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. 1997 Oct;63(10):4010–4014. doi: 10.1128/aem.63.10.4010-4014.1997

Gene sequence and expression of an analog of proliferating cell nuclear antigen (PCNA) in the alga Tetraselmis chui and detection of the encoded protein with anti-rat PCNA monoclonal antibody.

L C Cheng 1, S P Hwang 1, J Chang 1
PMCID: PMC168713  PMID: 9327566

Abstract

To identify a phytoplankton cell cycle marker detected by a monoclonal antibody against mammalian proliferating cell nuclear antigen (PCNA) (S. Lin, J. Chang, and E. J. Carpenter, J. Phycol. 30:449-456, 1994), a PCNA gene fragment was isolated by reverse transcription-PCR from the marine unicellular alga Tetraselmis chui Butcher (Prasinophyceae). The gene fragment was 616 bp in length and contained an open reading frame of 205 amino acids. The deduced amino acid sequence showed 80 and 88% similarity to human and rice PCNA, respectively. Southern hybridization indicated that the isolated gene fragment was part of the T. chui genome, with up to three copies in each haploid nucleus. Northern hybridization was used to detect a PCNA mRNA with a size of 1.2 kb from an exponentially growing algal culture. The T. chui gene fragment has been cloned into an expression vector, and a fusion protein was subsequently generated. Anti-rat PCNA simultaneously recognized the PCNA fusion protein and a single 33-kDa band in T.chui total protein extract. Our results indicate that T. chui PCNA is highly similar to its mammalian counterpart and that anti-rat PCNA is a good tool for detecting phytoplankton PCNA in general.

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Selected References

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  1. Achhammer G., Winkler A., Angerer B., Holler E. DNA polymerase delta of Physarum polycephalum. Curr Genet. 1995 Nov;28(6):534–545. doi: 10.1007/BF00518166. [DOI] [PubMed] [Google Scholar]
  2. Ball K. L., Lane D. P. Human and plant proliferating-cell nuclear antigen have a highly conserved binding site for the p53-inducible gene product p21WAF1. Eur J Biochem. 1996 May 1;237(3):854–861. doi: 10.1111/j.1432-1033.1996.0854p.x. [DOI] [PubMed] [Google Scholar]
  3. Bauer G. A., Burgers P. M. Molecular cloning, structure and expression of the yeast proliferating cell nuclear antigen gene. Nucleic Acids Res. 1990 Jan 25;18(2):261–265. doi: 10.1093/nar/18.2.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Celis J. E., Celis A. Cell cycle-dependent variations in the distribution of the nuclear protein cyclin proliferating cell nuclear antigen in cultured cells: subdivision of S phase. Proc Natl Acad Sci U S A. 1985 May;82(10):3262–3266. doi: 10.1073/pnas.82.10.3262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Celis J. E., Madsen P., Celis A., Nielsen H. V., Gesser B. Cyclin (PCNA, auxiliary protein of DNA polymerase delta) is a central component of the pathway(s) leading to DNA replication and cell division. FEBS Lett. 1987 Aug 10;220(1):1–7. doi: 10.1016/0014-5793(87)80865-7. [DOI] [PubMed] [Google Scholar]
  6. Chomczynski P. One-hour downward alkaline capillary transfer for blotting of DNA and RNA. Anal Biochem. 1992 Feb 14;201(1):134–139. doi: 10.1016/0003-2697(92)90185-a. [DOI] [PubMed] [Google Scholar]
  7. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  8. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. GUILLARD R. R., RYTHER J. H. Studies of marine planktonic diatoms. I. Cyclotella nana Hustedt, and Detonula confervacea (cleve) Gran. Can J Microbiol. 1962 Apr;8:229–239. doi: 10.1139/m62-029. [DOI] [PubMed] [Google Scholar]
  10. Hwang S. P., Partin J. S., Lennarz W. J. Characterization of a homolog of human bone morphogenetic protein 1 in the embryo of the sea urchin, Strongylocentrotus purpuratus. Development. 1994 Mar;120(3):559–568. doi: 10.1242/dev.120.3.559. [DOI] [PubMed] [Google Scholar]
  11. Laquel P., Litvak S., Castroviejo M. Mammalian proliferating cell nuclear antigen stimulates the processivity of two wheat embryo DNA polymerases. Plant Physiol. 1993 May;102(1):107–114. doi: 10.1104/pp.102.1.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. López I., Khan S., Vázquez-Ramos J., Hussey P. J. Molecular cloning of a maize cDNA clone encoding a putative proliferating cell nuclear antigen. Biochim Biophys Acta. 1995 Jan 2;1260(1):119–121. doi: 10.1016/0167-4781(94)00192-6. [DOI] [PubMed] [Google Scholar]
  13. Matsumoto K., Moriuchi T., Koji T., Nakane P. K. Molecular cloning of cDNA coding for rat proliferating cell nuclear antigen (PCNA)/cyclin. EMBO J. 1987 Mar;6(3):637–642. doi: 10.1002/j.1460-2075.1987.tb04802.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Matsumoto T., Hata S., Suzuka I., Hashimoto J. Expression of functional proliferating-cell nuclear antigen from rice (Oryza sativa) in Escherichia coli. Activity in association with human DNA polymerase delta. Eur J Biochem. 1994 Jul 1;223(1):179–187. doi: 10.1111/j.1432-1033.1994.tb18981.x. [DOI] [PubMed] [Google Scholar]
  15. Ng L., Prelich G., Anderson C. W., Stillman B., Fisher P. A. Drosophila proliferating cell nuclear antigen. Structural and functional homology with its mammalian counterpart. J Biol Chem. 1990 Jul 15;265(20):11948–11954. [PubMed] [Google Scholar]
  16. Ogata K., Ogata Y., Nakamura R. M., Tan E. M. Purification and N-terminal amino acid sequence of proliferating cell nuclear antigen (PCNA)/cyclin and development of ELISA for anti-PCNA antibodies. J Immunol. 1985 Oct;135(4):2623–2627. [PubMed] [Google Scholar]
  17. Shiina H., Igawa M., Nagami H., Yagi H., Urakami S., Yoneda T., Shirakawa H., Ishibe T., Kawanishi M. Immunohistochemical analysis of proliferating cell nuclear antigen, p53 protein and nm23 protein, and nuclear DNA content in transitional cell carcinoma of the bladder. Cancer. 1996 Oct 15;78(8):1762–1774. doi: 10.1002/(sici)1097-0142(19961015)78:8<1762::aid-cncr17>3.0.co;2-w. [DOI] [PubMed] [Google Scholar]
  18. Suzuka I., Daidoji H., Matsuoka M., Kadowaki K., Takasaki Y., Nakane P. K., Moriuchi T. Gene for proliferating-cell nuclear antigen (DNA polymerase delta auxiliary protein) is present in both mammalian and higher plant genomes. Proc Natl Acad Sci U S A. 1989 May;86(9):3189–3193. doi: 10.1073/pnas.86.9.3189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Vaulot D., Marie D., Olson R. J., Chisholm S. W. Growth of prochlorococcus, a photosynthetic prokaryote, in the equatorial pacific ocean. Science. 1995 Jun 9;268(5216):1480–1482. doi: 10.1126/science.268.5216.1480. [DOI] [PubMed] [Google Scholar]
  20. Warbrick E., Lane D. P., Glover D. M., Cox L. S. A small peptide inhibitor of DNA replication defines the site of interaction between the cyclin-dependent kinase inhibitor p21WAF1 and proliferating cell nuclear antigen. Curr Biol. 1995 Mar 1;5(3):275–282. doi: 10.1016/s0960-9822(95)00058-3. [DOI] [PubMed] [Google Scholar]
  21. Waseem N. H., Lane D. P. Monoclonal antibody analysis of the proliferating cell nuclear antigen (PCNA). Structural conservation and the detection of a nucleolar form. J Cell Sci. 1990 May;96(Pt 1):121–129. doi: 10.1242/jcs.96.1.121. [DOI] [PubMed] [Google Scholar]
  22. Wyman C., Botchan M. DNA replication. A familiar ring to DNA polymerase processivity. Curr Biol. 1995 Apr 1;5(4):334–337. doi: 10.1016/s0960-9822(95)00065-0. [DOI] [PubMed] [Google Scholar]

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