Key Cell Cycle Regulatory Genes Research Articles

Proton pump inhibitors reduce cell cycle abnormalities in Barrett's esophagus.

Neoplastic progression in Barrett's esophagus is a multi-step process in which the metaplastic columnar epithelium sequentially evolves through a metaplasia-dysplasia-carcinoma sequence. The expression and DNA copy number of key cell cycle regulatory genes in paired normal and Barrett's esophagus samples was evaluated. Protein levels were evaluated in 60 formalin-fixed, paraffin-embedded human tissues by immunohistochemistry. DNA copy number from 20 fresh tissue pairs was analysed by Southern blot analysis. All normal mucosal samples expressed the p27(kip1) protein, but did not display appreciable nuclear staining for p16(kip4), p21(cip1) or cyclins D1 and E. Barrett's metaplastic specimens displayed increased expression levels of p16(kip4) (74%), p21(cip1) (89%) and cyclins D1 (43%) and E (37%). p27 protein was absent in three cases. There was a significant correlation between the expression of p16(kip4) and cyclin E, and p21(cip1) and p27(kip4) with cyclin D1. DNA analysis did not reveal any amplification or deletion of these genes. Acid suppression, however, was associated with significantly lower expression levels of key cell cycle proteins. Increased expression of key cell cycle regulatory genes appears to occur early in the neoplastic progression associated with Barrett's esophagus. Treatment with proton pump inhibitors appears to alter this increased expression.

Inactivation of both Rb and p53 pathways in mouse lung epithelial cell lines.

Aberrant expression of key cell cycle regulatory genes is essential for the immortalization and transformation of cells in vitro. We examined 20 mouse lung epithelial cell lines (2 nontumorigenic, 5 nonmetastatic, and 13 metastatic) for mutations or alterations in the expression of key components of the Rb pathway (pRb and p16INK4a) and the p53 pathway (p53 and p19ARF). Seven cell lines had a mutation in exons 5 to 8 of p53. p19ARF was inactivated in the remaining 13 cell lines, primarily by homozygous deletion. Rb expression was present and unaltered in all cell lines, with both phosphorylated and unphosphorylated protein forms detectable. p16INK4a transcripts were undetectable in all cell lines tested except LM1. Loss of p16INK4a expression was a result of homozygous deletion in 11 out of 20 lung cell lines and promoter-exon 1 hypermethylation in 6 out of the remaining 8 cell lines. Other related components that were examined in this study included p21WAF1 and cyclin D1. Compared to normal lung tissue, p21WAF1 expression levels were reduced or undetectable in all cell lines, which did not correlate with loss of p53 function, but did correlate with inactivation of either p53 or p19ARF. Although cyclin D1 expression was variable between cell lines, transcript levels were decreased by at least 50% in the nontumorigenic lines C10 and E10 compared to the tumorigenic cell lines. These results demonstrate mutually exclusive relationships between p53 and p19ARF and between Rb and p16INK4a, but perhaps not between cyclin D1 and p16INK4a, and further describe the nature of involvement of both pathways in mouse lung tumorigenesis.

The peri-cell-cycle in Arabidopsis.

The root systems of plants proliferate via de novo formed meristems originating from differentiated pericycle cells. The identity of putative signals responsible for triggering some of the pericycle cells to re-enter the cell cycle remains unknown. Here, the cell cycle regulation in the pericycle of seedling roots of Arabidopsis thaliana (L.) HEYNH: is studied shortly after germination using various strategies. Based on the detailed analysis of the promoter-beta-glucuronidase activity of four key cell cycle regulatory genes, combined with cell length measurements, microdensitometry of DNA content, and experiments with a cell cycle-blocking agent, a model is proposed for cell cycle regulation in the pericycle at the onset of lateral root initiation. The results clearly show that before the first lateral root is initiated, the pericycle consists of dissimilar cell files in respect of their cell division history. Depending on the distance behind the root tip and on position in relation to the vascular tissue, particular pericycle cells remain in the G(2) phase of the cell cycle and are apparently more susceptible to lateral root initiation than others.

INACTIVATION OF BOTH Rb AND p53 PATHWAYS IN MOUSE LUNG EPITHELIAL CELL LINES

Aberrant expression of key cell cycle regulatory genes is essential for the immortalization and transformation of cells in vitro. We examined 20 mouse lung epithelial cell lines (2 nontumorigenic, 5 nonmetastatic, and 13 metastatic)for mutations or alterations in the expression of key components of the Rb pathway (pRb and p16INK4a) and the p53 pathway (p53 and p19ARF). Seven cell lines had a mutation in exons 5 to 8 of p53. p19ARF was inactivated in the remaining 13 cell lines, primarily by homozygous deletion. Rb expression was present and unaltered in all cell lines, with both phosphorylated and unphosphorylated protein forms detectable. p16INK4a transcripts were undetectable in all cell lines tested except LM1. Loss of p16INK4a expression was a result of homozygous deletion in 11 out of 20 lung cell lines and promoter-exon 1 hypermethylation in 6 out of the remaining 8 cell lines. Other related components that were examined in this study included p21WAF1 and cyclin D1. Compared to normal lung tissue, p21WAF1 expression levels were reduced or undetectable in all cell lines, which did not correlate with loss of p53 function, but did correlate with inactivation of either p53 or p19ARF. Although cyclin D1 expression was variable between cell lines, transcript levels were decreased by at least 50% in the nontumorigenic lines C10 and E10 compared to the tumorigenic cell lines. These results demonstrate mutually exclusive relationships between p53 and p19ARF and between Rb and p16INK4a, but perhaps not between cyclin D1 and p16INK4a, and further describe the nature of involvement of both pathways in mouse lung tumorigenesis.

Viral oncogenesis and cell cycle control

Increasing evidence suggests a critical role for cell cycle regulatory genes in tumorigenesis in mammals, and oncogenic viruses account for a considerable proportion of cancers in humans. In most cases, infection by oncogenic viruses is insufficient for malignant transformation; rather, the viruses provide host cells with additional growth stimuli, which extend the proliferative capacity of the infected cell. This implies that viral oncogenes can override cellular control mechanisms, which in untransformed cells regulate cell cycle progression in response to various antiproliferative signals. Recent progress has enabled the definition of several mammalian cell cycle regulatory genes as targets for viral oncoproteins. This new field of investigation was the central topic of a meeting (Viral Oncogenesis and Cell Cycle Control) held in Heidelberg in October 1995. In the following, major findings presented at this meeting will be summarized. The first part of the meeting was dedicated to the analysis of cell cycle regulation in normal untransformed mammalian cells, followed by an analysis of the interaction of tumor virus oncoproteins with mammalian cell cycle regulators. The latter interactions fall into three categories: viral oncoproteins were shown to interfere with various signal transduction pathways, to inactivate growth-suppressive nuclear proteins by direct binding, and to modulate the expression of key cell cycle regulatory genes.

Dissection of the Genetic Programs of p53-Mediated G1 Growth Arrest and Apoptosis: Blocking p53-Induced Apoptosis Unmasks G1 Arrest

Employing the myeloblastic leukemia M1 cell line, which does not express endogenous p53, and genetically engineered variants, it was recently shown that activation of p53, using a p53 temperature-sensitive mutant transgene (p53ts), resulted in rapid apoptosis that was delayed by high level ectopic expression of bcl-2. In this report, advantage has been taken of these M1 variants to investigate the relationship between p53-mediated G1 arrest and apoptosis. Flow cytometric cell cycle analysis has provided evidence that activation of wild-type (wt) p53 function in M1 cells resulted in the induction of G1 growth arrest; this was clearly seen in the M1p53/bcl-2 cells because of the delay in apoptosis that unmasked p53-induced G1 growth arrest. This finding was further corroborated at the molecular level by analysis of the expression and function of key cell cycle regulatory genes in M1p53 versus M1p53/bcl-2 cells after the activation of wt p53 function; events that take place at early times during the p53-induced G1 arrest occur in both the M1p53 and the M1p53/bcl-2 cells, whereas later events occur only in the M1p53/bcl-2 cells, which undergo delayed apoptosis, thereby allowing the cells to complete G1 arrest. Finally, it was observed that a spectrum of p53 target genes implicated in p53-induced growth suppression and apoptosis were similarly regulated, either induced (gadd45, waf1, mdm2, and bax) or suppressed (c-myc and bcl-2), after activation of wt p53 function in M1p53 and M1p53/bcl-2 cells. Taken together, these findings show that wt p53 can simultaneously induce the genetic programs of both G1 growth arrest and apoptosis within the same cell type, in which the genetic program of cell death can proceed in either G1-arrested (M1p53/bcl-2) or cycling (M1p53) cells. These findings increase our understanding of the functions of p53 as a tumor suppressor and how alterations in these functions could contribute to malignancy.