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R Marschalek

Publications and source records attributed to R Marschalek.

At least 37 records · Page 2Linked to original sources
PubMed1997

The structure of the human ALL-1/MLL/HRX gene.

The human ALL-1/MLL/HRX gene on chromosome 11q23 is the site of many locally clustered chromosomal alterations associated with several types of acute leukemias, including deletions. partial duplications and reciprocal translocations. Structurally variant proteins derived from an altered ALL-1 gene presumably make essential contributions to the malignant transformation of hematopoietic progenitor cells. The ALL-1 gene is spread over approximately 92 kb and consists of at least 37 exons. An exon/intron map including the position of the 3'-end of the gene and a detailed restriction map were produced and an updated map is presented. Data from other laboratories were incorporated where compatible. Exon/intron boundaries were sequenced and an intron-phase analysis was performed. The results are expected to contribute to a better understanding of those structural alterations of the gene that conserve the open reading frame and produce presumably oncogenic variants of the ALL-1 protein. They will also facilitate the rapid molecular diagnosis of structural alterations of this gene and the choice of therapeutic options. Mechanisms that may potentially account for the striking clustering of the translocation breakpoints in the breakpoint cluster region of the gene are discussed.

Base Sequence↗
PubMed1997

Hydroxyurea for treatment of unresectable and recurrent meningiomas. II. Decrease in the size of meningiomas in patients treated with hydroxyurea.

In this paper the authors present the first evidence that meningiomas respond to treatment with hydroxyurea. Hydroxyurea was administered as an adjunct chemotherapeutic treatment in patients with recurrent and unresectable meningiomas. Hydroxyurea was used because experimental data demonstrated that it inhibits growth of cultured human meningioma cells and meningioma transplants in nude mice by inducing apoptosis. The authors therefore treated four selected patients with hydroxyurea. All patients had undergone multiple gross resections and all except one received radiotherapy. Three patients with recurrent Grade I meningiomas assessed according to World Health Organization (WHO) guidelines received hydroxyurea because of an increased tumor growth rate, documented by magnetic resonance (MR) imaging, within a 6- or 12-month interval. A fourth patient with a malignant meningioma (WHO Grade III) began a course of treatment with hydroxyurea immediately after his sixth palliative operation without waiting for another relapse to be demonstrated on MR imaging. Because of their location and invasive growth behavior none of the meningiomas could have been removed completely by surgical intervention. All patients received hydroxyurea at a dosage level of 1000 to 1500 mg/day (approximately 20 mg/kg/day). In a man with a large sphenoid wing meningioma invading the right cavernous sinus and the temporal base, the intracranial tumor mass was reduced by 60% during 6 months of treatment. A woman with a large ball-shaped meningioma of the right sphenoid wing invading the cavernous sinus exhibited a 74% decrease of the initial tumor volume in 10 months of treatment with oral hydroxyurea. Serial MR images obtained monthly revealed that the process of size reduction was continuous and proportionate. The shrinkage of the tumor was accompanied by a complete remission of symptomatic trigeminal neuralgia after 2 months and by improved abducent paresis after 5 months. The third patient had a slowly growing meningioma that exhibited a 15% reduction in mass when reassessed after 5 months of hydroxyurea treatment. The fourth patient with the malignant meningioma in the left cerebellopontine angle has had no recurrence for 24 months. Long-term treatment with hydroxyurea may result in full remission of tumors in meningioma patients. The preliminary data indicate that hydroxyurea provides true medical treatment in patients with unresectable and recurrent meningiomas, replacing palliative surgery and radiotherapy in the management of this disease.

Adult↗
PubMed1997

Hydroxyurea for treatment of unresectable and recurrent meningiomas. I. Inhibition of primary human meningioma cells in culture and in meningioma transplants by induction of the apoptotic pathway.

Meningiomas, which invade intracranial bone structures and the adjacent connective tissue, are frequently unresectable because of their aggressive and recalcitrant growth behavior. They have a high recurrence rate, and in approximately 10% of these tumors there is an increased risk of malignancy. Significant morbidity and mortality rates associated with recurrent meningiomas demand nonsurgical approaches. To date, adjuvant hormonal treatment has not proven beneficial. The anticancer drug hydroxyurea was therefore tested for its potential use in the treatment of meningiomas. Early-passaged cell cultures were established from 20 different meningiomas. The addition of 5 x 10(-4) and 10(-3) M hydroxyurea over a period of 5 to 9 days resulted in a remarkable decrease in cell proliferation and even blocked tumor cell growth when compared with untreated cells. A significant arrest of meningioma cell growth in the S phase of the cell cycle was revealed on DNA flow cytometry. Electron micrographs of hydroxyurea-treated tumor cells showed ultrastructural features consistent with apoptosis, and light microscopy demonstrated DNA fragmentation by in situ DNA strand break labeling. Short-term treatment of meningioma cell cultures with hydroxyurea for 24 to 48 hours resulted in discrete oligonucleosomal fragments (DNA ladder), another characteristic sign of apoptosis. In addition to the in vitro studies, tissue from five different meningiomas was transplanted into nude mice followed by treatment with 0.5 mg/g body weight hydroxyurea over 15 days. In situ DNA strand break labeling demonstrated DNA fragmentation in distinct regions with different tumor cell densities in all hydroxyurea-treated meningioma transplants. These data provide evidence that hydroxyurea is a powerful inhibitor of meningioma cell growth, most likely by causing apoptosis in the tumor cells. Thus, hydroxyurea may be a suitable chemotherapeutic agent for the long-term treatment of unresectable or semi- to malignant meningiomas, or for preventing recurrent growth of meningiomas after resection.

Animals↗
PubMed1996

A specific deletion in the breakpoint cluster region of the ALL-1 gene is associated with acute lymphoblastic T-cell leukemias.

A variety of chromosomal translocations to the ALL-1 gene are regularly observed in acute leukemias and are thought to play a key role in the leukemogenic process. Chimeric proteins are encoded by the breakpoint regions of the derivative chromosomes have been proposed to be the relevant oncogenic agents. In addition, internal duplications of the ALL-1 gene have been observed in patients with specific acute myeloid leukemias. Thus, it has been hypothesized that oncogenic variants of the ALL-1 protein may be generated by both chimerization and self-fusion, but the critical structural features endowing the altered proteins with their oncogenic potential are still unknown. Here a novel structural alteration of the ALL-1 gene was observed in three patients presenting with acute T-cell leukemia (ALL) without chromosomal translocations or self-fusions of the ALL-1 gene. These unrelated patients carried an internal deletion in one of the two alleles of the ALL-1 gene that eliminated parts of introns 7 and 8, together with exon 8. The deletion was found in 3 of 74 ALL patients, but not in acute myeloid leukemias, follicular lymphomas, or peripheral blood leukocytes from healthy donors. One ALL patient showed the deletion at diagnosis but no longer at remission or at 9 months after remission. These findings support the hypothesis that the ALL-1 protein may be converted to an oncogenic variant, not only by chimerization or self-fusion, but also by deletion of sequences coded by exon 8. They further suggest that these three different types of structural alterations of the ALL-1 protein may each cause a distinct disease phenotype. Alternatively spliced mRNA species omitting exon 8 were observed in 14 of 24 ALL patients without detectable macroscopic alterations of the ALL-1 gene and also in peripheral blood leukocytes from healthy donors.

Adolescent↗
PubMed1996

Hormonal dependency of cerebral meningiomas.

Though meningiomas are benign intracranial tumors, a minor group invades the skull base and the connective tissue of the sinus cavernous inducing neurological deficits. These patients can not be cured by surgical treatment. Therefore, the development of an adjuvant medical therapy has been the goal during the last decade. We report here on different medical concepts which are based on steroids, amines, growth factor antagonists and cytokines. In addition, our data give evidence that the growth of intracranial meningiomas is under multifactorial proliferative control.

Brain↗
PubMed1996

Exon/intron structure of the human ALL-1 (MLL) gene involved in translocations to chromosomal region 11q23 and acute leukaemias.

The acute lymphoblastic leukaemia (ALL)-1 gene on human chromosome 11q23 is the site of many locally clustered chromosomal alterations associated with several types of acute leukaemias, including deletions, partial duplications and translocations. Structurally variant proteins derived from the altered gene presumably cause the malignant transformation of early haemopoietic progenitor cells. According to previously published reports, the gene consisted of at least 21 exons spread over approximately 100 kb. In this report a set of genomic fragments was isolated that represent a total of 35 exons (exons 3-37) encompassing > 95% of the protein-coding region (except exons 1 and 2) and the 3'-non-translated region of the gene. The distances between these exons were determined and a detailed restriction map was produced. The majority of the exon/intron boundaries were sequenced and an intron-phase analysis was performed. The results form the basis for a greater understanding of the translocations and other structural alterations of the gene that conserve the open reading frame and thus produce presumably oncogenic variants of the ALL-1 protein.

Base Sequence↗
PubMed1995

Isolation of two interleukin-6 response element binding proteins from acute phase rat livers.

Proteins binding at the IL-6 response element of the rat alpha 2 macroglobulin gene were purified by a combination of conventional chromatographic procedures and binding-site specific DNA affinity chromatography. The proteins were purified from the nuclei of rat livers, excised at the peak of an experimentally induced acute phase response. By this procedure three polypeptides of 92, 91 and 86 kD were enriched more than 6,000-fold. Partial proteolysis with lysyl endopeptidase and aminoacid sequence analysis of proteolytic peptides identified the 86 and 91 kD species as the transcription factor Stat3 and the 92 kD species as a Stat factor distinct from Stats 1 and 3. cDNA clones for Stats 1, 3 and this 92 kD factor were isolated from a cDNA library prepared with mRNA from acute phase rat livers. Parts of their DNA sequences were determined and the sequences of the purified peptides were found in these cDNA sequences. Thus, the identity of the factors as Stat3 and a Stat factor different from Stats 1 and 3 was confirmed. These results suggest, that APRF/Stat3 and p91/Stat1 are not the only factors mediating the effects of IL-6 on class 2 acute phase genes. The 92 kD Stat factor binding at the IL-6 RE probably also functions as a transcription factor in the cytokine-induced activation of the alpha 2M gene.

Acute-Phase Reaction↗
PubMed1995

Molecular analysis of the chromosomal breakpoint and fusion transcripts in the acute lymphoblastic SEM cell line with chromosomal translocation t(4;11).

The chromosomal breakpoint and fusion transcripts of the pre-B-leukaemia-derived SEM cell line carrying a reciprocal t(4;11)(q21;q23) translocation were analysed. The breakpoint from derivative chromosome der4 was cloned and sequenced. The crossover site was localized in intron 7 of the ALL-1 gene on chromosome 11q23 and in a large intron of the AF-4 (FEL) gene. RNA transcripts from both wild-type genes and both hybrid genes were detected by reverse transcriptase polymerase chain reaction (RT-PCR) assays. In addition, alternatively spliced mRNA species derived from the der4 chromosome were found. They were generated by using the exon 5' of the breakpoint on der4 as a common splice donor site and the 5' boundaries of exons 8 or 9 of the ALL-1 gene as alternative splice acceptor sites. The hypothesis is proposed that selective pressure operators to maintain the presence of both derivative chromosomes as important elements in the leukaemogenic process.

Base Sequence↗
PubMed1994

The acute lymphoblastic leukaemia cell line SEM with t(4;11) chromosomal rearrangement is biphenotypic and responsive to interleukin-7.

A cell line, designated SEM, was established from the peripheral blood of a 5-year-old girl in relapse with acute lymphoblastic leukaemia (ALL). Both the lymphoblasts of the patient and the cells of the cell line SEM showed the t(4;11) chromosomal rearrangement. The analysis of the immunophenotype of the SEM cell line revealed the B-cell differentiation antigens CD19, CD22 and CDw75 in the absence of CD20, CD24 and immunoglobulin expression. Besides B-lineage antigens, SEM cells were positive for the myeloid antigens CD13, CD15, CD33 and CDw65. Immunogenotypic analysis of SEM cells showed a monoclonal rearrangement of immunoglobulin heavy-chain (IgH). T-cell receptor (TCR) gamma and delta genes. Addition of interleukin (IL)-7 promoted the growth of the patient's lymphoblasts in culture and enhanced the proliferation of SEM cells. The SEM cells also express messenger RNA (mRNA) for the IL-7 receptor (IL-7R), but no evidence for autocrine production of IL-7 by the cell line was found. Addition of IL-4, tumour necrosis factor (TNF)-alpha, interferon (IFN)-alpha, or IFN-gamma resulted in a profound inhibition of SEM growth. Thus, these cytokines may have important growth regulatory activities for biphenotypic leukaemic ALL cells.

Antigens, CD↗
PubMed1994

Internally located and oppositely oriented polymerase II promoters direct convergent transcription of a LINE-like retroelement, the Dictyostelium repetitive element, from Dictyostelium discoideum.

The Dictyostelium discoideum NC4 genome harbors approximately 150 individual copies of a retrotransposable element called the Dictyostelium repetitive element (DRE). This element contains nonidentical terminal repeats (TRs) consisting of conserved building blocks A and B in the left TR and B and C in the right TR. Seven different-sized classes of RNA transcripts from these elements were resolved by Northern (RNA) blot analysis, but their combined abundance was very low. When D. discoideum cells were grown in the presence of the respiratory chain blocker antimycin A, steady-state concentrations of these RNA species increased 10- to 20-fold. The D. discoideum genome contains two DRE subtypes, the full-length 5.7-kb DREa and the internally deleted 2.4-kb DREb. Both subtypes are transcribed, as confirmed by analysis of cloned cDNA. Primary transcripts from the sense strand originate at nucleotide +1 and terminate at two dominant sites, located 21 or 28 nucleotides upstream from the 3' end of the elements. The activity of a reasonably strong polymerase II promoter in the 5'-terminal A module is slightly upregulated by the tRNA gene located 50 +/- 4 nucleotides upstream and drastically reduced by the adjacent B module of the DRE. Transcripts from the opposite DNA strand (complementary-sense transcripts) were also detected, directed by an internally located polymerase II promoter residing within the C module. This latter transcription was initiated at multiple sites within the oligo(dA12) stretch which terminates DREs.

Animals↗
PubMed1994

Characterization of transcripts from the Dictyostelium discoideum retrotransposable genetic element DRE.

All of the approximately 150 copies of the Dictyostelium discoideum retrotransposable element DRE are integrated 50 +/- 4 nucleotides upstream from different transfer RNA genes. These genomic regions are generally devoid of informative DNA why integration of the retrotransposon at these position never causes phenotypic mutations. This property makes DRE attractive as a tool in gene therapy where vectors with predictable integration specificities are still not available. Due to the nature as retrotransposon transcription is a prerequisite for the element to integrate at a new genomic position. Here we describe some transcription properties of DRE based on Northern blot analyses and on the characterization of in vitro synthesized cDNAs.

Animals↗
PubMed1993

Different organization of the tRNA-gene-associated repetitive element, DRE, in NC4-derived strains and in other wild-type Dictyostelium discoideum strains.

The retrotransposon DRE (Dictyostelium repetitive element) was discovered in the course of an extensive study concerning the genomic organization of tRNA genes in the NC4-derived strains AX2 and AX3 of the cellular slime mold Dictyostelium discoideum. As a striking feature, DRE was found exclusively in a constant orientation and at a constant distance upstream from different tRNA genes. About 150-200 DRE with intact 5'-terminal-repeat structures are present in NC4-derived strains. These strains were termed high-copy DRE strains (HCD strains) as opposed to low-copy DRE strains (LCD strains) such as the wild-type D. discoideum isolates DD61, WS380B, OHIO and V12. LCD strains contain only 3-15 DRE with intact 5'-terminal-repeat-structures. However, in addition to these few intact elements, many 5'-truncated DRE elements are present in LCD strains. In HCD strains, most DRE show typical structural characteristics of retrotransposons containing terminal repeats at both ends, which seems to be one prerequisite for active transposition. In LCD strains, however, most DRE elements are 5'-truncated, which is a common feature of eukaryotic LINE elements. Despite their truncated 5'-ends, DRE in LCD strains retain unique integration specificities, i.e. they are always found position-specifically and orientation-specifically integrated in front of tRNA genes, flanked by a 12-16-bp target-site duplication.

Animals↗
PubMed1993

The Dictyostelium discoideum 5S rDNA is organized in the same transcriptional orientation as the other rDNAs.

In eukaryotes 5S rRNA genes are transcribed by RNA polymerase III. These genes occur in D. discoideum on the ca. 90 copies of an extrachromosomal palindrom together with the other ribosomal RNAs, which are generally transcribed by RNA polymerase I. A 5S rRNA gene has been isolated and its nucleotide sequence as well as the organization of the gene relative to the RNA polymerase I operon has been determined. The sequence of the coding region corresponds exactly to an earlier published 5S rRNA sequence. The genes are located just downstream from the 26S RNA and transcription orientations of the pol I genes and the pol III gene point into the same direction, away from the centromer of the palindrom. The isolated gene appears to be functional since it serves as a specific target for a nuclear protein, most likely TFIIIA. A genomic copy of a 5S rRNA pseudogene has been isolated from the D. discoideum strain V12. This pseudocopy contains nucleotides 52 to 82 of a bona fide 5S rRNA gene with only three mismatches. It resides 78 nucleotides downstream from the glu13(UUC) tRNA gene which in the D. discoideum strain V12 is associated with the retrotransposable element DRE.

Animals↗
PubMed1993

Structure of the promoter region of the gene encoding cytochrome c oxidase subunit V in Dictyostelium.

A cDNA for the nuclear-encoded subunit V of Dictyostelium discoideum cytochrome-c oxidase was used as a probe to screen a genomic library and isolate the complete gene. Primer-extension analysis revealed two transcription start sites located 32 and 39 nucleotides upstream of the translation initiation codon. The chloramphenicol acetyltransferase assay in transient and stable Dictyostelium transformants indicated that the 400-bp dT-rich segment 5' to the transcription start sites retained promoter activity. This region contains an octanucleotide sequence similar to the yeast HAP2/3/4 responsive element.

Animals↗
PubMed1993

Nucleotide sequence of a Dictyostelium discoideum gene encoding a protein homologous to the yeast ribosomal protein S31.

A cDNA clone has been isolated whose coding potential is significantly homologous to the yeast ribosomal protein S31. The single copy genomic gene contains a 271 bp intron immediately downstream from the ATG translation initiation codon and is flanked by cannonical exon/intron junctions. The intron carries a CAATCAAT motif which has been described as inducer element for discoidin I gamma expression and which has also been found within the intron of the rp29 gene form D. discoideum. The deduced protein contains 110 amino acids and is slightly basic.

Amino Acid Sequence↗
PubMed1992

Two distinct subforms of the retrotransposable DRE element in NC4 strains of Dictyostelium discoideum.

Approximately 2% of the Dictyostelium discoideum genome consists of multiple copies of a retrotransposable element termed DRE (Dictyostelium Repetitive Element). These elements have always been found integrated in a position and orientation-specific manner 50 +/- 4 nucleotides upstream of the coding region of tRNA genes (tDNAs). An intact DRE is 5.7 kb long. It carries an extensive coding region flanked by non-identical long terminal repeats (LTRs), composed of three distinct modules A, B and C. The left LTR proximal to the tRNA gene contains one or several A-modules followed by a single B-module (AnB). By contrast, the right LTR is composed of a B-module followed by a C-module (BC). Approximately 50% of the DRE elements in NC4 derivatives of D. discoideum are structurally different from the 5.7 kb DRE described above. They carry the following alterations: a) a 3.1 kb deletion in the coding region; b) two small deletions of 8 and 29 nucleotides in the B-module of the right LTR; c) a 72 bp deletion in the B-C junction; and d) three distinct point mutations within the A-module of the left LTR. The deletion in the open reading frame encompasses the putative coding regions for reverse transcriptase adn integrase. At least 60 copies of this smaller 2.4 kb DRE subtype are found in the genome of D. discoideum NC4 strains associated with tRNA genes. Thus, inspite of their lack in reverse transcriptase and integrase those 2.4 kb elements are presumably transposable and at least all isolated copies are found exclusively in the proximity of tRNA gene loci. The enzymes needed for their replication and transposition are likely to be provided by the intact 5.7 kb DREs.

Animals↗
PubMed1992

RNA polymerase III catalysed transcription can be regulated in Saccharomyces cerevisiae by the bacterial tetracycline repressor-operator system.

We have investigated whether the RNA polymerase III-driven transcription of eukaryotic tRNA genes can be regulated by the prokaryotic tetracycline operator-repressor system. The bacterial tet operator (tetO) was inserted at two different positions (-7 and -46) upstream of a tRNA(Glu) (amber) suppressor gene. Both constructs are transcribed in Saccharomyces cerevisiae and yield functional tRNAs as scored by suppression of an amber nonsense mutation in the met8-1 allele. Controlled expression of Tet repressor was achieved by fusing the bacterial tetR gene to the yeast gal1 promoter. This leads to expression of Tet repressor in yeast on galactose--but not on glucose--containing media. Regulation of the su-tRNA gene with the tetO fragment inserted at position -7 has been demonstrated. Under conditions which allow tetR expression, cells exhibit a met- phenotype. This methionine auxotrophy can be conditionally reverted to prototrophy by adding tetracycline. However, a su-tRNA gene with the tetO fragment inserted at position -46 cannot be repressed. Our results demonstrate clearly that the bacterial repressor protein binds to its operator in the yeast genome. Formation of this complex in the vicinity of the pol III transcription initiation site reduces the level of su-tRNA at least 50-fold as concluded from quantitative primer extension analyses. This indicates for the first time that class III gene expression can be regulated by a DNA binding protein with its target site in the 5'-flanking region and that a prokaryotic repressor can confer regulation of a suitably engineered tRNA gene.

Alleles↗