ABSTRACT
We report on the discovery of interactions of porcine endometrial 17 beta-oestradiol dehydrogenase with actin. The 17 beta-oestradiol dehydrogenase of porcine uteri is an essentially unidirectional enzyme compounded in specialized organelles. The enzyme activity in Brij 35 extracts of the particulate fraction of epithelial cells sedimenting between 1800 and 11,000 g(av). was collected by immunoadsorption and eluted at low pH. The eluate contained three proteins of 32, 45 and 80 kDa as shown by SDS/PAGE and silver staining. They were identified by amino acid sequencing and immunotyping as oestradiol dehydrogenase (32 kDa), actin (45 kDa) and a covalent dehydrogenase-actin complex (80 kDa). Disulphides, aldimines, periodate-degradable bonds and hydrophobic interactions were excluded as linkages in the 80 kDa protein. The epsilon-(gamma-glutamyl)-lysine nature of the covalent cross-link was recognized by narrow-bore h.p.l.c. analysis of enzymic digests of electro-eluted 80 kDa material. An involvement of the actin anchor in positioning of the oestradiol dehydrogenase-containing organelles according to metabolic requirements is discussed.
Subject(s)
Actins/metabolism , Dipeptides/pharmacology , Endometrium/metabolism , Estradiol Dehydrogenases/metabolism , Amino Acid Sequence , Animals , Cells, Cultured , Collagen/metabolism , Disulfides , Endometrium/cytology , Female , Immunosorbent Techniques , Molecular Sequence Data , Peptide Mapping , SwineABSTRACT
Three hundred and twelve sera containing antibodies to smooth muscle (SMA) wer analysed for the immunofluorescence patterns they produced in various tissues. A classification is described based on the three main appearances in rat kidney. Some sera, mainly of low titre, reacted only with vessel walls (SMA-V), some stained vessels and renal glomeruli (SMA-G) and high titre sera, mainly from patients with chronic active hepatitis stained vessels, glomeruli, and intracellular fibrils in renal tubules (SMA-T). Peripheral staining in hepatocytes or thyroid cells was not a regular feature. 41/43 polyclonal SMA-T and -G sera were absorbed out completely by actin, and this also removed the pericullular staining in liver and thyroid when present. High titre SMA-V antibodies could not be absorbed by actin, and the antigen remains to be identified.
Subject(s)
Autoantibodies/classification , Muscle, Smooth/immunology , Animals , Autoantibodies/analysis , Cattle , Fluorescent Antibody Technique , Hepatitis/immunology , Humans , Kidney/immunology , Kidney Glomerulus/immunology , Kidney Tubules/immunology , Liver/immunology , Liver Diseases/immunology , Mice , Muscle Proteins/immunology , Muscles/immunology , Rats , T-Lymphocytes/immunology , Thymus Gland/immunology , Thyroid Gland/immunologyABSTRACT
Myosin in human, rat, mouse, and chicken fibroblasts was localized by indirect immunofluorescence microscopy using antibodies prepared in rabbits against highly purified chicken gizzard myosin. Filaments containing myosin span the interior of the cells and are often parallel to each other. The majority of the fibers are concentrated toward the adhesive side of the cell. Most of the myosin-containing filaments show "interruptions" or "striations." From a comparison of these fibers in fluorescence and phase microscopy and from previous results on actin-containing fibers, we conclude that at least some of the cytoplasmic myosin can be found in the actin-containing fibers, which themselves have been shown to be very similar or identical to the microfilament bundles. The occurrence of both myosin and actin in the microfilament bundles provides a basis for the motility and contractility of the cell.
