Skip to main content
Log in

Mapping and expression of a bifunctional thymidylate synthase, dihydrofolate reductase gene from maize

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

A bifunctional gene (ZmDHFR-TS) encoding dihydrofolate reductase (DHFR) and thymidylate synthase (TS) was cloned from a Zea mays cDNA library. Both of these enzymes are involved in nucleotide biosynthesis, specifically in the formation of thymidine monophosphate (TMP). Comparison of the deduced amino acid sequence with DHFR-TS sequences from three other plant sources revealed over 75% similarity and motifs typical of DHFR-TS proteins. Two copies of the gene were mapped to chromosomes 2 and 4. This represents the first DHFR-TS gene cloned from a monocotyledonous plant. Expression of ZmDHFR-TS was examined in developing kernels and various tissues of maize by RNA gel blot hybridization analysis in order to determine the relationship between expression of this gene and DNA synthesis. RNA transcripts for ZmDHFR-TS accumulated to high levels in developing maize kernels when endosperm cells were undergoing endoreduplication and cell division. Meristematic maize tissues had high levels of ZmDHFR-TS mRNA, but transcripts were barely detectable in RNA isolated from the root elongation zone and from mature leaf tissues.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Abbe, E.C. and Stein, O.L. 1954. The growth of the shoot apex in maize: embryogeny. Am. J. Bot. 41: 285–293.

    Google Scholar 

  • Ayusawa, D., Shimizu, K., Koyama, H., Kaneda, S., Tekeishi, K. and Seno, T. 1986. Cell-cycle directed regulation of thymidylate synthase messenger RNA in human diploid fibroblasts stimulated to proliferate. J. Mol Biol. 190: 559–567.

    PubMed  Google Scholar 

  • Balestrazzi, A., Branzoni, M., Carbonera, D., Parisi, B. and Cella, R. 1995. Biochemical evidence for the presence of a bifunctional dihydrofolate reductase-thymidylate synthase in plant species. J. Plant Physiol. 147: 263–266.

    Google Scholar 

  • Bass, H.W., Goode, J.H. and Greene, T.W. and Boston, R.S. 1994. Control of ribosome-inactivating protein (RIP) RNA levels during maize seed development. Plant Sci. 101: 17–30.

    Google Scholar 

  • Cella, R. and Parisi, B. 1993. Dihydrofolate reductase and thymidy-late synthase in plants: an open question. Physiol. Plant. 88: 509–521.

    Google Scholar 

  • Cossins, E.A. and Chen, L. 1997. Folates and one-carbon metabolism in plants and fungi. Phytochemistry 45: 437–452.

    PubMed  Google Scholar 

  • Grafi, G. and Larkins, B.A. 1995. Endoreduplication in maize endosperm: involvement of M phase-promoting factor inhibi-tion and induction of S phase-related kinases. Science 269: 1262–1264.

    Google Scholar 

  • Helentjaris, T. 1995. Atlas of duplicated sequences. Maize Genet. Coop. Newsl. 69: 67–81.

    Google Scholar 

  • Ivanitech, K.M. and Santi, D.V. 1990. Bifunctional thymidylate synthase-dihydrofolate reductase in protozoa. FASEB J 4: 1591–1597.

    PubMed  Google Scholar 

  • Jenh, C.H., Geyer, P.K. and Johnson, L.F. 1985. Control of thymidylate synthase mRNA content and gene transcription in an overproducing mouse cell line. Mol. Cell Biol. 10: 2527–2532.

    Google Scholar 

  • Kim, S.K., Nosaka, K., Downs, D.M., Kwak, J.M., Park, D., Chung, K. and Nam, H.G. 1998. A Brassica cDNA clone encoding a bifunctional hydroymethylpyrimidine kinase/thiamin-phosphate pyrophosphorylase involved in thiamin biosynthesis. Plant Mol. Biol. 37: 955–966.

    PubMed  Google Scholar 

  • Kornberg, A. 1980. DNA Replication. W.H. Freeman and Co., San Francisco.

    Google Scholar 

  • Kowles, R.V. and Phillips, R.L. 1985. DNA amplification patterns in maize endosperm nuclei during kernel development. Proc. Natl. Acad. Sci. USA 82: 7010–7015.

    Google Scholar 

  • Kowles, R.V. and Phillips, R.L. 1988. Endosperm development in maize. Int. Rev. Cytol. 112: 97–136.

    Google Scholar 

  • Kowles, R.V., Srienc, F. and Phillips, R.L. 1990. Endoreduplication of nuclear DNA in the developing maize endosperm. Dev. Genet. 11: 125–132.

    Google Scholar 

  • Lazar, G., Zhang, H. and Goodman, H.M. 1993. The origin of the bifunctional dihydrofolate reductase-thymidylate synthase isogenes of Arabidopsis thaliana. Plant J. 3: 657–668.

    PubMed  Google Scholar 

  • Leitch, I. and Bennett, M.D. 1997. Polyploidy in angiosperms. Trends Plant Sci. 2: 470–476.

    Google Scholar 

  • Luo, M., Piffanelli, P., Rastelli, L. and Cella, R. 1993. Molecu-lar cloning and analysis of a cDNA coding for the bifunctional dihydrofolate reductase-thymidylate synthase of Daucus carota. Plant Mol. Biol. 22: 427–435.

    PubMed  Google Scholar 

  • Luo, M., Orsi, R., Patrucco, E., Pancaldi, S. and Cella, R. 1997. Multiple transcription start sites of the carrot dihydro-folate reductase-thymidylate synthase gene, and sub-cellular localization of the bifunctional protein. Plant Mol. Biol 33: 709–722.

    PubMed  Google Scholar 

  • Neuburger, M., Rebeille, F., Jourdain, A., Nakamura, S. and Douce, R. 1996. Mitochondria are a major site for folate and thymidylate synthesis in plants. J. Biol. Chem. 271: 9466–9472.

    PubMed  Google Scholar 

  • Phillips, R.L., Wang, A.S. and Kowles, R.V. 1983. Molecular and developmental cytogenetics of gene multiplicity in maize. Stadler Genet. Symp. 15: 105–118.

    Google Scholar 

  • Phillips, R.L., Kowles, R.V., McMullen, M.D. and Rubenstein, E.S. 1985. Developmentally timed changes in maize endosperm DNA. Alan R. Liss., New York.

    Google Scholar 

  • Ratnam, S., Delcamp, T.J., Hynes, J.B. and Freisheim, J.H. 1987. Purification and characterization of dihydrofolate reductase from soybean seedlings. Arch. Biochem. Biophys. 255: 279–283.

    PubMed  Google Scholar 

  • Sambrook, J., Fritsch, E.F. and Maniatis, T. 1989. Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  • Senior, M.L., Chin, E.C.L., Lee, M., Smith, J.S.C. and Stuber, C.W. 1996. Simple sequence repeat markers developed from maize sequences found in the GenBank database: map construction. Crop Sci. 36: 1676–1683.

    Google Scholar 

  • Shah, D.M., Hightower, R.C. and Meagher, R.B. 1983. Genes encoding actin in higher plants: intron positions are highly con-served but the coding sequences are not. Mol. Appl. Genet. 2: 111–126.

    Google Scholar 

  • Storms, R.K., Ord, R.W., Grenwood, M.T., Mirdamadi, B., Chu, F.K. and Belfort, M. 1984. Cell cycle dependent expression of thymidylate synthase in Saccharomyces cerevisiae. Mol. Cell. Biol. 4: 2858–2864.

    PubMed  Google Scholar 

  • Thompson, W.F., Everett, M., Polans, N.O., Jorgensen, R.A. and Palmer, J.D. 1993. Phytochrome control of RNA levels in developing pea and mung bean leaves. Planta 158: 487–500.

    Google Scholar 

  • Toth, I., Lazar, G. and Goodman, H.M. 1987. Purification and immunochemical characterization of a dihydrofolate reductase-thymidylate synthase enzyme complex from wild carrot cells. EMBO J. 6: 1853–1858.

    Google Scholar 

  • Ullah, M.H., Robertson, D. and Fites, R. 1999. A gene for thymi-dine kinase in plants. (Accession No. AF066050) (PGR 99-048). Plant Physiol. 119: 1567.

    PubMed  Google Scholar 

  • Vandiver, V.V. and Fites, R.C. 1979. Thymidylate synthase activity from Chlamydomonas cells and cultured tissues of Nicotiana, Pinus,andDaucus. Plant Physiol. 64: 668–670.

    Google Scholar 

  • Wadsworth, G.J., Redinbaugh, M.G. and Scandalios, J.G. 1988. A procedure for the small-scale isolation of plant RNA suitable for RNA blot analysis. Anal. Biochem. 172: 279–283.

    PubMed  Google Scholar 

  • Wang, M., Ratnam, S. and Freisheim, J.H. 1995. Cloning, nu-cleotide sequence and expression of the bifunctional dihydrofo-late reductase-thymidylate synthase from Glycine max. Biochim. Biophys. Acta. 1261: 325–336.

    PubMed  Google Scholar 

  • Wrobel, R.L., O'Brian, G.R. and Boston, R.S. 1997. Comparative analysis of BiP gene expression in maize endosperm. Gene 204: 105–113.

    PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cox, K., Robertson, D. & Fites, R. Mapping and expression of a bifunctional thymidylate synthase, dihydrofolate reductase gene from maize. Plant Mol Biol 41, 733–739 (1999). https://doi.org/10.1023/A:1006324328355

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1006324328355

Navigation