Summary
Extractable nucleolar proteins from HeLa cells were used as a source of antigen to immunize mice for monoclonal antibody (MAb) production. Ten of the resulting MAbs shown to identify nucleolar phosphoprotein (110 kD/pI 5.5) were purified and used in immunochemical studies to further characterize protein C23. All ten MAbs showed nucleolar localization by indirect immunofluorescence; one antibody (FR2) also showed some nucleoplasmic localization that was attributed to a shared epitope between protein C23 and a 72 kD nuclear/nucleolar antigen. Reciprocal antibody cross blocking studies indicated that the ten MAbs identified nine distinct epitopes on protein C23. Interestingly, seven of the nine epitopes were shown by immunofluorescence and competitive ELISA studies to be species related. Immunostained patterns of exponentially growing HeLa cells suggest that protein C23 exists in vivo solely as a 110 kD peptide. However, protein C23 was subject to rapid degradation into a number of proteolytic fragments upon extraction or storage of isolated nucleoli. The failure to find protein C23 related peptides with molecular sizes less than 110 kD in exponentially growing cells and the lack of cytoplasmic localization of any of the ten MAbs suggests that protein C23 is not a prepro-protein processed in vivo to form ribosomal proteins as previously suggested (1).
Similar content being viewed by others
References
Bugler B, Caizergues-Ferrer M, Bouche G, Burbon H, Amalric F: Eur J Biochem 128:475–480, 1982.
Howell MW: In: (Busch H (Ed), The Cell Nucleus. Academic Press, New York, Vol 11, 1982, pp 89–105.
Hilgers J, Nowinsky RC, Geering G, Hardy W: Cancer Res 32:98–105, 1972.
Howell MW: Chromosoma 62:361–367, 1977.
Hubbell HR, Brown Y, Hsu T: Exp Cell Res 129:139–147, 1980.
Lischwe MA, Smetana K, Olson M, Busch H: Life Sci 25:701–708, 1979.
Freeman JW Chatterjee A, Ross B, Busch RK, Busch H: Proc Am Assoc Cancer Res, 26:1136, 1985.
Lischwe MA, Richards RL, Busch RL, Busch H: Exp Cell Res 136:101–109, 1981.
Olson MOJ, Guetzow K, Busch H: Exp Cell Res 135:259–265, 1981.
Ochs R, Lischwe M, O'Leary P, Busch H: Exp Cell Res 146:139–149, 1983.
Arriglio FE, Lau Y, Spallone A: Cytogenet Cell Genet 26:244–249, 1980.
Kacerovska H, Kovsky ZL, Smetana K: Neoplasma 28:513–516, 1981.
Smetana K, Gyorkey F, Gyorkey P, Busch H: Exp Cell Res 64:133–139, 1971.
Pfeifle J, Anderer FA: Biochem Biophys Res Com 116:106–112, 1983.
Spohn WH, Alm YS, Busch RK, Busch H: Cancer Invest, 1985, in press.
Lischwe MA, Yeoman LC, Busch H: J Cell Biol 99:12a, 1984.
Schroer J, Bender T, Feldmann RJ, Kim KJ: Eur J Immunol 13:693–700, 1983.
Freeman JW, Chatterjee A, Busch H: J Immunol Meth, 78:259–265, 1985.
Primus FJ, Newell KD, Blue A, Goldenberg DM: Cancer Res 43:686–692, 1983.
Nunberg JH, Rogers G, Gilbert JH, Snead RM: Proc Natl Acad Sci 81:3675–3679, 1984.
Thomas PE, Reih LM, Ryan DE, Levin W: J Biol Chem 259:3890–3899, 1984.
Busch H, Smetana K: The Nucleolus. Academic Press, New York, 1970.
Greenwood FC, Hunter WM: Biochem J 89:114–117, 1963.
Busch RK, Daskal I, Spohn WH, Kellermayer M, Busch H: Cancer Res 34:2362–2367, 1974.
Takacs B: In: Immunological Methods. Academic Press, New York, 1979, pp 81–105.
Towbin H, Staehelen T, Gordon J: Proc Natl Acad Sci 76:4350–4354, 1979.
Mamrack MD, Olson MO,Busch H: Biochem 18:3381–3386, 1979.
Olson MO, Guetzow K, Allan R, Busch H: Proc Am Assoc Cancer Res 20:118, 1979.
Tsutsui K, Tsutsui K, Oda T: Eur J Biochem 108:497–505, 1980.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Freeman, J.W., Chatterjee, A., Ross, B.E. et al. Epitope distribution and immunochemical characterization of nucleolar phosphoprotein C23 using ten monoclonal antibodies. Mol Cell Biochem 68, 87–96 (1985). https://doi.org/10.1007/BF00219392
Issue Date:
DOI: https://doi.org/10.1007/BF00219392