The positive-acting sulfur regulatory protein CYS3 of Neurospora crassa: nuclear localization, autogenous control, and regions required for transcriptional activation

The positive-acting global sulfur regulatory protein, CYS3, of Neurospora crassa turns on the expression of a family of unlinked structural genes that encode enzymes of sulfur catabolism. CYS3 contains a leucine zipper and an adjacent basic region (b-zip), which together constitute a bipartite sequence-specific DNA-binding domain. Specific anti-CYS3 antibodies detected a protein of the expected size in nuclear extracts of wild-type Neurospora under conditions in which the sulfur circuit is activated. The CYS3 protein was not observed in cys-3 mutants. Nuclear extracts of wild type, but not cys-3 mutants, also showed specific DNA-binding activity identical to that obtained with a CYS3 protein expressed in Escherichia coli. A truncated CYS3 protein that contains primarily the b-zip domain binds to DNA with high specificity and affinity in vitro, yet fails to activate gene expression in vivo, and instead inhibits the function of the wild-type CYS3 protein. Amino-terminal, carboxyterminal, and internal deletions as well as alanine scanning mutagenesis were employed to identify regions of the CYS3 protein that are required for its trans-activation function. Regions of CYS3 carboxy terminal to the b-zip motif are not completely essential for function although loss of an alanine-rich region results in decreased activity. All deletions amino terminal to the b-zip motif led to a complete loss of CYS3 function. Alanine scanning mutagenesis demonstrated that an unusual prolinerich domain of CYS3 appears to be very important for function and is presumed to constitute an activation domain. It is concluded that CYS3 displays nuclear localization and positive autogenous control in Neurospora and functions as a trans-acting DNA-binding protein.     

Aneurysms of splanchnic arteries

Aneurysms of splanchnic arteries represent an uncommon but important vascular disease, which many times presents itself as clinical emergency and often results in death. 11 patients with splanchnic aneurysms were treated in our Department during the last 15 years. These aneurysms were located in 5 cases in splenic artery, 4 cases in hepatic artery, 1 case in celiac axis and 1 case in right gastroepiploic artery. Surgical treatment of these aneurysms was successful in all but one patient (he died from rupture of a hepatic artery aneurysm). Giving an overall mortality similar to that reported in the literature. The treatment of these aneurysms is discussed, while literature about this uncommon disease is reviewed.     

Characterization of a baculovirus-encoded ATP-dependent DNA ligase

Sequence analysis of the Lymantria dispar multicapsid nucleopolyhedrovirus (LdMNPV) genome identified an open reading frame (ORF) encoding a 548-amino-acid (62-kDa) protein that showed 35% amino acid sequence identity with vaccinia virus ATP-dependent DNA ligase. Ligase homologs have not been reported from other baculoviruses. The ligase ORF was cloned and expressed as an N-terminal histidine-tagged fusion protein. Incubation of the purified protein with [alpha-32P]ATP resulted in formation of a covalent enzyme-adenylate intermediate which ran as a 62-kDa labeled band on a sodium dodecyl sulfate-polyacrylamide gel. Loss of the radiolabeled band occurred upon incubation of the intermediate with pyrophosphate, poly(dA) . poly(dT)12-18, or poly(rA) . poly(dT)12-18, characteristics of a DNA ligase II or III. The protein was able to ligate a double-stranded synthetic DNA substrate containing a single nick and inefficiently ligated a 1-nucleotide (nt) gap but did not ligate a 2-nt gap. It was able to ligate short, complementary overhangs but not blunt-ended double-stranded DNA. In a transient DNA replication assay employing six plasmids containing the LdMNPV homologs of the essential baculovirus replication genes, a plasmid containing the DNA ligase gene was neither essential nor stimulatory. All of these results are consistent with the activity of type III DNA ligases, which have been implicated in DNA repair and recombination.     

The key informant technique

The clinical outcome of 152 patients aged 65 years or over who were referred to the author's institute between August 1990 and August 1991 with certain specified gastrointestinal malignancies and acute, life-threatening abdominal conditions, were audited concurrently. Two groups were considered: patients aged 65-79 years and those over 80 years. The mortality rate within 30 days of surgery was 14 per cent in both age groups, although significantly fewer patients aged over 80 years (35 of 54) were considered suitable for surgery than in the 65-79 years age group (84 of 98) (0.01 > P > 0.001). Morbidity after operation and cost of treatment were not significantly different between the two groups. Two years after surgery 40 per cent of the patients aged over 80 years and 58 per cent of those aged 65-79 years were alive. Quality of life in these survivors was good with 85 per cent of those aged over 80 years living at home and 72 per cent fit enough to undertake light work.     

A region within murine chromosome 7F4, syntenic to the human 11q13 amplicon, is frequently amplified in 4NQO-induced oral cavity tumors

Our previous reports have shown that two thirds of 4-nitroquinoline-1-oxide (4NQO)-induced murine oral squamous cell carcinomas (SCC) have Hras1 mutations. Loss of heterozygosity (LOH) involving the distal portion of chromosome (Chr) 7 occurred in half of the tumors with Hras1 mutations. Here, we demonstrate that five of six tumors with LOH have 4-8-fold amplification involving the distal portion of Chr 7 (7F4). Ccnd1. Fgf4 and Fgf3, within the most telomeric region of Chr 7 (70.5 cM), are co-amplified. The region is syntenic to a previously identified human amplicon at 11q13. Only one out of eight tumors without LOH at Chr 7 had twofold amplification; the other seven had no detectable amplification. Significant amplification is restricted to the chromosome with the Hras1 mutation. Gene amplification occurred without overexpression since only one of five tumors with amplification and one of six tumors without Ccnd1 amplification expressed increased protein. Although amplification of 11q13 occurs rather frequently in human tumors, 4NQO-induced oral cavity tumors in inbred mice are the first example of a murine tumor with consistent amplification. Our observations are strikingly similar to human head and neck SCC where overexpression of genes within the 11q13 amplicon is inconsistently detected. The amplification of genes localized to human 11q13 and the syntenic region on murine Chr 7 during tumorigenesis suggests that similar structural elements are present which predispose these regions to amplification during malignant transformation.     

—Review Paper—Current State-Of-The-Art Measurements Of Ozone In The Gas Phase And In Solution

This paper is a critical review and summary of the analytical procedures currently used by operating water utilities to control ozone treatment processes, considering disinfection as well as the many oxidative applications of ozone in water treatment applications. The role of common interferences in the various methods is described.

In operating U S., Canadian, European, and Japanese water treatment plants currently employing ozone as a primary disinfectant, ozone nearly always is followed by the addition of small quantities of chlorine, chlorine dioxide, or chloramine to provide the residual in the distribution system and to act as a secondary disinfectant. Clearly, the analytical procedures used must distinguish between the various species. This review critically presents these various factors and their implication with respect to “true values” of ozone determined under widely varying conditions.