Increased Growth of NIH/3T3 Cells by Transfection with Human pi 20 Complementary DNA and Inhibition by a pi 20 Antisense Construct

Laszlo Perlaky, Benigno C. Valdez, Rose K. Busch, Robert G. Larson, Sissy M. Jhiang, Wei Wei Zhang, Michael Brattain, Harris Busch

Research output: Contribution to journalArticlepeer-review

52 Scopus citations

Abstract

The human nucleolar antigen pi 20 was detected with an anti-pi 20 monoclonal antibody in most human malignant tumors but not in most resting human tissues (J. W. Freeman et ai, Cancer Res., 48: 12441251, 1988) and has been used as a prognostic tumor marker in breast cancer patients (J. W. Freeman et al. Cancer Res., 51: 1973-1978, 1991). After the complementary DNA and gene for the human pi 20 protein were isolated and sequenced (review: H. Busch, Cancer Res., 50: 4830-4838, 1990), constructs were prepared to study the expression of the sense pi 20 and its antisense, p021 message. NIH/3T3 cells were transfected by electroporation with pSVX plasmids containing either the pi 20 complementary DNA (pSVX120) or the antisense, p021 DNA (pSVX021), and clones containing these constructs were selected. The expression of pi 20 or p021 in these constructs was regulated by Moloney murine leukemia virus long terminal repeats. In pSVX120-transfected NIH/3T3 cells, the expressed human pi 20 protein was localized to the nucleoli as shown by an ti-pi 20 monoclonal antibody immunofluorescence. Expression of the pi 20 message and protein was confirmed by Northern (mRNA) and Western (protein) blots. Transfection of the pi 20 complementary DNA in sense orientation caused malignant transformation of NIH/3T3 cells in vitro and produced rapidly growing tumors in nude mice. Transfection of the antisense pi 20 constructs markedly delayed the growth of these tumors in vitro and in vivo (L. Perlaky et al., Proc. Am. Assoc. Cancer Res., 32: 1682, 1991). When transformed 3T3/ pSVX120 cells were transfected with an inducible antisense pi 20 construct (pMSG021), dexamethasone induction decreased the growth rate by 62%, and the cell line returned to its normal phenotype. Northern blot analysis showed a decreased level of pi20 mRNA, and the immunofluorescence was also markedly reduced.

Original languageEnglish (US)
Pages (from-to)428-436
Number of pages9
JournalCancer Research
Volume52
Issue number2
StatePublished - Jan 1992

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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