Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1988 May;85(10):3541–3545. doi: 10.1073/pnas.85.10.3541

Influence of 5' flanking sequences on TL and H-2 expression in transfected L cells.

Y Obata 1, E Stockert 1, Y T Chen 1, T Takahashi 1, L J Old 1
PMCID: PMC280249  PMID: 3368465

Abstract

TL (thymus leukemia) antigens are encoded by genes in the major histocompatibility complex (MHC) of the mouse. Although similar in overall structure to other class I MHC antigens (H-2, Qa), TL expression is regulated in a highly distinctive fashion. In contrast to the broad distribution of H-2 and the intermediate distribution of Qa, TL expression is restricted to cells of T-cell derivation during development in the thymus and is lost when T cells migrate to the periphery. Some mouse strains do not express TL antigens on thymocytes (TL- strains), but leukemias occurring in these mice can have a TL+ phenotype, indicating activation of normally silent TL genes. In transfection studies with H-2 or TL genes in L cells (mouse fibroblasts), H-2 is expressed at high levels, whereas TL is poorly expressed. To identify genetic elements that regulate expression in transfected L cells, chimeric genes were constructed by transposing the 5' and 3' regions of TL and H-2 genes. Antigen expression was not influenced by transposing the cytoplasmic domain and 3' untranslated region. In contrast, interchanging the 5' flanking sequences and exon 1 had a marked influence on antigen expression, with 5' sequences from the H-2 gene increasing TL expression 10- to 50-fold, and 5' sequences from the TL gene markedly decreasing H-2 expression. With both the parental TL gene (p20-TL) and the highly expressed chimeric TL gene (construct 3), levels of TL mRNA and TL antigen correlated with the number of transfected gene copies. However, in cells transfected with equal copy numbers, much higher levels of TL mRNA and TL antigen were found in construct-3 transfectants than in p20-TL transfectants. In addition, there was marked heterogeneity in TL mRNA size in L cells transfected with p20-TL, in contrast to a more homogeneous transcript size in construct-3 transfectants. These results point to regulatory sequences in the 5' flanking region of class I genes that control proper initiation and processing of TL transcripts.

Full text

PDF
3541

Images in this article

3544Image
on p.3544
3544Image
on p.3544
3545Image
on p.3545

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allen H., Wraith D., Pala P., Askonas B., Flavell R. A. Domain interactions of H-2 class I antigens alter cytotoxic T-cell recognition sites. Nature. 1984 May 17;309(5965):279–281. doi: 10.1038/309279a0. [DOI] [PubMed] [Google Scholar]
  2. Arnold B., Burgert H. G., Hamann U., Hämmerling G., Kees U., Kvist S. Cytolytic T cells recognize the two amino-terminal domains of H-2 K antigens in tandem in influenza A infected cells. Cell. 1984 Aug;38(1):79–87. doi: 10.1016/0092-8674(84)90528-2. [DOI] [PubMed] [Google Scholar]
  3. Baldwin A. S., Jr, Sharp P. A. Binding of a nuclear factor to a regulatory sequence in the promoter of the mouse H-2Kb class I major histocompatibility gene. Mol Cell Biol. 1987 Jan;7(1):305–313. doi: 10.1128/mcb.7.1.305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Blin N., Stafford D. W. A general method for isolation of high molecular weight DNA from eukaryotes. Nucleic Acids Res. 1976 Sep;3(9):2303–2308. doi: 10.1093/nar/3.9.2303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen Y. T., Obata Y., Stockert E., Old L. J. Thymus-leukemia (TL) antigens of the mouse. Analysis of TL mRNA and TL cDNA TL+ and TL- strains. J Exp Med. 1985 Oct 1;162(4):1134–1148. doi: 10.1084/jem.162.4.1134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen Y. T., Obata Y., Stockert E., Takahashi T., Old L. J. Tla-region genes and their products. Immunol Res. 1987;6(1-2):30–45. doi: 10.1007/BF02918102. [DOI] [PubMed] [Google Scholar]
  8. Dynan W. S., Tjian R. Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins. 1985 Aug 29-Sep 4Nature. 316(6031):774–778. doi: 10.1038/316774a0. [DOI] [PubMed] [Google Scholar]
  9. Evans G. A., Margulies D. H., Shykind B., Seidman J. G., Ozato K. Exon shuffling: mapping polymorphic determinants on hybrid mouse transplantation antigens. Nature. 1982 Dec 23;300(5894):755–757. doi: 10.1038/300755a0. [DOI] [PubMed] [Google Scholar]
  10. Fisher D. A., Hunt S. W., 3rd, Hood L. Structure of a gene encoding a murine thymus leukemia antigen, and organization of Tla genes in the BALB/c mouse. J Exp Med. 1985 Aug 1;162(2):528–545. doi: 10.1084/jem.162.2.528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Goodenow R. S., McMillan M., Nicolson M., Sher B. T., Eakle K., Davidson N., Hood L. Identification of the class I genes of the mouse major histocompatibility complex by DNA-mediated gene transfer. Nature. 1982 Nov 18;300(5889):231–237. doi: 10.1038/300231a0. [DOI] [PubMed] [Google Scholar]
  12. Hämmerling G. J., Rüsch E., Tada N., Kimura S., Hämmerling U. Localization of allodeterminants on H-2Kb antigens determined with monoclonal antibodies and H-2 mutant mice. Proc Natl Acad Sci U S A. 1982 Aug;79(15):4737–4741. doi: 10.1073/pnas.79.15.4737. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kimura A., Israël A., Le Bail O., Kourilsky P. Detailed analysis of the mouse H-2Kb promoter: enhancer-like sequences and their role in the regulation of class I gene expression. Cell. 1986 Jan 31;44(2):261–272. doi: 10.1016/0092-8674(86)90760-9. [DOI] [PubMed] [Google Scholar]
  14. Margulies D. H., Ramsey A. L., Boyd L. F., McCluskey J. Genetic engineering of an H-2Dd/Q10b chimeric histocompatibility antigen: purification of soluble protein from transformant cell supernatants. Proc Natl Acad Sci U S A. 1986 Jul;83(14):5252–5256. doi: 10.1073/pnas.83.14.5252. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Obata Y., Chen Y. T., Stockert E., Old L. J. Structural analysis of TL genes of the mouse. Proc Natl Acad Sci U S A. 1985 Aug;82(16):5475–5479. doi: 10.1073/pnas.82.16.5475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Old L. J., Stockert E. Immunogenetics of cell surface antigens of mouse leukemia. Annu Rev Genet. 1977;11:127–160. doi: 10.1146/annurev.ge.11.120177.001015. [DOI] [PubMed] [Google Scholar]
  17. Ozato K., Evans G. A., Shykind B., Margulies D. H., Seidman J. G. Hybrid H-2 histocompatibility gene products assign domains recognized by alloreactive T cells. Proc Natl Acad Sci U S A. 1983 Apr;80(7):2040–2043. doi: 10.1073/pnas.80.7.2040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Pontarotti P. A., Mashimo H., Zeff R. A., Fisher D. A., Hood L., Mellor A., Flavell R. A., Nathenson S. G. Conservation and diversity in the class I genes of the major histocompatibility complex: sequence analysis of a Tlab gene and comparison with a Tlac gene. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1782–1786. doi: 10.1073/pnas.83.6.1782. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  20. Schulze D. H., Pease L. R., Obata Y., Nathenson S. G., Reyes A. A., Ikuta S., Wallace R. B. Identification of the cloned gene for the murine transplantation antigen H-2Kb by hybridization with synthetic oligonucleotides. Mol Cell Biol. 1983 Apr;3(4):750–755. doi: 10.1128/mcb.3.4.750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Straus D. S., Stroynowski I., Schiffer S. G., Hood L. Expression of hybrid class I genes of the major histocompatibility complex in mouse L cells. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6245–6249. doi: 10.1073/pnas.82.18.6245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Stroynowski I., Orn A., Goodenow R. S., McMillan M., Forman J., Brayton P. R., Frelinger J., Hood L. Cytotoxic T lymphocytes recognize determinants on the BALB/c-H-2Ld molecule controlled by alpha 1 and alpha 2 but not alpha 3 external domains. Immunogenetics. 1984;20(2):141–154. doi: 10.1007/BF00364486. [DOI] [PubMed] [Google Scholar]
  23. Stroynowski I., Soloski M., Low M. G., Hood L. A single gene encodes soluble and membrane-bound forms of the major histocompatibility Qa-2 antigen: anchoring of the product by a phospholipid tail. Cell. 1987 Aug 28;50(5):759–768. doi: 10.1016/0092-8674(87)90334-5. [DOI] [PubMed] [Google Scholar]
  24. Ullrich A., Shine J., Chirgwin J., Pictet R., Tischer E., Rutter W. J., Goodman H. M. Rat insulin genes: construction of plasmids containing the coding sequences. Science. 1977 Jun 17;196(4296):1313–1319. doi: 10.1126/science.325648. [DOI] [PubMed] [Google Scholar]
  25. Waneck G. L., Sherman D. H., Calvin S., Allen H., Flavell R. A. Tissue-specific expression of cell-surface Qa-2 antigen from a transfected Q7b gene of C57BL/10 mice. J Exp Med. 1987 May 1;165(5):1358–1370. doi: 10.1084/jem.165.5.1358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Weiss E., Golden L., Zakut R., Mellor A., Fahrner K., Kvist S., Flavell R. A. The DNA sequence of the H-2kb gene: evidence for gene conversion as a mechanism for the generation of polymorphism in histocompatibilty antigens. EMBO J. 1983;2(3):453–462. doi: 10.1002/j.1460-2075.1983.tb01444.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Wigler M., Silverstein S., Lee L. S., Pellicer A., Cheng Y. c., Axel R. Transfer of purified herpes virus thymidine kinase gene to cultured mouse cells. Cell. 1977 May;11(1):223–232. doi: 10.1016/0092-8674(77)90333-6. [DOI] [PubMed] [Google Scholar]
  28. Yano O., Kanellopoulos J., Kieran M., Le Bail O., Israël A., Kourilsky P. Purification of KBF1, a common factor binding to both H-2 and beta 2-microglobulin enhancers. EMBO J. 1987 Nov;6(11):3317–3324. doi: 10.1002/j.1460-2075.1987.tb02652.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES