Abstract
Patients with humoral autoimmune diseases such as systemic lupus erythematosus and Sjögren's syndrome contain antibodies in their sera directed against certain normal cellular components such as the La/SS-B autoantigen, an RNA-binding protein believed to function as a putative processor of RNA polymerase III precursor transcripts. We have identified cDNA clones from the fruit fly Drosophila melanogaster that encode a protein displaying significant sequence homology with human La/SS-B. The fly protein (which we refer to as D-La) contains a putative ribonucleoprotein 1 (RNP1) and RNP2 RNA-binding domain. D-La also possesses a leucine zipper motif, suggesting that it may interact with itself or other proteins. Using gel retardation analysis, we show that D-La can bind RNA; in addition, we demonstrate the first reported DNA-binding activity associated with a La protein. Northern (RNA) blot analysis revealed a single 1,600-nucleotide transcript expressed throughout embryonic, larval, pupal, and adult development. Surprisingly, whole-mount in situ hybridization experiments revealed that D-La transcripts are not present in all ovarian tissues. In addition, early expression throughout the embryo is followed by a restricted pattern of mesodermal expression that is later confined to the visceral mesoderm, gonads, gut, and salivary glands. These results suggest that D-La may play a more specialized role during fly development as opposed to a rather general role inferred by its homology to La proteins from other organisms.
Full text
PDF![5123](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/5de7091a166e/molcellb00008-0115.png)
![5124](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/bb4d49adbec2/molcellb00008-0116.png)
![5125](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/82fb838b653a/molcellb00008-0117.png)
![5126](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/6adac3fad65c/molcellb00008-0118.png)
![5127](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/6063943c2e0f/molcellb00008-0119.png)
![5128](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/4a66576ba050/molcellb00008-0120.png)
![5129](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/92de/359031/2ee69fbc5640/molcellb00008-0121.png)
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bachmann M., Falke D., Schröder H. C., Müller W. E. Intracellular distribution of the La antigen in CV-1 cells after herpes simplex virus type 1 infection compared with the localization of U small nuclear ribonucleoprotein particles. J Gen Virol. 1989 Apr;70(Pt 4):881–891. doi: 10.1099/0022-1317-70-4-881. [DOI] [PubMed] [Google Scholar]
- Bachmann M., Pfeifer K., Schröder H. C., Müller W. E. The La antigen shuttles between the nucleus and the cytoplasm in CV-1 cells. Mol Cell Biochem. 1989 Feb 21;85(2):103–114. doi: 10.1007/BF00577106. [DOI] [PubMed] [Google Scholar]
- Cavener D. R., Ray S. C. Eukaryotic start and stop translation sites. Nucleic Acids Res. 1991 Jun 25;19(12):3185–3192. doi: 10.1093/nar/19.12.3185. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chambers J. C., Kenan D., Martin B. J., Keene J. D. Genomic structure and amino acid sequence domains of the human La autoantigen. J Biol Chem. 1988 Dec 5;263(34):18043–18051. [PubMed] [Google Scholar]
- Chan E. K., Sullivan K. F., Tan E. M. Ribonucleoprotein SS-B/La belongs to a protein family with consensus sequences for RNA-binding. Nucleic Acids Res. 1989 Mar 25;17(6):2233–2244. doi: 10.1093/nar/17.6.2233. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dingwall C., Laskey R. A. Nuclear targeting sequences--a consensus? Trends Biochem Sci. 1991 Dec;16(12):478–481. doi: 10.1016/0968-0004(91)90184-w. [DOI] [PubMed] [Google Scholar]
- Fried M., Crothers D. M. Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Dec 11;9(23):6505–6525. doi: 10.1093/nar/9.23.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hultmark D. Immune reactions in Drosophila and other insects: a model for innate immunity. Trends Genet. 1993 May;9(5):178–183. doi: 10.1016/0168-9525(93)90165-e. [DOI] [PubMed] [Google Scholar]
- Ip Y. T., Reach M., Engstrom Y., Kadalayil L., Cai H., González-Crespo S., Tatei K., Levine M. Dif, a dorsal-related gene that mediates an immune response in Drosophila. Cell. 1993 Nov 19;75(4):753–763. doi: 10.1016/0092-8674(93)90495-c. [DOI] [PubMed] [Google Scholar]
- Johnson P. F., McKnight S. L. Eukaryotic transcriptional regulatory proteins. Annu Rev Biochem. 1989;58:799–839. doi: 10.1146/annurev.bi.58.070189.004055. [DOI] [PubMed] [Google Scholar]
- Lerner M. R., Andrews N. C., Miller G., Steitz J. A. Two small RNAs encoded by Epstein-Barr virus and complexed with protein are precipitated by antibodies from patients with systemic lupus erythematosus. Proc Natl Acad Sci U S A. 1981 Feb;78(2):805–809. doi: 10.1073/pnas.78.2.805. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Madore S. J., Wieben E. D., Pederson T. Eukaryotic small ribonucleoproteins. Anti-La human autoantibodies react with U1 RNA-protein complexes. J Biol Chem. 1984 Feb 10;259(3):1929–1933. [PubMed] [Google Scholar]
- Mariani B. D., Lingappa J. R., Kafatos F. C. Temporal regulation in development: negative and positive cis regulators dictate the precise timing of expression of a Drosophila chorion gene. Proc Natl Acad Sci U S A. 1988 May;85(9):3029–3033. doi: 10.1073/pnas.85.9.3029. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mathews M. B., Francoeur A. M. La antigen recognizes and binds to the 3'-oligouridylate tail of a small RNA. Mol Cell Biol. 1984 Jun;4(6):1134–1140. doi: 10.1128/mcb.4.6.1134. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nonaka M., Huang Z. M. Interleukin-1-mediated enhancement of mouse factor B gene expression via NF kappa B-like hepatoma nuclear factor. Mol Cell Biol. 1990 Dec;10(12):6283–6289. doi: 10.1128/mcb.10.12.6283. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Oikawa S., Nakazato H., Kosaki G. Primary structure of human carcinoembryonic antigen (CEA) deduced from cDNA sequence. Biochem Biophys Res Commun. 1987 Jan 30;142(2):511–518. doi: 10.1016/0006-291x(87)90304-4. [DOI] [PubMed] [Google Scholar]
- Pruijn G. J., Slobbe R. L., Van Venrooij W. J. Structure and function of La and Ro RNPs. Mol Biol Rep. 1990;14(2-3):43–48. doi: 10.1007/BF00360410. [DOI] [PubMed] [Google Scholar]
- Reddy R., Henning D., Tan E., Busch H. Identification of a La protein binding site in a RNA polymerase III transcript (4.5 I RNA). J Biol Chem. 1983 Jul 10;258(13):8352–8356. [PubMed] [Google Scholar]
- Rinke J., Steitz J. A. Association of the lupus antigen La with a subset of U6 snRNA molecules. Nucleic Acids Res. 1985 Apr 11;13(7):2617–2629. doi: 10.1093/nar/13.7.2617. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rinke J., Steitz J. A. Precursor molecules of both human 5S ribosomal RNA and transfer RNAs are bound by a cellular protein reactive with anti-La lupus antibodies. Cell. 1982 May;29(1):149–159. doi: 10.1016/0092-8674(82)90099-x. [DOI] [PubMed] [Google Scholar]
- Saunders R. D., Glover D. M., Ashburner M., Siden-Kiamos I., Louis C., Monastirioti M., Savakis C., Kafatos F. PCR amplification of DNA microdissected from a single polytene chromosome band: a comparison with conventional microcloning. Nucleic Acids Res. 1989 Nov 25;17(22):9027–9037. doi: 10.1093/nar/17.22.9027. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Scherly D., Stutz F., Lin-Marq N., Clarkson S. G. La proteins from Xenopus laevis. cDNA cloning and developmental expression. J Mol Biol. 1993 May 20;231(2):196–204. doi: 10.1006/jmbi.1993.1275. [DOI] [PubMed] [Google Scholar]
- Semsei I., Tröster H., Bartsch H., Schwemmle M., Igloi G. L., Bachmann M. Isolation of rat cDNA clones coding for the autoantigen SS-B/La: detection of species-specific variations. Gene. 1993 Apr 30;126(2):265–268. doi: 10.1016/0378-1119(93)90378-g. [DOI] [PubMed] [Google Scholar]
- Smith D. B., Johnson K. S. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. doi: 10.1016/0378-1119(88)90005-4. [DOI] [PubMed] [Google Scholar]
- Stefano J. E. Purified lupus antigen La recognizes an oligouridylate stretch common to the 3' termini of RNA polymerase III transcripts. Cell. 1984 Jan;36(1):145–154. doi: 10.1016/0092-8674(84)90083-7. [DOI] [PubMed] [Google Scholar]
- Suter B., Steward R. Requirement for phosphorylation and localization of the Bicaudal-D protein in Drosophila oocyte differentiation. Cell. 1991 Nov 29;67(5):917–926. doi: 10.1016/0092-8674(91)90365-6. [DOI] [PubMed] [Google Scholar]
- Tautz D., Pfeifle C. A non-radioactive in situ hybridization method for the localization of specific RNAs in Drosophila embryos reveals translational control of the segmentation gene hunchback. Chromosoma. 1989 Aug;98(2):81–85. doi: 10.1007/BF00291041. [DOI] [PubMed] [Google Scholar]
- Tolias P. P., Kafatos F. C. Functional dissection of an early Drosophila chorion gene promoter: expression throughout the follicular epithelium is under spatially composite regulation. EMBO J. 1990 May;9(5):1457–1464. doi: 10.1002/j.1460-2075.1990.tb08262.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Topfer F., Gordon T., McCluskey J. Characterization of the mouse autoantigen La (SS-B). Identification of conserved RNA-binding motifs, a putative ATP binding site and reactivity of recombinant protein with poly(U) and human autoantibodies. J Immunol. 1993 Apr 1;150(7):3091–3100. [PubMed] [Google Scholar]
- Vinson C. R., Sigler P. B., McKnight S. L. Scissors-grip model for DNA recognition by a family of leucine zipper proteins. Science. 1989 Nov 17;246(4932):911–916. doi: 10.1126/science.2683088. [DOI] [PubMed] [Google Scholar]