Trypanosoma cruzi TcSRPK, the first protozoan member of the SRPK family, is biochemically and functionally conserved with metazoan SR protein-specific kinases.

A novel SR protein-specific kinase (SRPK) from the SRPK family was identified for the first time in a protozoan organism. The primary structure of the protein, named TcSRPK, presents a significant degree of identity with other metazoan members of the family. In vitro phosphorylation experiments showed that TcSRPK has the same substrate specificity relative to other SRPKs.

The SR protein SRp38 represses splicing in M phase cells.

SR proteins constitute a family of pre-mRNA splicing factors that play important roles in both constitutive and regulated splicing. Here, we describe one member of the family, which we call SRp38, with unexpected properties. Unlike other SR proteins, SRp38 cannot activate splicing and is essentially inactive in splicing assays.

Pelle kinase is activated by autophosphorylation during Toll signaling in Drosophila.

The Drosophila Pelle kinase plays a key role in the evolutionarily conserved Toll signaling pathway, but the mechanism responsible for its activation has been unknown. We present in vivo and in vitro evidence establishing an important role for concentration-dependent autophosphorylation in the signaling process. We first show that Pelle phosphorylation can be detected transiently in early embryos, concomitant with activation of signaling.

Autoubiquitination of the BRCA1*BARD1 RING ubiquitin ligase.

The RING finger of BRCA1 confers ubiquitin ligase activity that is markedly enhanced when complexed with another RING-containing protein, BARD1, and is required for the function of this tumor suppressor protein in protecting genomic integrity. Here, we report that co-expression of BRCA1-(1-639) and BARD1 in bacteria can assemble a potent ubiquitin ligase activity. Purified BRCA1-(1-639)*BARD1 stimulated the Ubc5c-mediated monoubiquitination of histone H2A/H2AX in vitro, suggesting a possible role for BRCA1*BARD1 in modifying chromatin structure.

Tehao functions in the Toll pathway in Drosophila melanogaster: possible roles in development and innate immunity.

Toll and related proteins play important roles in innate immunity in both invertebrates and vertebrates. In Drosophila melanogaster, Tehao shares a striking similarity in its intracellular domain with Toll. In this paper, we show that Tehao is expressed throughout development and appears to be glycosylated.

Stability of a PKCI-1-related mRNA is controlled by the splicing factor ASF/SF2: a novel function for SR proteins.

Pre-mRNA splicing is a widely used regulatory mechanism for controlling gene expression, and a family of conserved proteins, SR proteins, participate in both constitutive and alternative splicing. Here we describe a novel function for the SR protein ASF/SF2. We used an embryonic chicken cDNA library to screen for differential mRNA expression in the chicken B-cell line DT40-ASF, expressing or not expressing ASF/SF2.

Requirements of the RNA polymerase II C-terminal domain for reconstituting pre-mRNA 3' cleavage.

RNA polymerase II (RNAP II) has previously been shown to be required for the pre-mRNA polyadenylation cleavage reaction in vitro. This activity was found to reside solely in the C-terminal domain (CTD) of the enzyme's largest subunit. Using a deletion analysis of glutathione S-transferase-CTD fusion proteins, we searched among the CTD's 52 imperfectly repetitive heptapeptides for the minimal subset that possesses this property.

Why is p53 acetylated?

Recent studies suggest that acetylation of the p53 tumor suppressor protein is not important for its DNA binding activity, as was previously thought. We discuss here a number of theories as to how this modification may serve to regulate the protein's functions.

Splicing-related catalysis by protein-free snRNAs.

Removal of intervening sequences from eukaryotic messenger RNA precursors is carried out by the spliceosome, a complex assembly of five small nuclear RNAs (snRNAs) and a large number of proteins. Although it has been suggested that the spliceosome might be an RNA enzyme, direct evidence for this has been lacking, and the identity of the catalytic domain of the spliceosome is unknown. Here we show that a protein-free complex of two snRNAs, U2 and U6, can bind and position a small RNA containing the sequence of the intron branch site, and activate the branch adenosine to attack a catalytically critical domain of U6 in a reaction that is related to the first step of splicing.

Identification and functional characterization of neo-poly(A) polymerase, an RNA processing enzyme overexpressed in human tumors.

Poly(A) polymerase (PAP) plays an essential role in polyadenylation of mRNA precursors, and it has long been thought that mammalian cells contain only a single PAP gene. We describe here the unexpected existence of a human PAP, which we call neo-PAP, encoded by a previously uncharacterized gene. cDNA was isolated from a tumor-derived cDNA library encoding an 82.

Physical and functional interactions between Drosophila TRAF2 and Pelle kinase contribute to Dorsal activation.

Signaling through the Toll receptor is required for dorsal/ventral polarity in Drosophila embryos, and also plays an evolutionarily conserved role in the immune response. Upon ligand binding, Toll appears to multimerize and activate the associated kinase, Pelle. However, the immediate downstream targets of Pelle have not been identified.

Evolutionarily conserved interaction between CstF-64 and PC4 links transcription, polyadenylation, and termination.

Tight connections exist between transcription and subsequent processing of mRNA precursors, and interactions between the transcription and polyadenylation machineries seem especially extensive. Using a yeast two-hybrid screen to identify factors that interact with the polyadenylation factor CstF-64, we uncovered an interaction with the transcriptional coactivator PC4. Both human proteins have yeast homologs, Rna15p and Sub1p, respectively, and we show that these two proteins also interact.

Early transvaginal ultrasonography versus early cerclage in women with an unclear history of incompetent cervix.

Objective: Our aim was to compare outcomes in women with a questionable history of incompetent cervix, followed up with early transvaginal ultrasonography, with outcomes in women who had early cerclage.

Study Design: Charts were reviewed and patients identified for incompetent cervix from our obstetric database from 1995 through 1997. We included women who had an unclear history of incompetent cervix as follows: second-trimester loss or termination, > or =3 first-trimester terminations, cone biopsy or loop electrosurgical excision, or exposure to diethylstilbestrol.

Porphyrins with exocyclic rings. 16. Synthesis and spectroscopic characterization of fluoranthoporphyrins, a new class of highly conjugated porphyrin chromophores.

Porphyrins with fused aromatic rings are under detailed investigation due to their unique spectroscopic properties. To gain more insights into the effects due to ring annealation on the porphyrin chromophore, a series of fluoranthoporphyrins have been synthesized. Reaction of 3-nitrofluoranthene with isocyanoacetate esters in the presence of a phosphazene base afforded good yields of the fluorantho[2,3-c]pyrrole esters 8.

A randomized comparison of transcervical Foley catheter to intravaginal misoprostol for preinduction cervical ripening.

Objective: To compare the efficacy of intravaginal misoprostol tablets with transcervical Foley catheter for preinduction cervical ripening.

Methods: Pregnant women who presented for induction of labor with unfavorable cervices (Bishop score less than 6) were assigned randomly to intravaginal misoprostol (50 microg tablet every 4 hours for a maximum of six doses) or 30-mL Foley catheter placed transcervically with maintenance of traction.

Results: Among 111 women, 53 were allocated to misoprostol and 58 to Foley bulb.

How echogenic is echogenic? Quantitative acoustics of the renal cortex.

The echogenicity of the cortex is an important parameter in interpreting renal sonograms that suggest changes in cortical structure. Echogenicity is currently measured qualitatively, and no attempts have been made at quantification. We developed a method to quantify renal cortical echogenicity in reference to the liver and evaluated its reproducibility, dependence on scanning variables, and potential utility.

Heterozygous disruption of the TATA-binding protein gene in DT40 cells causes reduced cdc25B phosphatase expression and delayed mitosis.

TATA-binding protein (TBP) is a key general transcription factor required for transcription by all three nuclear RNA polymerases. Although it has been intensively analyzed in vitro and in Saccharomyces cerevisiae, in vivo studies of vertebrate TBP have been limited. We applied gene-targeting techniques using chicken DT40 cells to generate heterozygous cells with one copy of the TBP gene disrupted.

The BARD1-CstF-50 interaction links mRNA 3' end formation to DNA damage and tumor suppression.

The mRNA polyadenylation factor CstF interacts with the BRCA1-associated protein BARD1, and this interaction represses the nuclear mRNA polyadenylation machinery in vitro. Given the suspected role of BRCA1/BARD1 in DNA repair, we tested whether inhibition of mRNA processing is linked to DNA damage. Strikingly, we found that 3' cleavage in extracts from cells treated with hydroxyurea or ultraviolet light was strongly, but transiently, inhibited.

The Drosophila TATA binding protein contains a strong but masked activation domain.

TATA binding protein (TBP) is a critical transcription factor involved in transcription by all three RNA polymerases (RNAPs). Studies using in vitro systems and yeast have shown that the C-terminal core domain (CTD) of TBP is necessary and sufficient for many TBP functions, but the significance of the N-terminal domain (NTD) of TBP is still obscure. Here, using transient expression assays in Drosophila Schneider cells, we show that the NTD of Drosophila TBP (dTBP) strongly activates transcription when fused to the GAL4 DNA binding domain (DBD).

Intracervical fibrin sealants: a potential treatment for early preterm premature rupture of the membranes.

Objective: We report our experience with a transvaginally applied intracervical fibrin sealant at <24 weeks' gestation.

Study Design: This is an observational study of a referred patient population, with preterm premature rupture of the membranes at <24 weeks' gestation.

Results: Twelve women consented to our protocol.

Phosphorylation of CPEB by Eg2 mediates the recruitment of CPSF into an active cytoplasmic polyadenylation complex.

The release of Xenopus oocytes from prophase I arrest is largely driven by the cytoplasmic polyadenylation-induced translation of dormant maternal mRNAs. Two cis elements, the CPE and the hexanucleotide AAUAAA, and their respective binding factors, CPEB and a cytoplasmic form of CPSF, control polyadenylation. The most proximal stimulus for polyadenylation is Eg2-catalyzed phosphorylation of CPEB serine 174.

Structural basis for signal transduction by the Toll/interleukin-1 receptor domains.

Toll-like receptors (TLRs) and the interleukin-1 receptor superfamily (IL-1Rs) are integral to both innate and adaptive immunity for host defence. These receptors share a conserved cytoplasmic domain, known as the TIR domain. A single-point mutation in the TIR domain of murine TLR4 (Pro712His, the Lps(d) mutation) abolishes the host immune response to lipopolysaccharide (LPS), and mutation of the equivalent residue in TLR2, Pro681His, disrupts signal transduction in response to stimulation by yeast and gram-positive bacteria.

The ends of the affair: capping and polyadenylation.

Nearly all mRNAs are post-transcriptionally modified at their 5' and 3' ends, by capping and polyadenylation, respectively. These essential modifications are of course chemically quite distinct, as are the enzymatic complexes responsible for their synthesis. But recent studies have uncovered some similarities as well.