Abstract
Irradiation of intact or EDTA-dissociated L-cell polyribosomes with 254 nm UV light at doses of 1-2 x 10(5) ergs/mm2 extensively crosslinks mRNA to proteins. The crosslinked mRNA-protein complexes can be isolated on the basis of buoyant density in urea-containing CS2SO4 gradients that dissociate non-covalent complexes. Crosslinking of mRNA can also be assayed by phenolchloroform extraction. mRNA recovered from the crosslinked complexes by digestion with proteinase K has the same electrophoretic mobility in polyacrylamide gels as unirradiated mRNA. Therefore, irradiation does not either crosslink RNA molecules to RNA molecules or break phosphodiester bonds. With these methods it has been found that more than 70% of high molecular weight polydisperse mRNA, but only 25-40% of histone mRNA, can be crosslinked to protein. On the basis of buoyant density the histone mRNA-protein complex had a protein content of 26%, whereas the mean protein content of most non-histone mRNA-protein complexes was 65%. It is concluded that most mRNA in polyribosomes is in close contact with proteins, and that histone mRNA can be crosslinked to many fewer proteins that most other mRNAs.
Full text
PDF

















Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Adesnik M., Darnell J. E. Biogenesis and characterization of histone messenger RNA in HeLa cells. J Mol Biol. 1972 Jun 28;67(3):397–406. doi: 10.1016/0022-2836(72)90458-5. [DOI] [PubMed] [Google Scholar]
- Anderson E., Nakashima Y., Konigsberg W. Photo-incuced cross-linkage of gene-5 protein and bacteriophage fd DNA+. Nucleic Acids Res. 1975 Mar;2(3):361–371. doi: 10.1093/nar/2.3.361. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Borun T. W., Scharff M. D., Robbins E. Rapidly labeled, polyribosome-associated RNA having the properties of histone messenger. Proc Natl Acad Sci U S A. 1967 Nov;58(5):1977–1983. doi: 10.1073/pnas.58.5.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Budzik G. P., Lam S. S., Schoemaker H. J., Schimmel P. R. Two photo-cross-linked complexes of isoleucine specific transfer ribonucleic acid with aminoacyl transfer ribonucleic acid synthetases. J Biol Chem. 1975 Jun 25;250(12):4433–4439. [PubMed] [Google Scholar]
- Cohen L. H., Newrock K. M., Zweidler A. Stage-specific switches in histone synthesis during embryogenesis of the sea urchin. Science. 1975 Dec 5;190(4218):994–997. doi: 10.1126/science.1237932. [DOI] [PubMed] [Google Scholar]
- Craig N. C. On the regulation of the synthesis of ribosomal proteins in L-cells. J Mol Biol. 1971 Jan 14;55(1):129–134. doi: 10.1016/0022-2836(71)90288-9. [DOI] [PubMed] [Google Scholar]
- Gallwitz D., Mueller G. C. Histone synthesis in vitro on HeLa cell microsomes. The nature of the coupling to deoxyribonucleic acid synthesis. J Biol Chem. 1969 Nov 10;244(21):5947–5952. [PubMed] [Google Scholar]
- Greenberg J. R. Isolation of L-cell messenger RNA which lacks poly(adenylate). Biochemistry. 1976 Aug 10;15(16):3516–3522. doi: 10.1021/bi00661a019. [DOI] [PubMed] [Google Scholar]
- Greenberg J. R. Isolation of messenger ribonucleoproteins in cesium sulfate density gradients: evidence that polyadenylated and non-polyadenylated messenger RNAs are associated with protein. J Mol Biol. 1976 Dec;108(2):403–416. doi: 10.1016/s0022-2836(76)80127-1. [DOI] [PubMed] [Google Scholar]
- Greenberg J. R. Messenger RNA metabolism of animal cells. Possible involvement of untranslated sequences and mRNA-associated proteins. J Cell Biol. 1975 Feb;64(2):269–288. doi: 10.1083/jcb.64.2.269. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kunkel N. S., Weinberg E. S. Histone gene transcripts in the cleavage and mesenchyme blastula embryo of the sea urchin, S. purpuratus. Cell. 1978 Jun;14(2):313–326. doi: 10.1016/0092-8674(78)90117-4. [DOI] [PubMed] [Google Scholar]
- Lin S. Y., Riggs A. D. Photochemical attachment of lac repressor to bromodeoxyuridine-substituted lac operator by ultraviolet radiation. Proc Natl Acad Sci U S A. 1974 Mar;71(3):947–951. doi: 10.1073/pnas.71.3.947. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Melli M., Spinelli G., Arnold E. Synthesis of histone messenger RNA of HeLa cells during the cell cycle. Cell. 1977 Sep;12(1):167–174. doi: 10.1016/0092-8674(77)90194-5. [DOI] [PubMed] [Google Scholar]
- Modak S. P., Commelin D., Grosset L., Imaizumi M. T., Monnat M., Scherrer K. DNA synthesis in circulating erythroblasts of anemic duck. Isolation and properties of nuclear and cytoplasmic-nonmitochondrial DNA. Eur J Biochem. 1975 Dec 15;60(2):407–421. doi: 10.1111/j.1432-1033.1975.tb21017.x. [DOI] [PubMed] [Google Scholar]
- Möller K., Brimacombe R. Specific cross-linking of proteins S7 and L4 to ribosomal RNA, by UV irradiation of Escherichia coli ribosomal subunits. Mol Gen Genet. 1975 Dec 9;141(4):343–355. doi: 10.1007/BF00331455. [DOI] [PubMed] [Google Scholar]
- Newrock K. M., Cohen L. H., Hendricks M. B., Donnelly R. J., Weinberg E. S. Stage-specific mRNAs coding for subtypes of H2A and H2B histones in the sea urchin embryo. Cell. 1978 Jun;14(2):327–336. doi: 10.1016/0092-8674(78)90118-6. [DOI] [PubMed] [Google Scholar]
- Perry R. P., Kelley D. E. Buoyant densities of cytoplasmic ribonucleoprotein particles of mammalian cells: distinctive character of ribosome subunits and the rapidly labeled components. J Mol Biol. 1966 Apr;16(2):255–268. doi: 10.1016/s0022-2836(66)80171-7. [DOI] [PubMed] [Google Scholar]
- Perry R. P., Kelley D. E. Messenger RNA turnover in mouse L cells. J Mol Biol. 1973 Oct 5;79(4):681–696. doi: 10.1016/0022-2836(73)90071-5. [DOI] [PubMed] [Google Scholar]
- Perry R. P., Kelley D. E. Messenger RNA-protein complexes and newly synthesized ribosomal subunits: analysis of free particles and components of polyribosomes. J Mol Biol. 1968 Jul 14;35(1):37–59. doi: 10.1016/s0022-2836(68)80035-x. [DOI] [PubMed] [Google Scholar]
- Perry R. P., La Torre J., Kelley D. E., Greenberg J. R. On the lability of poly(A) sequences during extraction of messenger RNA from polyribosomes. Biochim Biophys Acta. 1972 Mar 14;262(2):220–226. doi: 10.1016/0005-2787(72)90236-5. [DOI] [PubMed] [Google Scholar]
- Preobrazhensky A. A., Spirin A. S. Informosomes and their protein components: the present state of knowledge. Prog Nucleic Acid Res Mol Biol. 1978;21:1–38. doi: 10.1016/s0079-6603(08)60265-2. [DOI] [PubMed] [Google Scholar]
- Salomon J., Elad D. Photochemical reactions of nucleic acid constituents. Peroxide-initiated reactions of purines with alcohols. J Org Chem. 1973 Sep 21;38(19):3420–3421. doi: 10.1021/jo00959a044. [DOI] [PubMed] [Google Scholar]
- Schaffner W., Kunz G., Daetwyler H., Telford J., Smith H. O., Birnstiel M. L. Genes and spacers of cloned sea urchin histone DNA analyzed by sequencing. Cell. 1978 Jul;14(3):655–671. doi: 10.1016/0092-8674(78)90249-0. [DOI] [PubMed] [Google Scholar]
- Schodhetman G., Perry R. P. Early appearance of histone messenger RNA in polyribosomes of cultured L cells. J Mol Biol. 1972 Feb 14;63(3):591–596. doi: 10.1016/0022-2836(72)90450-0. [DOI] [PubMed] [Google Scholar]
- Schoemaker H. J., Schimmel P. R. Photo-induced joining of a transfer RNA with its cognate aminoacyl-transfer RNA synthetase. J Mol Biol. 1974 Apr 25;84(4):503–513. doi: 10.1016/0022-2836(74)90112-0. [DOI] [PubMed] [Google Scholar]
- Schott H. N., Shetlar M. D. Photochemical addition of amino acids to thymine. Biochem Biophys Res Commun. 1974 Aug 5;59(3):1112–1116. doi: 10.1016/s0006-291x(74)80093-8. [DOI] [PubMed] [Google Scholar]
- Sen A., Todaro G. J. The genome-associated, specific RNA binding proteins of avian and mammalian type C viruses. Cell. 1977 Jan;10(1):91–99. doi: 10.1016/0092-8674(77)90143-x. [DOI] [PubMed] [Google Scholar]
- Smith K. C. A mixed photoproduct of thymine and cysteine: 5-S-cysteine, 6-hydrothymine. Biochem Biophys Res Commun. 1970;39(6):1011–1016. doi: 10.1016/0006-291x(70)90658-3. [DOI] [PubMed] [Google Scholar]
- Smith K. C., Aplin R. T. A mixed photoproduct of uracil and cysteine (5-S-cysteine-6-hydrouracil). A possible model for the in vivo cross-linking of deoxyribonucleic acid and protein by ultraviolet light. Biochemistry. 1966 Jun;5(6):2125–2130. doi: 10.1021/bi00870a046. [DOI] [PubMed] [Google Scholar]
- Smith K. C. Photochemical addition of amino acids to 14C-uracil. Biochem Biophys Res Commun. 1969 Feb 7;34(3):354–357. doi: 10.1016/0006-291x(69)90840-7. [DOI] [PubMed] [Google Scholar]
- Wilt F. H., Anderson M., Ekenberg E. Centrifugation of nuclear ribonucleoprotein particles of sea urchin embryos in cesium sulfate. Biochemistry. 1973 Feb 27;12(5):959–966. doi: 10.1021/bi00729a027. [DOI] [PubMed] [Google Scholar]