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The development of cancer is the result of a series of molecular changes occurring in the cell. These events lead to changes in the expression level of numerous genes that result in different phenotypic characteristics of tumors. In this report we describe the assembly and utilization of a 5766 member cDNA microarray to study the differences in gene expression between normal and neoplastic human ovarian...
A novel method to transform the nematode Caenorhabditis elegans is described. DNA coprecipitated with gold particles is shot at worms by means of a helium beam. Transformed worms are either identified by a dominant visible marker or selected by a conditional lethal system.
The genomic organization of the heparin cofactor II (HCII) gene from rat and mouse was investigated and compared with their human counterpart. The genes share a common core structure consisting of five exons interrupted by four introns, but the mouse and rat gene reveal individual additional features. A unique differentially spliced exon is present in the 5'-untranslated region of the rat gene, which...
Four cDNA clones, rcs1, rcs2, rcs3 and rcs4, encoding cysteine synthase [O-acetylserine(thiol)lyase] were isolated from rice. The predicted amino acid sequences contain the conserved PXXSVKDR region characteristic of cysteine synthase, which includes the lysine residue that binds the cofactor, pyridoxal 5'-phosphate. Molecular phylogenic analysis suggests that, whereas rcs1 and rcs3 belong to the...
6-Phosphofructo-2-kinase/fructose 2,6-bisphosphatase (PFK-2/FBPase-2) is a bifunctional enzyme responsible for the synthesis and breakdown of Fru-2,6-P 2 , a key metabolite in the regulation of glycolysis. Several genes encode distinct PFK-2/FBPase-2 isozymes that differ in their tissue distribution and enzyme regulation. In this paper, we present the isolation of a cDNA from a human testis...
Translation elongation factor 2 (eEF2) is a large protein of more than 800 amino acids which establishes complex interactions with the ribosome in order to catalyze the conformational changes needed for translation elongation. Unlike other yeasts, the pathogenic fungus Candida albicans was found to have a single gene encoding this factor per haploid genome, located on chromosome 2. Expression of this...
Three muscle type-specific troponin T (TnT) genes are present in vertebrate to encode a number of protein isoforms via alternative mRNA splicing. While the genomic structures of cardiac and fast skeletal muscle TnT genes have been documented, this study cloned and characterized the slow skeletal muscle TnT (sTnT) gene. Complete nucleotide sequence and genomic organization revealed that the mouse sTnT...
The processes by which the kinase inactive receptor ErbB-3 transmits the signals of its ligand, heregulin (HRG), are incompletely understood. We used a yeast two-hybrid system to identify ErbB-3 interacting proteins that may participate in HRG signal transduction. We found that the protein p23, the human homolog of the mouse transplantation antigen P198, interacted with the cytoplasmic domain of ErbB-3...
The tetracycline-responsive expression system is based on the ability of the chimeric tTA and rtTA transactivators to stimulate specifically transcription from a companion synthetic CMV* or TK* promoter element, and can provide tightly regulated gene expression that can be induced up to five orders of magnitude in cultured cells and transgenic mice. A major problem with the system is that high level...
Recombinant immunotoxins (rITs) are highly specific anti-tumor agents composed of monoclonal antibody fragments or other specific carriers coupled to plant or bacterial toxins. A major problem in the purification of rITs is the low periplasmic yield in currently available expression systems. Thus, the aim of this study was the development of a new bacterial expression system for high-level production...
In the frame of the European Network for Functional Analysis (EUROFAN), two regions from chromosome XI covering 54kb have been subjected to 'mass-murder'. Ten deletions covering 23 novel open reading frames (ORFs) were constructed in haploid and diploid strains. Six deletions were lethal in haploid strains. One deletion caused slow germination of spores and slow cellular growth, and another one was...
Two regulatory genes (lcoR and lcoS) were identified from a plasmid-borne lactococcal copper resistance determinant and characterized by transcriptional fusion to the promoterless chloramphenicol acetyltransferase gene (cat). RT-PCR analysis indicates that lcoR and lcoS are organized within an operon, controlling the transcription of cat in a copper-inducible manner. The amino acid sequences deduced...
The mechanisms responsible for the development of the taxol resistance phenotype are unclear, and are likely explained by multiple mechanisms. To understand the molecular changes associated with drug resistance more fully, a taxol-resistant subline, derived from the human ovarian cancer cell line SKOV-3, was established through selection by culture in incrementally increasing taxol concentrations...
A new cassette (Er-Cm cassette) and mini-transposon (mTn) (TnMaxErCm) based on the previously described mTn, TnMax2 [Haas et al., Gene 130, 23-31.], have been constructed. The cassette and mTn make use of an erythromycin resistance (Er R ) marker encoded by ermC'. Both the Er-Cm cassette and TnMaxErCm also carry a promoterless cat gene to allow the construction of transcriptional fusions and...
GBP28 is a novel human plasma gelatin-binding protein that is encoded by apM1 mRNA, expressed specifically in adipose tissue. Three overlapping clones (two lambda clones and one BAC clone) containing the human plasma gelatin-binding protein (GBP28) gene were isolated and characterized. The GBP28 gene spans 16kb and is composed of three exons from 18bp to 4277bp in size with consensus splice sites...
The murine transcription factor murine cyclin D-binding Myb-like protein (mDmp1) arrests the cell cycle in G 1 phase, through an activity that can be overridden by direct interaction with the D-type cyclins. Here, we describe the identification, sequence, chromosomal localization, and expression of the human cognate, hDMP1. The hDMP1 cDNA contains a 2280bp open reading frame that shares a...
We isolated novel temperature-sensitive mutants of spo0H, spo0H1 and spo0H5, having E61K and G30E amino-acid substitutions within the σ H protein, respectively, and located in the highly conserved region, 2, among prokaryotic sigma factors that participates in binding to core enzyme of RNA polymerase. These mutants showed a sporulation-deficient phenotype at 43 o C. Moreover,...
Two distinct calmodulin (CaM) genes are isolated from the ascidian, Halocynthia roretzi, (Hr-CaM A and Hr-CaM B) and those structures are determined. There are three nucleotide substitutions, producing two amino acid differences between Hr-CaM A and Hr-CaM B, and those are corresponding to two of the known eight variable residues among metazoan CaMs. Both Hr-CaM A and Hr-CaM B are constructed from...
The double-stranded RNA-specific adenosine deaminase (ADAR1) is inducible by interferon (IFN) and is implicated in the editing of viral RNAs during lytic and persistent infection. We have now isolated and characterized human genomic clones that contain the promoter region required for transcription of the ADAR1 gene. Rapid amplification of cDNA 5'-ends (5'-RACE) identified additional upstream exon...
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