Growth state-dependent regulation of plasminogen activator inhibitor type-1 gene expression during epithelial cell stimulation by serum and transforming growth factor-beta1.
Transcription of the plasminogen activator inhibitor type-1 (PAI-1) gene appears to be growth state regulated in several cell types (e.g. , Ryan and Higgins, 1993, J Cell Physiol 155:376-384; Mu et al., 1998, J Cell Physiol 174:90-98). Transit of serum-stimulated normal rat kidney (NRK) epthelial cells through the first division cycle after release from quiescence (G(0)) provided a model system to assess the kinetics and mechanisms underlying PAI-1 expression in a growth "activated" phenotype. PAI-1 mRNA transcripts increased by more than 20-fold during the G(0)-->G(1) transition; induced expression had immediate-early response characteristics and abruptly declined prior to the onset of DNA synthesis. Transcriptional activity of the PAI-1 gene paralleled the steady-state mRNA abundance profile during this first synchronized growth cycle after release from quiescence. Although PAI-1 mRNA levels were up-regulated (approximately threefold) upon exposure to several different growth factors, neutralizing antibodies to transforming growth factor-beta1 (TGF-beta1) effectively attenuated the more than ninefold serum-associated PAI-1 inductive response by more than 70% (at both the mRNA transcript and protein levels). Similar to the metabolic requirements for serum-mediated PAI-1 transcription, PAI-1 induction upon addition of TGF-beta1 to quiescent NRK cell cultures was actinomycin D sensitive and resistant to cyclohexamide and puromycin, suggesting a primary mode of transcript control. The response to protein synthesis inhibitors, however, was complex. While cyclohexamide appeared to stabilize, or at least maintain, fetal bovine serum (FBS)- or TGF-beta1-stimulated PAI-1 mRNA levels, puromycin had no such affect. The amplitude and duration of induced PAI-1 expression were the same in either the presence or absence of puromycin. Cyclohexamide when used alone (i.e., in non-FBS- or TGF-beta1-treated cultures), moreover, effectively stimulated PAI-1 induction whereas puromycin was ineffective. Although TGF-beta1 was not a complete mitogen in the NRK cell system, incubation of quiescent renal cell cultures with TGF-beta1, prior to serum stimulation, resulted in a 10- to 12-fold increase in PAI-1 expression coincident with exit out of G(0). These data support a model in which PAI-1 gene expression is closely associated with creation of the growth-activated state and that cell cycle controls appear to be superimposed on the time course of the serum-induced expression of the PAI-1 gene.
Boehm
J R
JR
Program in Cell and Molecular Biology, Albany Medical College, Albany, New York 12208, USA.
Kutz
S M
SM
Sage
E H
EH
Staiano-Coico
L
L
Higgins
P J
PJ
D000818:Animals; D001790:Blood Physiological Phenomena; D002417:Cattle; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004847:Epithelial Cells / Q000187:drug effects*; D005786:Gene Expression Regulation / Q000187:drug effects*; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics*; D051381:Rats; D013268:Stimulation, Chemical; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D017395:Plasminogen Activator Inhibitor 1; D016212:Transforming Growth Factor beta