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J Mihaly

Publications and source records attributed to J Mihaly.

11 recordsLinked to original sources
PubMed2001

Variations of components of the plasminogen activation system with the cell cycle in benign prostate tissue and prostate cancer.

BACKGROUND: Components of the fibrinolytic system are involved in tumor cell invasion and metastasis. Previous investigations suggested a cell cycle-dependent expression of urokinase-type plasminogen activator (u-PA) in epithelial cells. In order to determine a correlation of cell cycle phases with the fibrinolytic system, we investigated the expression of u-PA, tissue-type plasminogen activator (t-PA), and plasminogen activator inhibitor type 1 (PAI-1) in normal and tumor-containing prostate extracts and analyzed a possible relationship with flow cytometry-determined proliferative activity of the samples. Cell cycle phases were correlated with fibrinolytic parameters in prostate tissue. METHODS: Samples were obtained from patients undergoing radical prostatectomy for prostate cancer and separated into two portions for DNA analysis and the detection of u-PA, t-PA, and PAI-1. Flow cytometric analysis was performed according to the Vindelov technique. The concentrations of u-PA, t-PA, and PAI-1 were determined from tissue extracts after homogenization by an enzyme-linked immunosorbent assay (ELISA) technique. RESULTS: Correlations of u-PA and t-PA expression with the frequency of G0/G1, S, G2M, S-phase fraction (SPF), and proliferation index (PI) for normal prostate and prostate cancer revealed no significant correlation. The only significant finding was observed in normal tissue revealing a positive correlation between PAI-1 expression and G0/G1 and a negative correlation with S-phase, SPF, and PI. No dependence of PAI-1 expression on different cell phases was found in prostate cancer. Furthermore, no significant correlation of u-PA, t-PA, and PAI-1 with cell cycles in organ-confined ( or = pT3a) tumors was found. No significant correlation in prostate cancer of components of the fibrinolytic system differentiated according to tumor grade or perineural tumor infiltration and cell cycle analysis was found. Only in highly differentiated G1 (Gleason 2-4) cancer, u-PA had a significant positive correlation with G2M-phase. CONCLUSION: Absence of a correlation between levels of components of the fibrinolytic system and cell cycle phases suggests that the reported association between increases of some of these components and aggressive biological behavior of prostate cancer is secondary to non-cell cycle-related mechanisms.

Cell Cycle↗
PubMed2001

The role of the Drosophila TAK homologue dTAK during development.

The TAK kinases belong to the MAPKKK group and have been implicated in a variety of signaling events. Originally described as a TGF-beta activated kinase (TAK) it has, however, subsequently been demonstrated to signal through p38, Jun N-terminal kinase (JNK) and Nemo types of MAP kinases, and the NFkappaB inducing kinase. Despite these multiple proposed functions, the in vivo role of TAK family kinases remains unclear. Here we report the isolation and genetic characterization of the Drosophila TAK homologue (dTAK). By employing overexpression and double-stranded RNA interference (RNAi) techniques we have analyzed its function during embryogenesis and larval development. Overexpression of dTAK in the embryonic epidermis is sufficient to induce the transcription of the JNK target genes decapentaplegic and puckered. Furthermore, overexpression of dominant negative (DN) or wild-type forms of dTAK in wing and eye imaginal discs, respectively, results in defects in thorax closure and ommatidial planar polarity, two well described phenotypes associated with JNK signaling activity. Surprisingly, RNAi and DN-dTAK expression studies in the embryo argue for a differential requirement of dTAK during developmental processes controlled by JNK signaling, and a redundant or minor role of dTAK in dorsal closure. In addition, dTAK-mediated activation of JNK in the Drosophila eye imaginal disc leads to an eye ablation phenotype due to ectopically induced apoptotic cell death. Genetic analyses in the eye indicate that dTAK can also act through the p38 and Nemo kinases in imaginal discs. Our results suggest that dTAK can act as a JNKKK upstream of JNK in multiple contexts and also other MAPKs in the eye. However, the loss-of-function RNAi studies indicate that it is not strictly required and thus either redundant or playing only a minor role in the context of embryonic dorsal closure.

Amino Acid Sequence↗
PubMed2001

Replacement of Fab-7 by the gypsy or scs insulator disrupts long-distance regulatory interactions in the Abd-B gene of the bithorax complex.

Chromatin domain boundaries, like scs or gypsy insulators in Drosophila, have been identified in transgene assays through their enhancer-blocking activity. Boundary elements in the bithorax complex (BX-C), such as Fab-7 and Fab-8, have been identified genetically and been shown to have insulator activity in transgene assays. However, it is not clear whether boundary elements identified in transgene assays will function appropriately in chromosomal contexts such as BX-C. Using gene conversion, we have substituted the scs or gypsy insulators for Fab-7. We find that both scs and gypsy are very potent insulators in the ectoderm, but surprisingly, the insulating activity of gypsy (but not scs) is lost in the CNS. Our results reveal that the Fab-7 boundary must have special properties that scs and gypsy lack, which allow it to function appropriately in BX-C regulation.

Abdomen↗
PubMed2001

The iab-7 polycomb response element maps to a nucleosome-free region of chromatin and requires both GAGA and pleiohomeotic for silencing activity.

In the work reported here we have undertaken a functional dissection of a Polycomb response element (PRE) from the iab-7 cis-regulatory domain of the Drosophila melanogaster bithorax complex (BX-C). Previous studies mapped the iab-7 PRE to an 860-bp fragment located just distal to the Fab-7 boundary. Located within this fragment is an approximately 230-bp chromatin-specific nuclease-hypersensitive region called HS3. We have shown that HS3 is capable of functioning as a Polycomb-dependent silencer in vivo, inducing pairing-dependent silencing of a mini-white reporter. The HS3 sequence contains consensus binding sites for the GAGA factor, a protein implicated in the formation of nucleosome-free regions of chromatin, and Pleiohomeotic (Pho), a Polycomb group protein that is related to the mammalian transcription factor YY1. We show that GAGA and Pho interact with these sequences in vitro and that the consensus binding sites for the two proteins are critical for the silencing activity of the iab-7 PRE in vivo.

Amino Acid Motifs↗
PubMed2000

Signaling specificity by Frizzled receptors in Drosophila.

Wnt-Frizzled (Fz) signaling pathways play recurring important roles during the development and homeostasis of vertebrates and invertebrates. Fz receptors can signal through beta-catenin-dependent and -independent pathways. In Drosophila, Fz and Fz2 are redundant receptors for Wg. In addition, Fz conveys signals through a distinct pathway to organize planar polarization of epithelial structures. We demonstrate that the cytoplasmic sequences of Fz2 and Fz preferentially activate the beta-catenin and planar polarity cascade, respectively. Both receptors activate either pathway, but with different efficiencies. Intrinsic differences in signaling efficiency in closely related receptors might be a general mechanism for generating signaling specificity in vivo.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Involvement of low-density lipoprotein receptor-related protein (LRP) in the clearance of factor VIII in von Willebrand factor-deficient mice.

Factor VIII is tightly noncovalently linked to von Willebrand factor (vWF) in plasma with a stoichiometry of 1:50, and vWF deficiency results in secondary factor VIII deficiency, with accelerated clearance of factor VIII from the circulation. We used a murine model of severe von Willebrand disease (vWF knockout mice) to study the effect of a recombinant vWF/pro-vWF preparation (rpvWF) on factor VIII survival and to investigate whether low-density lipoprotein receptor-related protein (LRP) might be involved in the in vivo clearance of factor VIII in the absence of vWF. vWF-deficient mice received 70 U/kg rpvWF in the first series of experiments, and in a second series, 80 mg/kg receptor-associated protein (RAP) as a recombinant fusion protein to block the action of LRP. Factor VIII levels were measured at time 0, or 1 or 3 hours after administration of rpvWF or RAP. RAP induced a sustained rise in factor VIII levels comparable to that induced by rpvWF. In a third series, the preadministration of RAP resulted in a slower disappearance of factor VIII antigen (measured by an enzyme-linked immunosorbent assay specific for human factor VIII) after infusion of recombinant factor VIII. These findings suggest that the accelerated clearance of factor VIII seen in the absence of vWF may be a result of the involvement of LRP in factor VIII metabolism. (Blood. 2000;95:1703-1708)

Animals↗
PubMed2000

The Fab-8 boundary defines the distal limit of the bithorax complex iab-7 domain and insulates iab-7 from initiation elements and a PRE in the adjacent iab-8 domain.

The Drosophila bithorax complex Abdominal-B (Abd-B) gene specifies parasegmental identity at the posterior end of the fly. The specific pattern of Abd-B expression in each parasegment (PS) determines its identity and, in PS10-13, Abd-B expression is controlled by four parasegment-specific cis-regulatory domains, iab-5 to iab-8, respectively. In order to properly determine parasegmental identity, these four cis-regulatory domains must function autonomously during both the initiation and maintenance phases of BX-C regulation. The studies reported here demonstrate that the (centromere) distal end of iab-7 domain is delimited by the Fab-8 boundary. Initiators that specify PS12 identity are located on the proximal iab-7 side of Fab-8, while initiators that specify PS13 identity are located on the distal side of Fab-8, in iab-8. We use transgene assays to demonstrate that Fab-8 has enhancer blocking activity and that it can insulate reporter constructs from the regulatory action of the iab-7 and iab-8 initiators. We also show that the Fab-8 boundary defines the realm of action of a nearby iab-8 Polycomb Response Element, preventing this element from ectopically silencing the adjacent domain. Finally, we demonstrate that the insulating activity of the Fab-8 boundary in BX-C is absolutely essential for the proper specification of parasegmental identity by the iab-7 and iab-8 cis-regulatory domains. Fab-8 together with the previously identified Fab-7 boundary delimit the first genetically defined higher order domain in a multicellular eukaryote.

Animals↗
PubMed1998

Analysis of fibrinolytic proteins in relation to DNA ploidy in prostate cancer.

The tissue concentrations of urokinase-type plasminogen activator (u-PA), urokinase-type plasminogen activator receptor (u-PAR), plasminogen activator inhibitor type 1 (PAI-1) and tissue-type plasminogen activator (t-PA) were investigated by an ELISA technique in normal and malignant samples of the prostate from 24 patients undergoing radical prostatectomy for organ-confined prostate cancer. The median concentration of u-PA was significantly higher in cancerous than in normal prostate tissue (p = 0.006). No significant increase of u-PAR, PAI-1 and t-PA was found in cancer tissue in comparison with the benign samples (p > 0.05). Assessment of the relationship between fibrinolytic proteins and DNA ploidy revealed an increased u-PA, u-PAR and PAI-1 in diploid prostate cancer as compared with the normal controls. However, in aneuploid cancer u-PA remained high but u-PAR and PAI-1 were decreased. This led to a higher local concentration of u-PA in aneuploid samples than in normal prostate and in diploid prostate cancer. No alteration of median t-PA was found in benign prostate or in diploid or aneuploid prostate cancer. The altered expression of u-PA, u-PAR and PAI-1 in diploid and aneuploid prostate cancer suggests a possible role of fibrinolytic proteins in the different biologic behavior of tumors, and may be one explanation for the higher metastatic potential of aneuploid tumors.

Aged↗
PubMed1998

Chromatin domain boundaries in the Bithorax complex.

Eukaryotic chromosomes are thought to be organized into a series of discrete higher-order chromatin domains. This organization is believed to be important not only in the compaction of the chromatin fibre, but also in the utilization of genetic information. Critical to this model are the domain boundaries that delimit and segregate the chromosomes into units of independent gene activity. In Drosophila, such domain boundaries have been identified through two different approaches. On the one hand, elements like scs/scs' and the reiterated binding site for the SU(HW) protein have been characterized through their activity of impeding enhancer-promoter interactions when intercalated between them. Their role of chromatin insulators can protect transgenes from genomic position effects, thereby establishing independent functional domains within the chromosome. On the other hand, domain boundaries of the Bithorax complex (BX-C) like Fab-7 and Mcp have been identified through mutational analysis. Mcp and Fab-7, however, may represent a specific class of boundary elements; instead of separating adjacent domains that contain separate structural genes. Mcp and Fab-7 delimit adjacent cis-regulatory domains, each of which interacts independently with their target promoters. In this article, we review the genetic and molecular characteristics of the domain boundaries of the BX-C. We describe how Fab-7 functions to confine activating as well as repressive signals to the flanking regulatory domains. Although the mechanisms by which Fab-7 works as a domain boundary remain an open issue, we provide preliminary evidence that Fab-7 is not a mere insulator like scs or the reiterated binding site for the SU(HW) protein.

Animals↗
PubMed1997

In situ dissection of the Fab-7 region of the bithorax complex into a chromatin domain boundary and a Polycomb-response element.

Parasegmental (PS)-specific expression of the homeotic genes of the bithorax-complex (BX-C) appears to depend upon the subdivision of the complex into a series of functionally independent cis-regulatory domains. Fab-7 is a regulatory element that lies between iab-6 and iab-7 (the PS11- and PS12-specific cis-regulatory domains, respectively). Deletion of Fab-7 causes ectopic expression of iab-7 in PS11 (where normally only iab-6 is active). Two models have been proposed to account for the dominant Fab-7 phenotype. The first considers that Fab-7 functions as a boundary element that insulates iab-6 and iab-7. The second model envisages that Fab-7 contains a silencer element that keeps iab-7 repressed in parasegments anterior to PS12. Using a P-element inserted in the middle of the Fab-7 region (the bit transposon), we have generated an extensive collection of new Fab-7 mutations that allow us to subdivide Fab-7 into a boundary element and a Polycomb-respond element (PRE). The boundary lies within 1 kb of DNA on the proximal side of the bit transposon (towards iab-6). Deletions removing this element alone cause a complex gain- and loss-of-function phenotype in PS11; in some groups of cells, both iab-6 and iab-7 are active, while in others both iab-6 and iab-7 are inactive. Thus, deletion of the boundary allows activating as well as repressing activities to travel between iab-6 and iab-7. We also provide evidences that the boundary region contains an enhancer blocker element. The Polycomb-response element lies within 0.5 kb of DNA immediately distal to the boundary (towards iab-7). Deletions removing the PRE alone do not typically cause any visible phenotype as homozygotes. Interestingly, weak ectopic activation of iab-7 is observed in hemizygous PRE deletions, suggesting that the mechanisms that keep iab-7 repressed in the absence of this element may depend upon chromosome pairing. These results help to reconcile the previously contradictory models on Fab-7 function and to shed light on how a chromatin domain boundary and a nearby PRE concur in the setting up of the appropriate PS-specific expression of the Abd-B gene of the BX-C.

Abdomen↗