Opinions of Zagreb University students on professor-student dating.

AIM: To investigate what Zagreb University students think about professor-student dating. METHODS: We surveyed 500 students from ten different Zagreb University schools using anonymous questionnaires. Twenty-five female and 25 male students from each school were asked to assess professor-student dating as either acceptable, reckless, unethical, or shameful, and decide how severely such behavior should be punished, if it should be punished at all. Sex of the professor and student was irrelevant. RESULTS: Students were equally strict towards the student and the professor. They judged professor-student dating more strictly if dating started before the student passed the exam in the subject taught by the professor than if it started afterwards. More female than male students had no opinion on whether the professor should be punished. When dating started after the student passed the exam, male students judged the student's behavior more strictly than female students; if the dating started before, the Economics students were stricter and the Music Academy students milder than average in their judgements. In the case when dating started after the student passed the exam, students of the Faculty of Physical Education judged both dating parties more strictly, Political Sciences students were milder toward a professor, whereas more students of the Faculty of Philosophy thought that it should be punished. Most students with no opinion on punishing dating were found among Agronomy students. CONCLUSION: Most Zagreb University students think that professor-student dating should not be punished and that both dating parties take equal responsibility for such behavior.

TGFalpha is required for full expression of the transformed growth phenotype of NIH 3T3 cells overexpressing ornithine decarboxylase.

Ornithine decarboxylase (ODC) overexpressed from a heterologous promoter drives the tumorigenic transformation of NIH 3T3 cells and provides a model to investigate the underlying molecular mechanisms. These transformed cells, designated NODC cells, exhibit elevated levels of epidermal growth factor receptor (EGFR) tyrosine kinase (Tyr-k) activity relative to control transfected cells and inhibition of EGFR Tyr-k activation suppresses the transformed growth phenotype of these cells. Thus, ODC-induced transformation of NIH 3T3 cells appears to be mediated, at least in part, by enhanced signaling through the EGFR pathway. Here we extend these studies by evaluating: (i) the effects on growth regulation of overexpressing ODC in EGFR-deficient NIH 3T3 cells; (ii) the potential role of TGFalpha in mediating the EGFR-dependent transformation of NIH 3T3 cells by ODC. Disruption of EGFR-TGFalpha interactions either by deleting EGFR, by treatment with anti-TGFalpha neutralizing antibody or by transfection with a TGFalpha antisense expression vector suppressed acquisition of the full transformed growth phenotype. Specifically, the loss of contact inhibition and the capacity for clonogenic growth appear more dependent on EGFR-TGFalpha interactions than anchorage-independent growth in ODC-overexpressing cells. ODC overexpression does not alter the amount, localization or secretion of TGFalpha. Thus, TGFalpha is not the ODC-responsive component of the EGFR signaling pathway but appears to be critically involved in development of the transformed phenotype of NODC cells.

Thymosin alpha 1 does not promote growth or oncogenic transformation.

Thymosin alpha 1 (T alpha 1) is an immune modulatory peptide which has been evaluated in a variety of clinical trials. Although no in vivo adverse effects, including enhancement of tumor growth, have been noted, in vitro studies suggesting a role for T alpha 1 in cell growth have been reported. The studies presented in this report evaluated both exogenously added T alpha 1 and endogenously expressed T alpha 1 as factors which could either promote growth of tumor cells or induce transformation. No effect of exogenous T alpha 1 on cell growth was found. NIH-3T3 cells transfected with cDNA for the precursor ProThymosin alpha (Pro T alpha) expressed elevated levels of authentic T alpha 1 but did not demonstrate either enhanced proliferation in liquid culture or transformation as defined by the loss of contact inhibition or anchorage independent growth in soft agar. Thus these studies argue against the hypothesis that T alpha 1 is either an intracellular or extracellular growth promoter.

Regulation of ornithine decarboxylase gene expression by the Wilms' tumor suppressor WT1.

The importance of ornithine decarboxylase (ODC) to cell proliferation is underscored by the complex array of cell-specific mechanisms invoked to regulate its synthesis and activity. Misregulation of ODC has severe negative consequences on normal cell function, including the acquisition of tumorigenic growth properties by cells overexpressing ODC. We hypothesize that ODC gene expression is a candidate target for the anti-proliferative function of certain tumor suppressors. Here we show that the Wilms' tumor suppressor WT1 binds to multiple sites within the human ODC promoter, as determined by DNase I protection and methylation interference assays. The expression of WT1 in transfected HCT 116, NIH/3T3 and HepG2 cells represses activity of the ODC promoter controlling expression of a luciferase reporter gene. In contrast WT1 expression enhances ODC promoter activity in SV40-transfected HepG2 cells. Both the extent of modulation of ODC gene expression and the mediating WT1 binding elements are cell specific. Constructs expressing WT1 deletion mutants implicate two regions required for repressor function, as well as an intrinsic activation domain. Understanding the regulation of ODC gene expression by WT1 may provide valuable insights into the roles of both WT1 and ODC in development and tumorigenesis.

Ornithine decarboxylase transformation of NIH/3T3 cells is mediated by altered epidermal growth factor receptor activity.

Ornithine decarboxylase (ODC) has been shown to be oncogenic in transfected NIH/3T3 cells overexpressing the enzyme from a heterologous promoter. These cells, designated as NODC-2 cells, acquire proliferative properties associated with tumorigenic transformation such as loss of contact inhibition, decreased population doubling time, anchorage-independent growth, and tumor production in nude mice. At least one of these parameters, loss of contact inhibition, remains dependent on elevated ODC levels. We have used these cells to investigate the molecular mechanisms by which ODC overexpression drives cell transformation and to examine the involvement of other proto-oncogene products in this process. An interaction between ODC overexpression and the epidermal growth factor receptor (EGF-R) was suggested initially by the elevation of both basal (300%) and ligand-induced (457%) EGF-R tyrosine kinase activities in NODC-2 cells compared to similarly treated control NLK cells. Disruption of EGF-R mediated signal transduction in NODC-2 cells both by treatment with tyrphostin-25 or by transfection with a vector expressing a dominant negative EGF-R mutant resulted in reacquisition of contact-inhibited growth and suppression of anchorage-independent, clonogenic growth in soft agar. We conclude that ODC-induced transformation of NIH/3T3 cells is mediated, at least partly, by alterations in EGF-R signal transduction activity.

Transformation of NIH/3T3 cells by ornithine decarboxylase overexpression.

Ornithine decarboxylase (ODC) plays a rate-limiting role in polyamine biosynthesis and is intimately associated with cell proliferation and function. Although elevated levels of ODC mRNA, protein, and enzyme activity are consistently detected in transformed cells and tumors, the question remains as to whether ODC gene overexpression has a causative role in tumorigenesis. We have stably transfected NIH/3T3 fibroblasts with an expression construct containing human ODC complementary DNA under transcriptional control of the human beta-actin promoter. Cells transfected with the beta-actin/ODC DNA construct, designated NODC cells, and control transfectants, termed NLK cells, were analyzed for ODC gene expression and cell growth characteristics. ODC activity and mRNA levels were elevated 3-6-fold in NODC cells relative to NLK cells. NODC cells, in contrast to NLK control cells, are not contact inhibited, exhibit anchorage-independent growth, cycle more rapidly, and induce tumors in nude mice more efficiently and rapidly. These results directly establish a causative role for the misregulation of ODC gene expression in the acquisition of a transformation phenotype and provide a model to examine the interaction of ODC and other gene products in neoplastic development.

Multiple promoter elements govern expression of the human ornithine decarboxylase gene in colon carcinoma cells.

Overexpression of the ornithine decarboxylase (ODC) gene may be important to the development and maintenance of colonic neoplasms, as well as tumors in general. In this study, we examined the promoter elements governing constitutive expression of the human ODC gene in HCT 116 human colon carcinoma cells and, for comparison, K562 human erythro-leukemia cells. It was determined by functional analysis that the promoter elements responsible reside within the 378 bp immediately upstream from the transcription start site. Within this sequence, there are at least three regions that modulate the efficiency of the ODC promoter cooperatively. Both DNA bandshift and footprint assays demonstrated all three regions to be rich in sites that bind to nuclear proteins isolated from HCT 116 and K562 cells; the protein binding pattern of non-transformed, diploid fibroblasts was found to be much less complex. Several of the protein binding sequences have little or no homology to common regulatory elements. We suggest that the constitutive activity of the ODC gene in HCT 116 colon carcinoma cells, and perhaps transformed cells in general, involves a complex interaction of multiple regulatory sequences and their associated nuclear proteins. Finally, the saturation of the promoter in these transformed cell lines suggests that high levels of protein binding in the ODC promoter may contribute to elevated constitutive expression of this gene.

Isolation and expression of a human ornithine decarboxylase gene.

We have isolated a human ornithine decarboxylase (ODC) gene from a leukocyte genomic DNA library in order to examine the mechanisms involved in the regulation of ODC gene expression in normal and neoplastic cell growth. Nucleotide sequence analysis shows that the human ODC gene in clone ODC709-A2 consists of 12 exons which encode a protein identical to that inferred from a human ODC cDNA sequence. The 5' end of the gene was determined by S1 nuclease and primer extension mapping. The high G + C content and small open reading frame found in exon 1 may be pertinent to translation regulation of ODC. Conserved sequences and potential promoter elements including a TATA box, a possible CCAAT element, SP1 and AP-2 transcription factor binding sites, and cAMP response elements were identified in the 5'-flanking region. Transfection of mouse LM (tk-) cells with ODC709-A2 DNA resulted in the production of human ODC mRNA approximately 2.25 kilobases in length. Evidence that the protein synthesized from the human gene is functional is provided by "rescue" transfection of a Chinese hamster ovary mutant cell line, C55.7, which is ODC-deficient. C55.7 cells transfected with ODC709-A2 DNA expressed ODC enzyme activity and proliferated without exogenous putrescine.

Nucleotide sequence and transcription analysis of the gene coding for subunit III of soybean chloroplast proton-translocating ATPase.

The gene coding for subunit III of CF0-ATPase complex (atpH gene) has been localized on the soybean chloroplast genome by heterologous hybridization with the atpH gene of wheat chloroplast. The gene, 243 bp in length, is located in a 7.5-kb BamHI fragment generated from the 14.8-kb PvuII fragment in the large single-copy region of the chloroplast genome. The nucleotide sequence of the gene has been determined. The deduced amino acid (aa) sequence shows 2 aa changes relative to its homologue from wheat and spinach and 1 aa with respect to that found in tobacco chloroplast. The gene is apparently transcribed independently as an 850-nt transcript.