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Biomedical subjects

R J Rooney

Publications and source records attributed to R J Rooney.

At least 19 recordsLinked to original sources
PubMed2001

Cell cycle attenuation by p120E4F is accompanied by increased mitotic dysfunction.

In addition to their well-established roles at the G1-S checkpoint, recent reports support a role for universal cyclin-dependent kinase (CDK) inhibitors in the control of G2-M and suggest that their induction may stimulate the occurrence of endomitosis or polyploidy in a number of physiological settings. In this report, the stable expression of the p120E4F transcription factor, which attenuates G1-S progression by elevating p21WAF1 and p27KIP1 protein levels, was shown to also interfere with the regulation of G2-M and cytokinesis. Exponentially growing cultures of p120E4F-expressing fibroblast cell lines had reduced levels of CDC2 kinase activity, elevated levels of Cyclin B1 protein, and continuously generated a subpopulation of tetraploid cells and elevated numbers of multinucleated cells. Coexpression of activated Ras, which stimulates Cyclin D1 expression and G1-S-specific cyclin-CDK kinase activities, alleviated these effects without reducing p21WAF1 or p27KIP1 protein levels; p120E4F/ras-expressing cell lines contained reduced levels of Cyclin B1 protein, a restoration of Cyclin B-CDC2 kinase activity to control levels, and exhibited no increase of tetraploid or multinucleated cells. Interestingly, changes in the expression of Cyclin B1 and, to a lesser extent, CDC2 were primarily regulated by post-transcriptional mechanisms. The results indicate that mechanisms which moderately elevate CDK inhibitor levels can reduce CDC2 kinase activity to the point of impeding normal G2-M function and suggest that two molecular determinants commonly associated with the induction of polyploidy in a number of tissues, i.e., elevated levels of universal CDK inhibitors and sustained CDK2 kinase activity, may be solely sufficient to initiate endomitosis.

3T3 Cells↗
PubMed2000

p53 is involved in the p120E4F-mediated growth arrest.

Control of cell growth and division by the p53 tumor suppressor protein requires its abilities to transactivate and repress specific target genes and to associate in complex with other proteins. Here we demonstrate that p53 binds to the E1A-regulated transcription factor p120E4F, a transcriptional repressor of the adenovirus E4 promoter. The interaction involves carboxy-terminal half of p120E4F and sequences located at the end of the sequence-specific DNA-binding domain of p53. Ectopic expression of p120E4F leads to a block of cell proliferation in several human and murine cell lines and this effect requires the association with wild-type (wt) p53. Although p120E4F can also bind to mutant p53, the growth suppression induced by overexpression of the protein is severely reduced in a cell line that contains mutant p53. These data suggest that p120E4F may represent an important element within the complex network of p53 checkpoint functions.

3T3 Cells↗
PubMed2000

TEL, a putative tumor suppressor, modulates cell growth and cell morphology of ras-transformed cells while repressing the transcription of stromelysin-1.

TEL is a member of the ETS family of transcription factors that interacts with the mSin3 and SMRT corepressors to regulate transcription. TEL is biallelically disrupted in acute leukemia, and loss of heterozygosity at the TEL locus has been observed in various cancers. Here we show that expression of TEL in Ras-transformed NIH 3T3 cells inhibits cell growth in soft agar and in normal cultures. Unexpectedly, cells expressing both Ras and TEL grew as aggregates. To begin to explain the morphology of Ras-plus TEL-expressing cells, we demonstrated that the endogenous matrix metalloproteinase stromelysin-1 was repressed by TEL. TEL bound sequences in the stromelysin-1 promoter and repressed the promoter in transient-expression assays, suggesting that it is a direct target for TEL-mediated regulation. Mutants of TEL that removed a binding site for the mSin3A corepressor but retained the ETS domain failed to repress stromelysin-1. When BB-94, a matrix metalloproteinase inhibitor, was added to the culture medium of Ras-expressing cells, it caused a cell aggregation phenotype similar to that caused by TEL expression. In addition, TEL inhibited the invasiveness of Ras-transformed cells in vitro and in vivo. Our results suggest that TEL acts as a tumor suppressor, in part, by transcriptional repression of stromelysin-1.

3T3 Cells↗
PubMed1999

Suppression of E1A-mediated transformation by the p50E4F transcription factor.

The adenovirus E1A gene can act as an oncogene or a tumor suppressor, with the latter effect generally arising from the induction of apoptosis or the repression of genes that provide oncogenic growth stimuli (e.g., HER-2/c-erbB2/neu) or increased metastatic invasiveness (e.g., metalloproteases). In this study, coexpression of E1A and p50E4F, a cellular transcription factor whose DNA binding activity is stimulated by E1A, suppressed colony formation by NIH 3T3 cells and transformation of primary rat embryo fibroblasts but had no observed effect in the absence of E1A. Domains in p50E4F required for stimulation of the adenovirus E4 promoter were required for the suppressive effect, indicating a transcriptional mechanism. In serum-containing media, retroviral expression of p50E4F in E1A13S/ras-transformed NIH 3T3 fibroblasts had little effect on subconfluent cultures but accelerated a decline in viability after the cultures reached confluence. Cell death occurred by both apoptosis and necrosis, with the predominance of each process determined by culture conditions. In serum-free media, p50E4F accelerated E1A-induced apoptosis. The results suggest that p50E4F sensitizes cells to signals or conditions that cause cell death.

3T3 Cells↗
PubMed1998

Mutational analysis of p50E4F suggests that DNA binding activity is mediated through an alternative structure in a zinc finger domain that is regulated by phosphorylation.

p50E4F is a cellular transcription factor whose DNA binding activity is stimulated in a phosphorylation-dependent manner by products of the adenovirus E1A oncogene. Although p50E4F does not contain a bZIP DNA binding motif, it binds a tandemly repeated palindromic sequence in the adenovirus E4 promoter that is recognized by a large number of bZIP proteins, but with much greater stability. Analysis of deletions in the p50E4F sequence identified the regions that are responsible for its unique DNA binding properties. Sequence-specific DNA binding and factor dimerization were localized to a C-terminal region containing two C2H2and one CCHC zinc finger motifs; the phosphorylation site critical for DNA binding activity was also localized to this domain. The high stability of p50E4F binding also required residues within the first 83 amino acids of the N-terminus. Analysis of single and double amino acid substitutions in the C-terminal zinc finger domain demonstrated that while the second C2H2zinc finger was required for DNA binding activity, the putative structures of the first C2H2and the CCHC zinc fingers were not. Instead, residues from these other zinc finger motifs appeared to participate in an alternative structure that mediates DNA binding activity and is regulated by phosphorylation.

Amino Acid Sequence↗
PubMed1998

Adenovirus E1A-regulated transcription factor p120E4F inhibits cell growth and induces the stabilization of the cdk inhibitor p21WAF1.

Adenovirus E1A proteins influence cell growth and phenotype through physical interactions with cellular proteins that regulate basic processes such as cell cycle progression, DNA synthesis, and differentiation. p120E4F is a low-abundance cellular transcription factor that represses the adenovirus E4 promoter and is regulated by E1A, through a phosphorylation-induced reduction of its DNA binding activity, to permit activation of the E4 promoter during early infection. To determine the normal biological role of p120E4F, we assessed its ability to influence fibroblast cell growth and transformation. p120E4F suppressed NIH 3T3 fibroblast colony formation but had little effect when coexpressed with E1A and/or activated ras. Cells that overexpressed p120E4F were inhibited in their ability to enter S phase, had elevated levels of the cdk inhibitor p21WAF1, and reduced cyclin D-cdk4/6 kinase activity. The increase of p21WAF1 levels occurred through a p53-independent posttranscriptional mechanism that included a three- to fourfold increase in the half-life of p21WAF1 protein. Coexpression of activated ras with p120E4F stimulated cyclin D1 expression, elevated cyclin D-cdk4/6 kinase activity, and accelerated cell growth. These data suggest an important role for p120E4F in normal cell division and demonstrate that p21WAF1 can be regulated by protein turnover.

Adenoviridae↗
PubMed1997

The adenovirus E1A-regulated transcription factor E4F is generated from the human homolog of nuclear factor phiAP3.

A 50-kDa cellular factor, E4F, has been implicated in mediating trans activation of the adenovirus E4 gene by the 289R E1A(13S) protein. Previous experiments demonstrated an E1A-dependent increase in E4F DNA binding activity, dependent on phosphorylation, that correlated with the activation of E4 transcription. Using expression screening, we isolated a cDNA clone encoding the E4F protein, as judged by DNA binding characteristics, transcriptional activation, and immunological criteria. The E4F-1 cDNA encodes a 783-amino-acid polypeptide that has 86% sequence identity with the murine nuclear factor phiAP3, a GLI-krüppel-related protein. E4F DNA binding activity is encoded within an amino-terminal region of E4F-1 that contains a zinc finger domain and, as with endogenous E4F, is phosphatase sensitive. We found that E4F was generated from the full-length E4F-1-encoded protein as a 50-kDa amino-terminal fragment. Moreover, E1A(13S) expression induced the phosphorylation of both forms of E4F-1 but differentially regulated their DNA binding activities, stimulating the 50-kDa fragment while reducing the activity of the full-length protein. In transient-transfection assays, the E4F-1 amino-terminal fragment stimulated the adenovirus E4 promoter in the presence of E1A(13S), whereas the full-length protein repressed the promoter in the absence, but not the presence, of E1A. The results indicate that the 50-kDa polypeptide responsible for E4F DNA binding activity is a fragment generated from the human homolog of phiAP3 and that the two forms of the E4F-1 protein are differentially regulated by E1A through phosphorylation.

Adenovirus E1A Proteins↗
PubMed1994

Case report 876: Aneurysmal bone cyst of the patella.

A case of aneurysmal bone cyst of the patella in a 15-year-old woman was presented. An ABC occurring in a female patient is rare, and to our knowledge this is the first report in the literature. All other reports of patellar ABCs reported have occurred in young black men [2, 3]. The radiographic findings and clinical presentation were stressed since they are typical of all benign patellar tumors. Although radiographic findings are usually typical of patellar tumors, CT and MRI may be used for staging and standardization of operative treatment. MRI is also helpful for examining the integrity of the patellar tendon and quadriceps since many of these patients present with malfunction of the extensor mechanism [14].

Adolescent↗
PubMed1993

Giant cell tumour of bone. A surgical approach to grade III tumours.

Thirty-one patients with giant cell tumours of bone (BGCT) were treated at our hospital by a conservative approach between 1975 and 1988. 94% of the tumours were grade III (Campanacci) and the follow up was from 2 to 13 years. In the 20 patients who had a modified intralesional excision (curettage) with bone grafting and cement, 5 recurrences occurred. There was one soft tissue recurrence in the 11 patients who had marginal or wide excisions. All recurrences were in grade III lesions. We conclude that grade III BGCT can be treated by modified intralesional excision provided the articular surfaces and part of the metaphysis are intact.

Adolescent↗
PubMed1993

Bilateral dysplasia epiphysealis hemimelica: a case report.

Dysplasia epiphysealis hemimelica is a rare disorder of cartilage growth. Classically, the disease involves a single limb and is isolated to the medial or lateral half of that limb. A patient is described who presented with classic findings of this disease but involvement of both ankles.

Adult↗
PubMed1992

Stabilization of the interphalangeal joint of the big toe: comparison of three methods.

Three methods of stabilizing the IP of the big toe were compared. In group A, 10 patients underwent tenodesis of the extensor hallucis longus to the extensor digitorum brevis tendon. All of them developed a toe-drop; two patients had significant symptoms that required IP fusion. In group B, 19 patients underwent IP fusion using smooth or threaded intramedullary Kirschner wire fixation. There were nine nonunions, three requiring refusion. In group C, 32 patients underwent IP fusion using intramedullary screw fixation. There was one nonunion with screw failure that required revision. Although none of our patients considered the toe-drop after extensor hallucis longus tenodesis cosmetically unacceptable, this may not be so in other cultures. All complications following IP fusion with screw fixation were technical and are avoidable. When stabilization of IP is required, we recommend fusion of IP with screw fixation.

Adolescent↗
PubMed1992

Hydatid bone disease of the pelvis. A report of two cases and review of the literature.

Hydatid disease is caused by a parasitic tapeworm Echinococcus. This parasite in larval stage can thrive in many parts of the body, most frequently in the liver. Hydatid disease in bone is rare, but in areas where hydatid disease is endemic, it must always be considered in the differential diagnosis. Two cases of hydatid bone disease of the pelvis are reported. The treatment of choice is a combination of chemotherapy and surgery.

Aged↗
PubMed1991

Brucellosis complicating bilateral total knee arthroplasty.

Infection after arthroplasty is a serious complication associated with a high incidence of morbidity. A 24-year-old woman developed brucella septic arthritis in both knees following bilateral total knee arthroplasty. This complication appears not to have been previously reported in the literature.

Adult↗
PubMed1990

E1A-dependent trans-activation of the c-fos promoter requires the TATAA sequence.

Previous experiments have demonstrated that transcription of the human c-fos oncogene is activated through the action of the 289-amino acid adenovirus E1A gene product. In this study we have utilized a series of c-fos promoter deletion and substitution mutants to define regulatory sequences that allow the induction by E1A. Although the deletion of upstream promoter sequences has varying degrees of effect on overall promoter activity, these deletions retain inducibility by E1A. This includes the deletion of the serum response element and two elements that bind the ATF transcription factor. In fact, a c-fos promoter deleted to position -53, which leaves the TATA element but no other known functional element, retains inducibility, indicating a role for the TATA element in E1A control. Indeed, substitution of the c-fos TATA element (TATAA) with a TATA sequence from the simian virus 40 early promoter (TATTTAT) abolishes E1A inducibility; this promoter does retain responsiveness to cAMP induction, however, demonstrating that this TATTTAT substitution is functional. We conclude that the E1A-dependent activation of c-fos transcription is mediated through an effect on a TATA-binding protein that has specificity for the TATAA sequence.

Adenovirus Early Proteins↗
PubMed1990

E4F and ATF, two transcription factors that recognize the same site, can be distinguished both physically and functionally: a role for E4F in E1A trans activation.

Previous experiments have identified an element in the adenovirus E4 promoter that is critical for E1A-dependent trans activation and that can confer inducibility to a heterologous promoter. This DNA element is a recognition site for multiple nuclear factors, including ATF, which is likely a family of DNA-binding factors with similar DNA recognition properties. However, ATF activity was found not to be altered in any demonstrable way as a result of adenovirus infection. In contrast, another factor that recognizes this element, termed E4F, was found at only very low levels in uninfected cells but was increased markedly upon adenovirus infection, as measured in DNA-binding assays. Although both the ATF activity and the E4F activity recognized and bound to the same two sites in the E4 promoter, they differed in their sequence recognition of these sites. Furthermore, E4F bound only to a small subset of the ATF recognition sites; for instance, E4F did not recognize the ATF sites in the E2 or E3 promoters. Various E4F and ATF binding sites were inserted into an expression vector and tested by cotransfection assays for responsiveness to E1A. We found that a sequence capable of binding E4F could confer E1A inducibility. In contrast, a sequence that could bind ATF but not E4F did not confer E1A inducibility. We also found that E4F formed a stable complex with the E4 promoter, whereas the ATF DNA complex was unstable and rapidly dissociated. We conclude that the DNA-binding specificity of E4F as well as the alterations in DNA-binding activity of E4F closely correlates with E1A stimulation of the E4 promoter.

Activating Transcription Factors↗