Expression of the transforming growth factor-beta gene during growth inhibition following polyamine depletion.
Polyamine depletion and cytokine transforming growth factor-beta (TGF-beta) inhibit cell proliferation. The current study tests the hypothesis that polyamine depletion results in growth inhibition by altering expression of the TGF-beta gene in intestinal epithelial cells. Studies were conducted in the IEC-6 cell line derived from rat small intestinal crypt cells. Cells were grown in DMEM in the presence or absence of alpha-difluoromethylornithine (DFMO), a specific inhibitor of polyamine biosynthesis, for 6 and 12 days. Administration of DFMO not only depleted intracellular polyamines but also significantly increased the mRNA levels of TGF-beta. Increased TGF-beta mRNA in DFMO-treated cells was paralleled by an increase in TGF-beta content. Depletion of intracellular polyamines by DFMO had no effect on the rate of TGF-beta gene transcription, as measured by nuclear run-on assay. The half-life of mRNA for TGF-beta in normal cells was approximately 65 min and increased to >16 h in cells treated with DFMO for 6 or 12 days. Exogenous polyamine, when given together with DFMO, prevented the increased half-life of TGF-beta mRNA in IEC-6 cells. TGF-beta added to the culture medium significantly decreased the rate of DNA synthesis and final cell number in normal and polyamine-deficient cells. Furthermore, growth inhibition caused by polyamine depletion was partially but significantly blocked by addition of immunoneutralizing anti-TGF-beta antibody. These results indicate that 1) depletion of intracellular polyamines induces the activation of the TGF-beta gene through posttranscriptional regulation and 2) increased expression of the TGF-beta gene plays an important role in the process of growth inhibition following polyamine depletion.
Department of Surgery, University of Maryland Medical School and Baltimore Veterans Affairs Medical Center, Baltimore, Maryland 21201, USA.
A R
AR
Patel
J
J
Li
B L
BL
Bass
J Y
JY
Wang
D000818:Animals; D000906:Antibodies; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D000518:Eflornithine / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D006207:Half-Life; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D007422:Intestines; D007700:Kinetics; D011073:Polyamines / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000096:biosynthesis* / Q000235:genetics* / Q000502:physiology
D000906:Antibodies; D011073:Polyamines; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D003609:Dactinomycin; D000518:Eflornithine