Background: Previous work has implicated isolated, control human lung mast cell granules in RNA metabolism using multiple methods of high-magnification imaging based on different mechanistic principles. These methods have demonstrated ribosomes, RNA, U1snRNP and uridine in, around and attached to secretory granules. Methods: Here, we have extended these studies using ultrastructural autoradiography of radiolabeled uridine incorporation in degranulating and recovering mast cells. Results: We found that control cells incorporated uridine into granules, with values that decreased dramatically in conjunction with stimulated histamine secretion and granule extrusion, and that granule stores of tritiated uridine increased together with the reconstitution of secretory granules in recovering mast cells. Conclusion: These findings support a possible new role for secretory granules in RNA metabolism in mast cell biology.

Keywords:
Human mast cell, Granules, RNA, Ribosomes, Uridine, Ultrastructural autoradiography, Secretion, Recovery
1.
Dvorak AM, Morgan ES, Lichtenstein LM, Weller PF, Schleimer RP: RNA is closely associated with human mast cell secretory granules, suggesting a role(s) for granules in synthetic processes. J Histochem Cytochem 2000;48:1–12.
2.
Davis L, Banker GA, Steward O: Selective dendritic transport of RNA in hippocampal neurons in culture. Nature 1987;330:477–479.
3.
Ornelles DA, Fey EG, Penman S: Cytochalasin releases mRNA from the cytoskeletal framework and inhibits protein synthesis. Mol Cell Biol 1986;6:1650–1662.
4.
Eliceiri GL: Short-lived, small RNAs in the cytoplasm of HeLa cells. Cell 1974;3:11–14.
5.
Soeiro R, Birnboim HC, Darnell JE: Rapidly labeled HeLa cell nuclear RNA. 2. Base composition and cellular localization of a heterogeneous RNA fraction. J Mol Biol 1966;19:362–372.
6.
Herman R, Weymouth L, Penman S: Heterogeneous nuclear RNA-protein fibers in chromatin-depleted nuclei. J Cell Biol 1978;78:663–674.
7.
Fakan S: Perichromatin fibrils are in situ forms of nascent transcripts. Trends Cell Biol 1994;4:86–90.
8.
Fakan S, Puvion E: The ultrastructural visualization of nucleolar and extranucleolar RNA synthesis and distribution. Int Rev Cytol 1980;65:255–299.
9.
Fakan S, Bernhard W: Localisation of rapidly and slowly labelled nuclear RNA as visualized by high resolution autoradiography. Exp Cell Res 1971;67:129–141.
10.
Fakan S, Bernhard W: Nuclear labelling after prolonged 3H-uridine incorporation as visualized by high resolution autoradiography. Exp Cell Res 1973;79:431–444.
11.
Fakan S, Puvion E, Spohr G: Localization and characterization of newly synthesized nuclear RNA in isolated rat hepatocytes. Exp Cell Res 1976;99:155–164.
12.
Zieve G, Penman S: Subnuclear particles containing a small nuclear RNA and heterogeneous nuclear RNA. J Mol Biol 1981;145:501–523.
13.
Fey EG, Krochmalnic G, Penman S: The nonchromatin substructures of the nucleus: The ribonucleoprotein (RNP)-containing and RNP-depleted matrices analyzed by sequential fractionation and resinless section electron microscopy. J Cell Biol 1986;102:1654–1665.
14.
Scherrer K, Darnell JE: Sedimentation characteristics of rapidly labelled RNA from HeLa cells. Biochem Biophys Res Commun 1962;7:486–490.
15.
Angelier N, Bouteille M, Charret R, Curgy J-J, Delain E, Fakan S, Geuskens M, Guelin M, Lacroix J-C, Laval M, Steinert G, Van Assel S: Detection of nucleic acids by means of ultrastructural autoradiography. J Microsc Biol Cell 1976;27:215–230.
16.
Amano M, LeBlond CP: Comparison of the specific activity time curves of ribonucleic acid in chromatin, nucleolus and cytoplasm. Exp Cell Res 1960;20:250–253.
17.
Warner JR, Soeiro R, Birnboim HC, Girard M, Darnell JE: Rapidly labeled HeLa cell nuclear RNA. 1. Identification by zone sedimentation of a heterogeneous fraction separate from ribosomal precursor RNA. J Mol Biol 1966;19:349–361.
18.
Gambetti P, Autilio-Gambetti L, Shafer B, Pfaff L: Quantitative autoradiographic study of labeled RNA in rabbit optic nerve after intraocular injection of [3H]uridine. J Cell Biol 1973;59:677–684.
19.
Abelson HT, Johnson LF, Penman S, Green H: Changes in RNA in relation to growth of the fibroblast. 2. The lifetime of mRNA, rRNA, and tRNA in resting and growing cells. Cell 1974;1:161–165.
20.
Liau MC, Perry RP: Ribosome precursor particles in nucleoli. J Cell Biol 1969;42:272–283.
21.
Dvorak AM: Human Mast Cells; in Beck F, Hild W, Kriz W, Ortmann R, Pauly JE, Schiebler TH (eds): Advances in Anatomy, Embryology, and Cell Biology. Berlin, Springer, 1989, vol 114.
22.
Asboe-Hansen G: Mast cells and the skin; in Helwig EB, Mostofi FK (eds): Monographs in Pathology. Baltimore, Williams & Wilkins, 1971, vol 10: The Skin, pp 83–111.
23.
Kobayasi T, Midtgård K, Asboe-Hansen G: Ultrastructure of human mast-cell granules. J Ultrastruct Res 1968;23:153–165.
24.
Schulman ES, MacGlashan DW Jr, Peters SP, Schleimer RP, Newball HH, Lichtenstein LM: Human lung mast cells: Purification and characterization. J Immunol 1982;129:2662–2667.
25.
Dvorak AM, Hammel I, Schulman ES, Peters SP, MacGlashan DW Jr, Schleimer RP, Newball HH, Pyne K, Dvorak HF, Lichtenstein LM, Galli SJ: Differences in the behavior of cytoplasmic granules and lipid bodies during human lung mast cell degranulation. J Cell Biol 1984;99:1678–1687.
26.
Dvorak AM, Schulman ES, Peters SP, MacGlashan DW Jr, Newball HH, Schleimer RP, Lichtenstein LM: Immunoglobulin E-mediated degranulation of isolated human lung mast cells. Lab Invest 1985;53:45–56.
27.
Peters SP, MacGlashan DW Jr, Schulman ES, Schleimer RP, Hayes EC, Rokach J, Adkinson NF Jr, Lichtenstein LM: Arachidonic acid metabolism in purified human lung mast cells. J Immunol 1984;132:1972–1979.
28.
Siraganian RP: An automated continuous-flow system for the extraction and fluorometric analysis of histamine. Anal Biochem 1974;57:383–394.
29.
Dvorak AM, Schleimer RP, Schulman ES, Lichtenstein LM: Human mast cells use conservation and condensation mechanisms during recovery from degranulation. In vitro studies with mast cells purified from human lungs. Lab Invest 1986;54:663–678.
30.
Dvorak AM: Procedural guide to specimen handling for the ultrastructural pathology service laboratory (monograph). J Electron Microsc Tech 1987;6:255–301.
31.
Bachmann L, Salpeter MM: Autoradiography with the electron microscope. A quantitative evaluation. Lab Invest 1965;14:1041–1053.
32.
Caro LG, van Tubergen RP, Kolb JA: High-resolution autoradiography. 1. Methods. J Cell Biol 1962;15:173–188.
33.
Dvorak AM, Dvorak HF, Peters SP, Schulman ES, MacGlashan DW Jr, Pyne K, Harvey VS, Galli SJ, Lichtenstein LM: Lipid bodies: Cytoplasmic organelles important to arachidonate metabolism in macrophages and mast cells. J Immunol 1983;131:2965–2976 [republished in J Immunol 1984;132:1586–1597].
34.
Kopriwa B: A comparison of various procedures for fine grain development in electron microscopic radioautography. Histochemistry 1975;44:201–224.
35.
Kopriwa BM: A reliable, standardized method for ultrastructural electron microscopic radioautography. Histochemie 1973;37:1–17.
36.
Kopriwa BM, Levine GM, Nadler NJ: Assessment of resolution by half distance values for tritium and radioiodine in electron microscopic radioautographs using Ilford L4 emulsion developed by ‘solution physical’ or D-19b methods. Histochemistry 1984;80:519–522.
37.
Dvorak AM: Recovery of human lung mast cells from anaphylactic degranulation utilizes a mixture of conservation and synthetic mechanisms, in Galli SJ, Austen KF (eds): Mast Cell and Basophil Differentiation and Function in Health and Disease. New York, Raven, 1989, pp 119–148.
38.
Dvorak AM: Basophil and Mast Cell Degranulation and Recovery; in Harris JR (ed): Blood Cell Biochemistry. New York, Plenum, 1991, vol 4.
39.
Dvorak AM: Human mast cells. Ultrastructural observations of in situ, ex vivo, and in vitro sites, sources, and systems; in Kaliner MA, Metcalfe DD (eds): The Mast Cell in Health and Disease, vol 62 in Lenfant C (series ed): Lung Biology in Health and Disease. New York, Marcel Dekker, 1992, pp 1–90.
40.
Dvorak AM, Schleimer RP, Lichtenstein LM: Morphologic mast cell cycles. Cell Immunol 1987;105:199–204.
41.
Dvorak AM, Schleimer RP, Lichtenstein LM: Human mast cells synthesize new granules during recovery from degranulation. In vitro studies with mast cells purified from human lungs. Blood 1988;71:76–85.
42.
Bernhard W: A new staining procedure for electron microscopical cytology. J Ultrastruct Res 1969;27:250–265.
43.
Hinterberger M, Pettersson I, Steitz JA: Isolation of small nuclear ribonucleoproteins containing U1, U2, U4, U5, and U6 RNAs. J Biol Chem 1983;258:2604–2613.
44.
Pettersson I, Hinterberger M, Mimori T, Gottlieb E, Steitz JA: The structure of mammalian small nuclear ribonucleoproteins. Identification of multiple protein components reactive with anti-(U1)ribonucleoprotein and anti-Sm autoantibodies. J Biol Chem 1984;259:5907–5914.
45.
Padgett RA, Mount SM, Steitz JA, Sharp PA: Splicing of messenger RNA precursors is inhibited by antisera to small nuclear ribonucleoprotein. Cell 1983;35:101–107.
46.
Krúmer A, Keller W, Appel B, Luhrmann R: The 5′ terminus of the RNA moiety of U1 small nuclear ribonucleoprotein particles is required for the splicing of messenger RNA precursors. Cell 1984;38:299–307.
47.
Kuo H-C, Nasim F-UH, Grabowski PJ: Control of alternative splicing by the differential binding of U1 small nuclear ribonucleoprotein particle. Science 1991;251:1045–1050.
48.
Mattaj IW: Splicing stories and poly(A) tales: An update on RNA processing and transport. Curr Opin Cell Biol 1990;2:528–538.
49.
Stevens BJ, Swift H: RNA transport from nucleus to cytoplasm in Chironomus salivary glands. J Cell Biol 1966;31:55–77.
50.
Mehlin H, Lönnroth A, Skoglund U, Daneholt B: Structure and transport of a specific premessenger RNP particle. Cell Biol Int Rep 1988;12:729–736.
51.
Mehlin H, Daneholt B, Skoglund U: Translocation of a specific premessenger ribonucleoprotein particle through the nuclear pore studied with electron microscope tomography. Cell 1992;69:605–613.
52.
Visa N, Izaurralde E, Ferreira J, Daneholt B, Mattaj IW: A nuclear cap-binding complex binds Balbiani ring pre-mRNA cotranscriptionally and accompanies the ribonucleoprotein particle during nuclear export. J Cell Biol 1996;133:5–14.
53.
Sun X, Alzhanova-Ericsson AT, Visa N, Aissouni Y, Zhao J, Daneholt B: The hrp23 protein in the Balbiani ring pre-mRNP particles is released just before or at the binding of the particles to the nuclear pore complex. J Cell Biol 1998;142:1181–1193.
54.
Fakan S: High-resolution autoradiography as a tool for the localization of nucleic acid synthesis and distribution in the mammalian cell nucleus. J Microsc 1976;106:159–171.
55.
Penman S, Vesco C, Penman M: Localization and kinetics of formation of nuclear heterodisperse RNA, cytoplasmic heterodisperse RNA and polyribosome-associated messenger RNA in HeLa cells. J Mol Biol 1968;34:49–69.
56.
Lewin B: Units of transcription and translation: The relationship between heterogeneous nuclear RNA and messenger RNA. Cell 1975;4:11–20.
57.
Bassell G, Singer RH: mRNA and cytoskeletal filaments. Curr Opin Cell Biol 1997;9:109–115.
58.
Bassell GJ, Oleynikov Y, Singer RH: The travels of mRNAs through all cells large and small. FASEB J 1999;13:447–454.
59.
Etkin LD, Lipshitz HD: RNA localization. FASEB J 1999;13:419–420.
60.
Gottlieb E: Messenger RNA transport and localization. Curr Opin Cell Biol 1990;2:1080–1086.
61.
Jansen R-P: RNA-cytoskeletal associations. FASEB J 1999;13:455–466.
62.
Lasko P: RNA sorting in Drosophila oocytes and embryos. FASEB J 1999;13:421–433.
63.
Mowry KL, Cote CA: RNA sorting in Xenopus oocytes and embryos. FASEB J 1999;13:435–445.
64.
Singer RH: The cytoskeleton and mRNA localization. Curr Opin Cell Biol 1992;4:15–19.
65.
Singer RH: RNA zipcodes for cytoplasmic addresses. Intracellular localization of mRNAs appears to be determined by sequences in their 3′ untranslated regions that are composed of multiple elements. Curr Biol 1993;3:719–721.
66.
Wilhelm JE, Vale RD: RNA on the move: The mRNA localization pathway. J Cell Biol 1993;123:269–274.
67.
St. Johnston D: The intracellular localization of messenger RNAs. Cell 1995;81:161–170.
68.
Oleynikov Y, Singer RH: RNA localization: Different zipcodes, same postman? Trends Cell Biol 1998;8:381–383.
69.
Bassell GJ: High resolution distribution of mRNA within the cytoskeleton. J Cell Biochem 1993;52:127–133.
70.
Fulton AB, Wan KM, Penman S: The spatial distribution of polyribosomes in 3T3 cells and the associated assembly of proteins into the skeletal framework. Cell 1980;20:849–857.
71.
Isaacs WB, Fulton AB: Cotranslational assembly of myosin heavy chain in developing cultured skeletal muscle. Proc Natl Acad Sci USA 1987;84:6174–6178.
72.
Lenk R, Ransom L, Kaufmann Y, Penman S: A cytoskeletal structure with associated polyribosomes obtained from HeLa cells. Cell 1977;10:67–78.
73.
Hill MA, Schedlich L, Gunning P: Serum-induced signal transduction determines the peripheral location of β-actin mRNA within the cell. J Cell Biol 1994;126:1221–1230.
74.
Kislauskis EH, Zhu X, Singer RH: Sequences responsible for intracellular localization of b-actin messenger RNA also affect cell phenotype. J Cell Biol 1994;127:441–451.
75.
Knowles RB, Sabry JH, Martone ME, Deerinck TJ, Ellisman MH, Bassell GJ, Kosik KS: Translocation of RNA granules in living neurons. J Neurosci 1996;16:7812–7820.
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