Acute intermittent porphyria: alternative splicing of hydroxymethylbilane synthase mRNA excludes exons 3 and 12

The hydroxymethylbilane synthase (HMBS) mRNAs from 44 control individuals and 30 patients suffering from acute intermittent porphyria (AIP), were screened for length differences by reverse transcriptase polymerase chain reaction (RT-PCR) and any abnormalities were characterized by direct sequencing. Examination of the mRNAs extracted from the peripheral blood lymphocytes of the samples revealed varying degrees of alternative splicing, involving the removal of exons 3 and 12. Approximately 10-50% of the mRNA molecules were affected, despite the absence of genomic splice site mutations or any major deviance from consensus splice sequence values. The preliminary data obtained from this study suggest that this event is a normal occurrence in peripheral blood lymphocytes, and may not be associated with the molecular pathology responsible for AIP. (C) 1998 Academic Press Limited.

Human GC-AG alternative intron isoforms with weak donor sites show enhanced consensus at acceptor exon positions

It has been previously observed that the intrinsically weak variant GC donor sites, in order to be recognized by the U2-type spliceosome, possess strong consensus sequences maximized for base pair formation with U1 and U5/U6 snRNAs. However, variability in signal strength is a fundamental mechanism for splice site selection in alternative splicing. Here we report human alternative GC-AG introns (for the first time from any species), and show that while constitutive GC-AG introns do possess strong signals at their donor sites, a large subset of alternative GC-AG introns possess weak consensus sequences at their donor sites. Surprisingly, this subset of alternative isoforms shows strong consensus at acceptor exon positions 1 and 2. The improved consensus at the acceptor exon can facilitate a strong interaction with U5 snRNA, which tethers the two exons for ligation during the second step of splicing. Further, these isoforms nearly always possess alternative acceptor sites and always possess alternative acceptor sites and exhibit particularly weak polypyrimidine tracts characteristic of AG-dependent introns. The acceptor exon nucleotides are part of the consensus required for the U2AF(35)-mediated recognition of AG in such introns. Such improved consensus at acceptor exons is not found in either normal or alternative GT-AG introns having weak donor sites or weak polypyrimidine,tracts. The changes probably reflect mechanisms that allow GC-AG alternative intron isoforms to cope with two conflicting requirements, namely an apparent need for differential splice strength to direct the choice of alternative sites and a need for improved donor signals to compensate for the central mismatch base pair (C-A) in the RNA duplex of U1 snRNA and the pre-mRNA. The other important findings include (i) one in every twenty alternative introns is a GC-AG intron, and (ii) three of every five observed GC-AG introns are alternative isoforms.

Impact of alternative initiation, splicing, and termination on the diversity of the mRNA transcripts encoded by the mouse transcriptome

We analyzed the FANTOM2 clone set of 60,770 RIKEN full-length mouse cDNA sequences and 44,122 public mRNA sequences. We developed a new computational procedure to identify and classify the forms of splice variation evident in this data set and organized the results into a publicly accessible database that can be used for future expression array construction, structural genomics, and analyses of the mechanism and regulation of alternative splicing. Statistical analysis shows that at least 41% and possibly as much as 60% of multiexon genes in mouse have multiple splice forms. Of the transcription units with multiple splice forms, 49% contain transcripts in which the apparent use of an alternative transcription start (stop) is accompanied by alternative splicing of the initial (terminal) exon. This implies that alternative transcription may frequently induce alternative splicing. The fact that 73% of all exons with splice variation fall within the annotated coding region indicates that most splice variation is likely to affect the protein form. Finally, we compared the set of constitutive (present in all transcripts) exons with the set of cryptic (present only in some transcripts) exons and found statistically significant differences in their length distributions, the nucleoticle distributions around their splice junctions, and the frequencies of occurrence of several short sequence motifs.

ASD: the Alternative Splicing Database

Alternative splicing is widespread in mammalian gene expression, and variant splice patterns are often specific to different stages of development, particular tissues or a disease state. There is a need to systematically collect data on alternatively spliced exons, introns and splice isoforms, and to annotate this data. The Alternative Splicing Database consortium has been addressing this need, and is committed to maintaining and developing a value-added database of alternative splice events, and of experimentally verified regulatory mechanisms that mediate splice variants. In this paper we present two of the products from this project: namely, a database of computationally delineated alternative splice events as seen in alignments of EST/cDNA sequences with genome sequences, and a database of alternatively spliced exons collected from literature. The reported splice events are from nine different organisms and are annotated for various biological features including expression states and cross-species conservation. The data are presented on our ASD web pages (http://www.ebi.ac.uk/asd).

Genomic organization of human arylamine N-acetyltransferase Type I reveals alternative promoters that generate different 5'-UTR splice variants with altered translational activities

In humans, a polymorphic gene encodes the drug-metabolizing enzyme NATI (arylamine N-acetyltransferase Type 1), which is widely expressed throughout the body. While the protein-coding region of NATI is contained within a single exon, examination of the human EST (expressed sequence tag) database at the NCBI revealed the presence of nine separate exons, eight of which were located in the 5'non-coding region of NATI. Differential splicing produced at least eight unique mRNA isoforms that could be grouped according to the location of the first exon, which suggested that NATI expression occurs from three alternative promoters. Using RT (reverse transcriptase)-PCR, we identified one major transcript in various epithelial cells derived from different tissues. In contrast, multiple transcripts were observed in blood-derived cell lines (CEM, THP-1 and Jurkat), with a novel variant, not identified in the EST database, found in CEM cells only. The major splice variant increased gene expression 9-11-fold in a luciferase reporter assay, while the other isoforrns were similar or slightly greater than the control. We examined the upstream region of the most active splice variant in a promoter-reporter assay, and isolated a 257 bp sequence that produced maximal promoter activity. This sequence lacked a TATA box, but contained a consensus Sp1 site and a CAAT box, as well as several other putative transcription-factor-binding sites. Cell-specific expression of the different NATI transcripts may contribute to the variation in NATI activity in vivo.

The use of complementary and alternative therapies by patients with cancer

The aim of this research was to assess the prevalence and predictors of complementary and alternative therapy (CAT) use among cancer patients in Australia. A total of 1492 cancer patients attending nine major public cancer treatment centers in New South Wales, Australia, were asked to complete the Supportive Care Needs Survey. Of the 1354 consenting patients, 888 (65%) returned a completed survey. This article reports the secondary analyses of the survey data, specifically focusing on CAT use. For all cancers, 17.1% of patients were using at least one CAT. The two main demographic characteristics of CAT users were gender and age, where females were more likely to use CAT than males and that CAT use declined as age increased. Time since diagnosis was identified as the only significant clinical predictor of CAT use, where CAT use increased with time until 5 years since diagnosis. Our research shows that herbal treatments and naturopathy are the most popular CAT used by cancer patients (constituting over 30% of all CAT use recorded). The use of CAT among cancer patients is a significant issue in cancer care, especially considering the potential interactions between CAT and conventional medicines. Given that many cancer patients may not be aware of potential risks associated with these interactions it is important that oncologists and others involved in cancer patient care are informed about CAT and its use amongst their patients.

Demonstrators and Semper press car during Moratorium in Brisbane 1970

Demonstrators in Brisbane, Australia with banners and flags during Moratorium march 1970. Semper press car (Volkswagon Kombi) can be seen. Protestors are outside the Presbyterian Church of Queensland (now Uniting Church) in Ann Street, Brisbane. In the background, scaffolding can be seen at the construction site of the Crest Hotel. Semper Floreat is the student newspaper of the University of Queensland.

The Swiss heroin trials: testing alternative approaches

Over half a million heroin misusers receive oral methadone maintenance treatment world-wide1 but the maintenance prescription of injectable opioid drugs, like heroin, remains controversial. In 1992 Switzerland began a large scale evaluation of heroin and other injectable opiate prescribing that eventually involved 1035 misusers. 2 3 The results of the evaluation have recently been reported.4 These show that it was feasible to provide heroin by intravenous injection at a clinic, up to three times a day, for seven days a week. This was done while maintaining good drug control, good order, client safety, and staff morale. Patients were stabilised on 500 to 600 mg heroin daily without evidence of increasing tolerance. Retention in treatment was 89% at six months and 69% at 18 months.4 The self reported use of non-prescribed heroin fell signifianctly, but other drug use was minimally affected. The death rate was 1% per year, and there were no deaths from overdose among participants . . . [Full text of this article]

Effect of oral creatine supplementation on near-maximal strength and repeated sets of high-intensity bench press exercise

This study examined the effects of 26 days of oral creatine monohydrate (Cr) supplementation on near-maximal muscular strength, high-intensity bench press performance, and body composition. Eighteen male powerlifters with at least 2 years resistance training experience took part in this 28-day experiment. Pre and postmeasurements (Days 1 and 28) were taken of near-maximal muscular strength, body mass, and % body fat. There were two periods of supplementation Days 2 to 6 and Days 7 to 27. ANOVA and t-tests revealed that Cr supplementation significantly increased body mass and lean body mass with no changes in % body fat. Significant increases in 3-RM strength occurred in both groups, both absolute and relative to body mass; the increases were greater in the Cr group. The change in total repetitions also increased significantly with Cr supplementation both in absolute terms and relative to body mass, while no significant change was seen in the placebo (P) group. Creatine supplementation caused significant changes in the number of BP reps in Sets 1, 4, and 5. No changes occurred in the P group. It appears that 26 days of Cr supplementation significantly improves muscular strength and repeated near-maximal BP performance, and induces changes in body composition.