Two evolutionarily conserved repression domains in the Drosophila Kruppel protein differ in activator specificity.

To identify biologically functional regions in the product of the Drosophila melanogaster gene Kruppel, we cloned the Kruppel homolog from Drosophila virilis. Both the previously identified amino (N)-terminal repression region and the DNA-binding region of the D. virilis Kruppel protein are greater than 96% identical to those of the D.

Two N-terminal self-association domains are required for the dominant negative transcriptional activity of WT1 Denys-Drash mutant proteins.

Patients with Denys-Drash syndrome (DDS) have been shown to be constitutionally heterozygous for mutations of the WT1 gene. Almost all DDS mutations inactivate or remove the DNA-binding zinc finger region of WT1 and the resulting mutant proteins appear to act in a dominant negative manner. This may occur via WT1 self-association, which has been shown to involve the first 180 amino acids.

A novel repressor, par-4, modulates transcription and growth suppression functions of the Wilms' tumor suppressor WT1.

The tumor suppressor WT1 represses and activates transcription. The loss and/or imbalance of the dual transcriptional activity of WT1 may contribute to Wilms' tumor. In this study, we identified par-4 (for prostate apoptosis response) as a WT1-interacting protein that itself functions as a transcriptional repressor.

The online patient meeting.

Studies have shown the importance of social support for health, and the value of patient support groups. Today we are seeing a further development of the idea of the patient meeting: online discussions on the internet and other computer networks. This paper reports on the online activities of patients with amyotrophic lateral sclerosis.

Amino-terminal protein-protein interaction motif (POZ-domain) is responsible for activities of the promyelocytic leukemia zinc finger-retinoic acid receptor-alpha fusion protein.

Promyelocytic leukemia zinc finger-retinoic acid receptor a (PLZF-RARalpha), a fusion receptor generated as a result of a variant t(11;17) chromosomal translocation that occurs in a small subset of acute promyelocytic leukemia (APL) patients, has been shown to display a dominant-negative effect against the wild-type RARalpha/retinoid X receptor alpha (RXRalpha). We now show that its N-terminal region (called the POZ-domain), which mediates protein-protein interaction as well as specific nuclear localization of the wild-type PLZF and chimeric PLZF-RARalpha proteins, is primarily responsible for this activity. To further investigate the mechanisms of PLZF-RARalpha action, we have also studied its ligand-receptor, protein-protein, and protein-DNA interaction properties and compared them with those of the promyelocytic leukemia gene (PML)-RARalpha, which is expressed in the majority of APLs as a result of t(15;17) translocation.

Reduced and altered DNA-binding and transcriptional properties of the PLZF-retinoic acid receptor-alpha chimera generated in t(11;17)-associated acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) associated with chromosomal rearrangement t(11;17) is a distinct syndrome which, unlike typical t(15;17) APL, fails to respond to all-trans retinoic acid (ATRA) therapy. In t(11;17) the PLZF gene, encoding a Krüppel-like zinc finger protein, is fused to the retinoic acid receptor-alpha (RAR alpha) gene, yielding two classes of chimeric proteins. PLZF protein was found in the nucleus in a punctate speckled pattern that differed from the nuclear body expression pattern of the PML protein affected in t(15;17) APL.

Leukemia translocation gene, PLZF, is expressed with a speckled nuclear pattern in early hematopoietic progenitors.

The PLZF gene was discovered by studying a rearrangement of the RAR alpha locus in a patient with acute promyelocytic leukemia and a t(11;17) chromosomal translocation. To understand further the potential role(s) of the PLZF gene product in hematopoiesis, we have examined its expression levels in a variety of murine tissues and in established cell lines that are representative of various stages of myeloid and lymphoid development. We show that murine PLZF(mPLZF) is expressed at the highest levels in undifferentiated, multipotential hematopoietic progenitor cells and that its expression declines as cells become more mature and committed to various hematopoietic lineages.

The transcriptional effect of WT1 is modulated by choice of expression vector.

The WT1 Wilms' tumor suppressor gene encodes a zinc finger transcription factor which plays a critical role in renal and genitourinary development. The WT1 protein was reported to both activate and repress transcription. We found that the transcriptional effect of WT1 on the Egr1 promoter could be modulated by the use of expression vectors containing different promoters.

The tumor suppressor gene WT1 inhibits ras-mediated transformation.

Wilms' tumor belongs to a small group of pediatric neoplasms that have served as paradigms of human cancers in which recessive mutations play a primary role in tumorigenesis. WT1 is a candidate tumor suppressor gene that is mutationally inactivated in a proportion of both familial and sporadic Wilms' tumors. Recent studies demonstrated that WT1 can partially suppress growth of a Wilms' tumor cell line in vitro and in vivo.

WT1-mediated transcriptional activation is inhibited by dominant negative mutant proteins.

The WT1 tumor suppressor gene encodes four isoforms of a zinc finger transcription factor with both activation and repression functions which are dependent upon promoter architecture. Using a simple HSV-tk promoter containing 5'-Egr-1/WT1-binding sites, we found that WT1 isoforms (A) and (B) strongly activated transcription. WT1(A) and (B) bound equally well to the Egr-1/WT1-binding site, but WT1(B), which contains a 17 amino acid insertion compared to WT1(A), was a consistently stronger activator of transcription than WT1(A).

Expression of the zinc-finger gene PLZF at rhombomere boundaries in the vertebrate hindbrain.

To investigate the potential biological role(s) of the PLZF gene, discovered as a fusion with the RARA locus in a patient with acute promyelocytic leukemia harboring a t(11;17) chromosomal translocation, we have isolated its murine homologue (mPLZF) and studied its patterns of developmental expression. The levels of mPLZF mRNAs increased perinatally in the liver, heart, and kidney, but with the exception of the heart, they were either absent or very low in the adult tissues. In situ analysis of mPLZF expression in mouse embryos between 7.

Clinical and molecular characterization of a rare syndrome of acute promyelocytic leukemia associated with translocation (11;17).

Analysis of a variant translocation t(11;17) in a case of acute promyelocytic leukemia (APL) led to discovery of a novel zinc finger gene, PLZF, fused to the retinoic acid receptor-alpha (RAR alpha) gene. We reviewed the clinical and molecular features of five additional patients with t(11;17)-associated APL. The clinical course of three patients was characterized by early death and three experienced disseminated intravascular coagulation.

Retinoic acid is required for and potentiates differentiation of acute promyelocytic leukemia cells by nonretinoid agents.

Patients with acute promyelocytic leukemia (APL) associated with the t(15;17) translocation and fusion of the promyelocytic leukemia (PML) and retinoic acid receptor-alpha (RAR-alpha) genes achieve complete remission but not cure with all-trans retinoic acid (RA), NB4, a cell line derived from a patient with t(15;17) APL that undergoes granulocytic differentiation when treated with pharmacologic doses of RA, was used as a model for differentiation therapy of APL. We found that NB4 cells are resistant to differentiation by nonretinoid inducers such as hexamethylene bisacetamide (HMBA), butyrates, vitamin D3, or hypoxanthine, all of which can induce differentiation in the commonly used HL60 leukemia cell line. Preexposure of NB4 cells to low concentrations of RA for a period as short as 30 minutes abolished resistance to nonretinoids and potentiated differentiation.

HLA-DR-, CD33+, CD56+, CD16- myeloid/natural killer cell acute leukemia: a previously unrecognized form of acute leukemia potentially misdiagnosed as French-American-British acute myeloid leukemia-M3.

We have identified and characterized a previously unrecognized form of acute leukemia that shares features of both myeloid and natural killer (NK) cells. From a consecutive series of 350 cases of adult de novo acute myeloid leukemia (AML), we identified 20 cases (6%) with a unique immunophenotype: CD33+, CD56+, CD11a+, CD13lo, CD15lo, CD34+/-, HLA-DR-, CD16-. Multicolor flow cytometric assays confirmed the coexpression of myeloid (CD33, CD13, CD15) and NK cell-associated (CD56) antigens in each case, whereas reverse transcription polymerase chain reaction (RT-PCR) assays confirmed the identity of CD56 (neural cell adhesion molecule) in leukemic blasts.

Mapping and mutagenesis of the amino-terminal transcriptional repression domain of the Drosophila Krüppel protein.

We previously demonstrated that the Drosophila Krüppel protein is a transcriptional repressor with separable DNA-binding and transcriptional repression activities. In this study, the minimal amino (N)-terminal repression region of the Krüppel protein was defined by transferring regions of the Krüppel protein to a heterologous DNA-binding protein, the lacI protein. Fusion of a predicted alpha-helical region from amino acids 62 to 92 in the N terminus of the Krüppel protein was sufficient to transfer repression activity.

Phase IV trial of daily oral etoposide in the treatment of advanced soft-tissue sarcoma.

Intravenous-bolus etoposide has modest activity in sarcomas when given daily for 3-5 days. Low frequent doses theoretically inhibit topoisomerase II activity over a longer duration and have been reported to have increased activity in small-cell lung cancer. A phase I trial of oral etoposide resulted in partial responses in two patients with soft-tissue sarcomas.

Predictive statistics and artificial intelligence in the U.S. National Cancer Institute's Drug Discovery Program for Cancer and AIDS.

The National Cancer Institute's drug discovery program screens more than 20,000 chemical compounds and natural products a year for activity against a panel of 60 tumor cell lines in vitro. The result is an information-rich database of patterns that form the basis for what we term an "information-intensive" approach to the process of drug discovery. The first step was a demonstration, both by statistical methods (including the program COMPARE) and by neural networks, that patterns of activity in the screen can be used to predict a compound's mechanism of action.

Selective repression of transcriptional activators at a distance by the Drosophila Krüppel protein.

The Krüppel (Kr) protein, bound at kilobase distances from the start site of transcription, represses transcription by RNA polymerase II in mammalian cells. Repression is monotonically dependent on the dose of Kr protein and the presence of Kr binding site(s) on the DNA. These data suggest an inhibitory protein-protein interaction between the Kr protein and proximal transcription factors.

NGFIA (EGR1) contains transcription activating domains in both the amino terminal and carboxyl terminal regions of the protein.

We constructed and tested a number of lac repressor fusion proteins containing various portions of the zinc-finger containing protein NGFIA for their ability to stimulate transcription of a reporter gene containing lac operators. NGFIA contains two transcription activation regions, found in two distinct regions of the protein. The carboxyl (C) terminal portion of the molecule contains a weak activation domain, including five tandem copies of an eight amino acid repeat (T/S,T/S,F/Y,P,S,P,X,X).

Effect of mecillinam on peptidoglycan synthesis during the division cycle of Salmonella typhimurium 2616.

The effects of mecillinam, ampicillin and cephalexin on peptidoglycan synthesis in Salmonella typhimurium 2616 have been studied at equivalent concentrations or "isoactivities". Using antibiotics at isoactivities allows a direct comparison of the biochemical effects of different antibiotics. When mecillinam was added at different times during the division cycle at a concentration that produced 50% inhibition of peptidoglycan synthesis in an exponential culture over a short period of time, the inhibition of synthesis was greatest in the newborn cells and least in the dividing cells.

Phase II trial of trimetrexate in patients with advanced soft-tissue sarcoma.

Trimetrexate, a lipophilic, 2,4-diaminoquinazoline derivative of methotrexate, enters cells by passive diffusion rather than via a transport system. Trimetrexate has shown promising activity in animal model systems. A total of 16 patients with metastatic soft-tissue sarcoma who had received only one prior chemotherapy regimen were treated with trimetrexate (8 mg/m2 given intravenously daily for 5 days) every 3 weeks.

Vasculitis with recurrent pulmonary hemorrhage in a long-term survivor after autologous bone marrow transplantation.

We report an unusual vasculitic syndrome in a long-term survivor of autologous bone marrow transplant. Clinical and pathologic studies revealed a cutaneous and pulmonary leukocytoclastic vasculitis complicated by recurrent pulmonary hemorrhage. Serologic studies revealed an elevated anti-neutrophil cytoplasmic antibody titer.

Treatment of low-grade and intermediate-grade lymphoma with intensive combination chemotherapy results in long-term, disease-free survival.

To define the role of intensive combination chemotherapy in the treatment of low-grade or intermediate-grade lymphomas, the authors report results in 49 patients treated with intermediate-dose or high-dose methotrexate, bleomycin, Adriamycin (doxorubicin; Adria Laboratories, Columbus, OH), cytoxan (cyclophosphamide), vincristine, and dexamethasone (m/M-BACOD) with long-term follow-up. The complete response rate was 59% (29 of 49), including 67% (eight of 12) with low-grade and 57% (21 of 37) with intermediate-grade disease. The median survival for the entire group was 81 months.