rough endoplasmic reticulum, nuclear membranes, and chromatin; and immunoreactive TXB2 was present primarily in different-size vesicles and nuclear chromatin. In summary, the present studies demonstrate for the first time that primarily small and large luteal cells and secondarily blood vessels and nonluteal cells in bovine corpora lutea express TXA2 receptor gene along with the functional TXA2 synthase. The presence of functional enzyme in luteal cell nuclei suggests that the enzyme and/or its product may have previously unrecognized functions in nuclei.
Lei
Z M
ZM
Department of Obstetrics and Gynecology, University of Louisville, School of Medicine, Kentucky 40292.
Rao
C V
CV
Chakraborty
C
C
D000818:Animals; D001345:Autoradiography; D015152:Blotting, Northern; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure; D003338:Corpus Luteum / Q000166:cytology / Q000201:enzymology* / Q000648:ultrastructure*; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000648:ultrastructure; D005260:Female; D015870:Gene Expression; D006056:Golgi Apparatus / Q000201:enzymology / Q000648:ultrastructure; D007150:Immunohistochemistry; D008168:Lung / Q000166:cytology / Q000201:enzymology / Q000648:ultrastructure; D008247:Lysosomes / Q000201:enzymology / Q000648:ultrastructure; D008854:Microscopy, Electron; D008928:Mitochondria / Q000201:enzymology / Q000648:ultrastructure; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D017482:Receptors, Thromboxane / Q000032:analysis* / Q000235:genetics*; D013929:Thromboxane B2 / Q000032:analysis; D013930:Thromboxane-A Synthase / Q000032:analysis*; D014158:Transcription, Genetic
D012333:RNA, Messenger; D017482:Receptors, Thromboxane; D013929:Thromboxane B2; D013930:Thromboxane-A Synthase