Skip to main content
SpringerLink
Log in

Abundance and state of phosphorylation of the retinoblastoma susceptibility gene product in human colon cancer

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

In an effort to understand the possible role of Rb in cellular growth control, we have investigated the abundance and the state of phosphorylation of Rb protein (pRb) in normal and colon tumor cell lines as well as in matched colon tumors, adenomas and adjoining normal colonic mucosa. Resting normal human fibroblast cell lines were found to have only unphosphorylated pRb and phosphorylation of pRb occurred when the cells entered G1-S phase. In general, the colon tumor tissues had atleast 1.5–2.0 fold increase in the abundance of pRb and 1.5–2.5 fold increase in the percentage of its phosphorylation as compared to the corresponding normal colonic mucosa. Whereas, the adenomas had similar pRb level and its phosphorylation status as observed in the normal colonic mucosa. The actively growing tumor cell lines had approximately two fold higher total pRb than normal cell lines. Although, the percentage of phosphorylated form in growing tumor cell lines as well as normal cell lines were almost equal, it was still considerably higher than normal colonic mucosa. Moreover, DNA binding assay revealed reduced binding affinity of pRb from colon tumor cell line SW480 as compared to the normal cell line W138. These results suggest that the abundance of pRb and its phosphorylation level may have a role in the cellular growth control in human colonic epithelium.

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

Access this article

Log in via an institution

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

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

pRb:

Retinoblastoma Protein

FBS:

Fetal Bovine Serum

DMEM:

Dulbecco's Modified Eagle's Medium

TBS:

Tris Buffered Saline

PBS:

Phosphate Buffered Saline

References

  1. Knudson AG, Jr: Mutation and cancer. Proc Natl Acad Sci USA 68: 820–823, 1971

    Article  PubMed  Google Scholar 

  2. cavanee WK, Dryja TP, Phillips RA, Benedict WF, Gobout R, Gallie BL, Murphree AL, Strong LC, White RL: Expression of recessive alleles by chromosomal mechanisms in retinoblastoma. Nature 305: 779–784, 1983

    Article  Google Scholar 

  3. Dryja TP, Rapaport JM, Joyce JM, Peterson RA: Molecular detection of deletions involving band Q14 of chromosome 13 in retinoblastoma. Proc Natl Acad Sci USA 83: 7391–7394, 1986

    Article  PubMed  CAS  Google Scholar 

  4. Friend SH, Bernards R, Rogelj S, Weinberg RA, Rappaport JM, Albert DM, Dryja TP: A human DNA segment with properties of the gene that predisposes to retinoblastoma and osteosarcoma. Nature 323: 643–646, 1986

    Article  PubMed  CAS  Google Scholar 

  5. Lee WH, Bookstein R, Hong F, Young LJ, Shew JY, Lee EYHP: Human retinoblastoma susceptibility gene: Cloning, identification and sequence. Science 235: 1394–1399, 1987a

    PubMed  CAS  Google Scholar 

  6. Fung YKT, Murphree AL, T'Ang A, Qian J, Hinrics SH, Benedict WF: Structural evidence for the authencity of the human retinoblastoma gene. Science 236: 1657–1661, 1987

    PubMed  CAS  Google Scholar 

  7. Toguchida J, Ishizaki K, Sasaki MS, Nakamura Y, Ikenaga M, Kato M, Sugimoto M, Kotoura Y, Yamamuro T: Preferential mutation of paternally derived RB gene as the initial event in sporadic osteosarcoma. Nature 338: 156–158, 1989

    Article  PubMed  CAS  Google Scholar 

  8. Harbour JW, Lai SL, Whang-Peng J, Gazdar AF, Minna JD, Kaye FJ: Abnormalities in structure and expression of the human retinoblastoma gene in SCLC. Science 241: 353–357, 1988

    PubMed  CAS  Google Scholar 

  9. Yokota J, Akiyama T, Fung YKT, Benedict WF, Namba Y, Hanaoka M, Wada M, Terasaki T, Shimosato Y, Sugimura T, Terada M: Altered expression of the retinoblastoma (RB) gene in small cell carcinoma of the lung. Oncogene 3: 471–475, 1988

    PubMed  CAS  Google Scholar 

  10. Lee EYHP, To H, Shew JY, Bookstein R, Scully P, Lee WH: Inactivation of the retinoblastoms susceptibility gene in human breast cancers. Science 241: 218–221, 1988

    PubMed  CAS  Google Scholar 

  11. T'Ang A, Varley JM, Chakraborty S, Murphree AL, Fung YKT: Structural rearrangement of the retinoblastoma gene in human breast carcinoma. Science 242: 263–266, 1988

    PubMed  Google Scholar 

  12. Horowitz JM, Yandell DW, Park SH, Canning S, Whyte P, Buchkovich KJ, Harlow E, Weinberg RA, Dryja TP: Point mutational inactivation of the Retinoblastoma antioncogene. Science 243: 937–940, 1989

    PubMed  CAS  Google Scholar 

  13. Dyson N, Howley PM, Munger K, Harlow E: The human papillomavirus-16 E7 oncoprotein is able to bind to the retinoblastoma gene product. Science 242: 934–937, 1989

    Google Scholar 

  14. Huang HJS, Yee JK, Shew JY, Chen PL, Bookstein R, Friedman T, Lee EYHP, Lee WH: Suppression of the neoplastic phenotype by replacement of the RB gene in human cancer cells. Science 242: 1563–1566, 1988

    PubMed  CAS  Google Scholar 

  15. Bookstein R, Shew JY, Chen PL, Scully P, Lee WH: Suppression of tumorogenicity of human prostate carcinoma cells by replacing a mutated RB gene. Science 247: 712–715, 1990

    PubMed  CAS  Google Scholar 

  16. Whyte P, Buchkovich KJ, Horowitz JM, Friend SH, Raybuck M, Weinberg RA, Harlow E. Association between an oncogene and an antioncogene: the adenovirus E1A proteins bind to the retinoblastoma gene products. Nature 334: 124–129, 1988

    Article  PubMed  CAS  Google Scholar 

  17. DeCaprio JA, Ludlow JW, Figge J, Shew JY, Huang CM, Lee WH, Marsilio E, Paucha E, Livingston DM: SV40 large tumor antigen forms a specific complex with the product of the retinoblastoma susceptibility gene. Cell 54: 275–282, 1988

    Article  PubMed  CAS  Google Scholar 

  18. Ludlow JW, DeCaprio JA, Huang CM, Lee WH, Paucha E, Livingston DM: SV40 large T antigen binds preferentially to an underphosphorylated member of the retinoblastoma susceptibility gene product family. Cell 56: 57–65, 1989

    Article  PubMed  CAS  Google Scholar 

  19. Egan C, Bayley ST, Branlon PE: Binding of the Rb1 protein to E1A product is required for adenovirus transformation. Oncogene 4: 383–388, 1989

    PubMed  CAS  Google Scholar 

  20. Whyte P, Williamson NM, Harlow E: Cellular targets for transformation by the E1A proteins. Cell 56: 67–75, 1989

    Article  PubMed  CAS  Google Scholar 

  21. DeCaprio JA, Ludlow JW, Lynch D, Furukawa Y, Griffin JD, Piwnica-Worms H, Huang CM, Livingston DM: The product of the retinoblastoma susceptibility gene has properties of a cell cycle regulatory element. Cell 58: 1085–1095, 1989

    Article  PubMed  CAS  Google Scholar 

  22. Buchkovich K, Duffy LA, Harlow E. The retinoblastoma protein is phosphorylated during specific phases of the cell cycle. Cell 58: 1097–1105, 1989

    Article  PubMed  CAS  Google Scholar 

  23. Chen PL, Scully P, Shew JY, Wang JYJ, Lee WH: Phosphorylation of the retinoblastoma gene product is modulated during the cell cycle and cellular differentiation. Cell 58: 1193–1198, 1989

    Article  PubMed  CAS  Google Scholar 

  24. Mihara K, Cao XR, Yen A, Chandler S, Driscoll B, Murphree AL, T'Ang A, Fung YKT: Cell cycle-dependent regulation of phosphorylation of the human retinoblastoma gene product. Science 246: 1300–1303, 1989

    PubMed  CAS  Google Scholar 

  25. Robins PD, Horowitz JM, Mulligan RC: Negative regulation of human c-fos expression by the retinoblastom gene product. Nature 346: 668–671, 1990

    Article  Google Scholar 

  26. Lee WH, Shew JY, Hong FD, Sery TW, Donoso LA, Young LJ, Bookstein R, Lee EHYP: The retinoblastoma susceptibility gene encodes a nuclear phosphoprotein associated with DNA binding activity. Nature 329: 642–645, 1987(b)

    Article  PubMed  CAS  Google Scholar 

  27. Bernards R, Schackleford GM, Gerber MR, Horowitz JM, Friend SH, Schartl M, Bogenmann E, Rapaport JM, McGee T, Dryja TP, Weinberg RA: Structure and expression of the murine retinoblastoma gene and characterization of its encoded protein. Proc Natl Acad Sci USA 86: 6474–6478, 1989

    Article  PubMed  CAS  Google Scholar 

  28. Grand RJA, Byrd PJ, Grabham PW, Gregory CD, Huen DS, Merrick RM, Young LS, Gallimore PH: The expression of the retinoblastoma gene product Rb1 in primary and adenovirus-transformed human cells. Oncogene 4: 1291–1298, 1989

    PubMed  CAS  Google Scholar 

  29. Gope R, Christensen MA, Thorson A, Lynch HT, Smyrk T, Hodgson C, Wildrick DM, Gope ML: Increased expression of retinoblastoma gene in human colorectal carcinomas relative to normal colonic mucosa. J Natl Can Inst 82: 310–314, 1990

    CAS  Google Scholar 

  30. Gope R, Christensen MA, Thorson A, Lynch HT, Smyrk T, Gope ML, Boman BM: Molecular analysis of retinoblastoma (Rb) gene in human colorectal carcinomas. In: J Utsunomiya, HT Lynch (eds.) Hereditary Colorectal Cancer, Springer-Verlag, Tokyo, 1990, pp 489–496

    Google Scholar 

  31. Xu HG, Hu SX, Hashimoto T, Takahashi R, Benedict WF: The retinoblastoma susceptibility gene product: a characteristic pattern in normal cells and abnormal expression in malignant cells. Oncogene 4: 807–812, 1989

    PubMed  CAS  Google Scholar 

  32. Bombik BM, Baserga R: Increased RNA synthesis in nuclear monolayers of W138 cells stimulated to proliferate. Proc Natl Acad Sci USA 71: 2038–2042, 1974

    Article  PubMed  CAS  Google Scholar 

  33. Horowitz JM, Park SH, Bogenmann E, Cheng JC, Yandell DW, Kaye FJ, Minna JD, Dryja TP, Weinberg RA: Frequent inactivation of the retinoblastoma anti-oncogene is restricted to a subset of human tumor cells. Proc Natl Acad Sci USA 87: 2775–2779, 1990

    Article  PubMed  CAS  Google Scholar 

  34. Furukawa Y, DeCaprio JA, Freedman A, Kanakura Y, Nakamura M, Ernst TJ, Livingston DM, Griffin JD: Expression and state of phosphorylation of the retinoblastoma susceptibility gene product in cycling and noncycling human hematopoietic cells. Proc Natl Acad Sci USA 87: 2770–2774, 1990

    Article  PubMed  CAS  Google Scholar 

  35. Cooper JA, Whyte P: Rb and the cell cycle: entrance or exit? Cell 58: 1009–1011, 1989

    Article  PubMed  CAS  Google Scholar 

  36. Laiho M, DeCaprio JA, Ludlow JW, Livingston DM, Massaague J: Growth inhibition by TGF-beta linked to suppression of retinoblastoma protein phosphorylation. Cell 62: 175–185, 1990

    Article  PubMed  CAS  Google Scholar 

  37. Moritsuka T, Kobori O, Morioka Y, Watanabe S, Yamaguchi N, Sugano S: Expression level of the retinoblastoma susceptibility gene is elevated in simian virus 40-transformed cells. Oncogene Res 5: 155–159, 1989

    PubMed  CAS  Google Scholar 

  38. Tuck SP, Crawford L: Overexpression of normal human p53 in established fibroblasts leads to their tumrigeneic conversion. Oncogene Res 4: 81–96, 1989

    PubMed  CAS  Google Scholar 

  39. Grinnel RS, Lane A: Benign and adenomatous polyps and papillary adenomas of colon and rectum—Analysis of 1856 tumors in 1355 patients. Int Abstr Surg 106: 519–538, 1958

    Google Scholar 

  40. Morson BC: Factors influencing the prognosis of the early cancer of the rectum. Proc R Soc Med 59: 607–608, 1966

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Creighton Cancer Center, Creighton University, 24th at California Street, Criss III, Rm #358, 68178, Omaha, NE, USA

    Rajalakshmi Gope & Mohan L. Gope

Authors
  1. Rajalakshmi Gope
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mohan L. Gope
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Gope, R., Gope, M.L. Abundance and state of phosphorylation of the retinoblastoma susceptibility gene product in human colon cancer. Mol Cell Biochem 110, 123–133 (1992). https://doi.org/10.1007/BF02454189

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF02454189

Key words

Search

Navigation