RNA editing, introns and evolution

Caries activity tests are an essential part of any programme concerned with the study or treatment of dental caries. Most of the caries activity tests require expensive kits or specially prepared media or facilities for incubation which limits their use in clinical practice. The present study sample consisted of twenty five children with dental caries and twenty five controls, free of caries, gingivitis and other oral ailments. The test is simple and consists of rinsing the mouth with 10 ml of sterile milk, 3 ml of which is mixed with 0.12 ml of 0.1% methylene blue dye and observed for colour change. The present study demonstrated the potential uses of this test in a pediatric dental clinic setup.     

Coincident loss of consciousness and ventricular tachycardia during +GZ stress

The environment of the advanced fighter aircraft represents a unique combination of stressful factors. Each of the individual stresses is hazardous, with the summation of these factors possibly resulting in an additional risk for sudden in-flight incapacitation. Advanced fighter aircraft are capabble of producing both rapid onset and high sustained +GZ forces which, on occasion, can exceed the tolerance limits of the pilot. The +GZ forces encountered during aerial combat maneuvering are physiologically stressful and have a profound effect on the regulatory mechanisms of the body. The influence of these stresses, including +GZ stress, on the autonomic nervous system is complex. The overall normal regulation of the cardiovascular system depends on a balance between both branches of the autonomic nervous system. An imbalance between sympathetic and parasympathetic tone can result in cardiac dysrhythmias and symptoms not conducive to safe and effective flight. An episode of ventricular tachycardia, coincident with an episode of loss of consciousness, was observed in an apparently healthy aircrewman during +GZ stress on the USAF School of Aerospace Medicine human centrifuge. The implications of autonomic imbalance in the production of similar potentially hazardous dysrhythmias and symptoms in the multistress environment deserve more in-depth investigation.     

Coincident DiGeorge anomaly and renal agenesis and its relation to maternal diabetes

We describe 2 cases of DiGeorge anomaly with bilateral renal agenesis-one, who also had hemivertebrae, in an infant of an insulin-dependent diabetic mother (IDDM). In a review we identified 2 other instances of this combination, both in IDDMs. The currently accepted notion that DiGeorge anomaly and renal agenesis are developmental field defects, coupled with their coincidence in IDDM, suggests that this combination is an association.     

The recent origins of spliceosomal introns revisited

Does the intron/exon structure of eukaryotic genes belie their ancient assembly by exon-shuffling or have introns been inserted into preformed genes during eukaryotic evolution? These are the central questions in the ongoing 'introns-early' versus 'introns-late' controversy. The phylogenetic distribution of spliceosomal introns continues to strongly favor the intronslate theory. The introns-early theory, however, has claimed support from intron phase and protein structure correlations.     

Grain-boundary structures in hexagonal materials: Coincident and near coincident grain boundaries

Embedded atom method (EAM) simulations of the structure of grain boundaries in hexagonal metals are presented. The simulations use recently developed interatomic potentials for Ti and Co. Structures were calculated for various symmetrical tilt boundaries with the [1ˉ100] tilt axis. The structures obtained for both metals are very similar. The energies for the Co boundaries are higher than those for Ti by a factor of 2. The structural unit model was applied to the computed grain-boundary structures in these hexagonal materials. As in cubic materials, the structural unit model can describe a series of symmetrical tilt coincident boundaries. In addition, when the coincidence ratio in the grain-boundary plane varies with thec/a ratio, a structural unit-type model can describe the variation of grain-boundary structure withc/a ratio. This model is adequate for describing series of symmetrical tilt boundaries with the grain-boundary plane oriented perpendicular to a fixed crystallographic direction and varyingc/a ratios. For the structures of the so-called near coincident boundaries that appear in these materials, it was concluded that near coincident boundaries behave similarly to exact coincidence boundaries if there is a coincident periodic structure in the grain-boundary plane. This may occur even without a three-dimensional (3-D) coincident site lattice.     

The correlation between introns and the three-dimensional structure of proteins

We test the hypothesis that introns were used to construct the first genes from small exons, whose protein products represent compact elements of structure. For any three-dimensional structure, a computer program analyzes the structure into a set of modules, segments of the polypeptide chain bounded in space by a maximum diameter, separated by a set of 'boundary regions'. The 'boundary regions' are such that if the gene were divided by an intron in each 'boundary region', the protein would be divided into modules less than the specified diameter. Using a set of 32 ancient proteins, which have no introns in prokaryotes, we examine the intron positions in their eukaryotic homologs and show that the introns are correlated with modules of diameter 21, 28 and 33 A, with P values below 0.001.     

Archaeal introns: splicing, intercellular mobility and evolution

Until recently, it appeared that archaeal introns were spliced by a process specific to the archaeal domain in which an endoribonuclease cuts a 'bulge-helix-bulge' motif that forms at exon-intron junctions. Recent results, however, have shown that the endoribonuclease involved in archaeal intron splicing is a homologue of two subunits of the enzyme complex that excises eukaryotic nuclear tRNA introns. Moreover, some archaeal introns encode homing enzymes that are also encoded by group I introns.     

Asymmetric contribution to Ig repertoire diversity by V kappa exons: differences in the utilization of V kappa 10 exons

The mouse has approximately 140 germline V kappa genes, and functional V kappa exons are expressed at roughly equivalent levels in the preimmune repertoire. We have examined the expression of individual members of the V kappa 10 family. V kappa 10A and V kappa 10B genes have been utilized in numerous hybridomas and myelomas, while V kappa 10C has not. In this study, we have cloned the V kappa 10C gene and shown that it is structurally functional, has the expected promoter elements and recombination signal sequences, and that it is capable of recombination. V kappa 10C mRNA, however, is present at levels at least 1000-fold lower than V kappa 10A and V kappa 10B in adult spleens. While there are no sequence differences in the octamer or TATA box between V kappa 10C and V kappa 10A, there are three nucleotide changes in the promoter region. These promoters equally drive the expression of a reporter gene in B cells or plasma cells, but the V kappa 10A promoter is able to drive expression in pre-B cell lines significantly better than the V kappa 10C promoter (p < 0.05). V kappa 10C rearrangements can be detected in bone marrow and splenic DNA. Therefore, the lack of V kappa 10C expression may reflect the inability of V kappa 10C-rearranged cells to undergo positive or negative selection. Our results suggest that the available Ab repertoire is shaped not only by the number of structurally functional genes, but also by the ability of assembled genes to be expressed at critical points during B cell maturation.     

Paramecium tetraurelia encodes unconventional actin containing short introns

The polymerase chain reaction was used to amplify and clone an actin gene fragment from Paramecium tetraurelia. This DNA fragment was 1,138 bp long, more than 96% of the actin coding sequence, and contained four in-frame UAA codons and two small introns located at positions unique in the actin intron catalogue. This is the first report for the phylum Ciliophora of an actin gene containing introns. The deduced amino acid sequence of this actin fragment shared 58-77% identity with other actins. When compared with rabbit alpha-muscle actin, similarities were observed mainly in subdomains 1 and 3, whereas subdomains 2 and 4 appeared to be more divergent.     

U1 small nuclear RNA and spliceosomal introns in Euglena gracilis

In the flagellated protozoon Euglena gracilis, characterized nuclear genes harbor atypical introns that usually are flanked by short repeats, adopt complex secondary structures in pre-mRNA, and do not obey the GT-AG rule of conventional cis-spliced introns. In the nuclear fibrillarin gene of E. gracilis, we have identified three spliceosomal-type introns that have GT-AG consensus borders. Furthermore, we have isolated a small RNA from E. gracilis and propose, on the basis of primary and secondary structure comparisons, that it is a homolog of U1 small nuclear RNA, an essential component of the cis-spliceosome in higher eukaryotes. Conserved sequences at the 5' splice sites of the fibrillarin introns can potentially base pair with Euglena U1 small nuclear RNA. Our observations demonstrate that spliceosomal GT-AG cis-splicing occurs in Euglena, in addition to the nonconventional cis-splicing and spliced leader trans-splicing previously recognized in this early diverging unicellular eukaryote.     

Toward a resolution of the introns early/late debate: only phase zero introns are correlated with the structure of ancient proteins

We present evidence that a well defined subset of intron positions shows a non-random distribution in ancient genes. We analyze a database of ancient conserved regions drawn from GenBank 101 to retest two predictions of the theory that the first genes were constructed by exon shuffling. These predictions are that there should be an excess of symmetric exons (and sets of exons) flanked by introns of the same phase (positions within the codon) and that intron positions in ancient proteins should correlate with the boundaries of compact protein modules. Both these predictions are supported by the data, with considerable statistical force (P values < 0.0001). Intron positions correlate to modules of diameters around 21, 27, and 33 A, and this correlation is due to phase zero introns. We suggest that 30-40% of present day intron positions in ancient genes correspond to phase zero introns originally present in the progenote, while almost all of the remaining intron positions correspond to introns added, or moved, appearing equally in all three intron phases. This proposal provides a resolution for many of the arguments of the introns-early/introns-late debate.     

U1-mediated exon definition interactions between AT-AC and GT-AG introns

A minor class of metazoan introns has well-conserved splice sites with 5'-AU-AC-3' boundaries, compared to the 5'-GU-AG-3' boundaries and degenerate splice sites of conventional introns. Splicing of the AT-AC intron 2 of a sodium channel (SCN4A) precursor messenger RNA in vitro did not require inhibition of conventional splicing and required adenosine triphosphate, magnesium, and U12 small nuclear RNA (snRNA). When exon 3 was followed by the 5' splice site from the downstream conventional intron, splicing of intron 2 was greatly stimulated. This effect was U1 snRNA-dependent, unlike the basal AT-AC splicing reaction. Therefore, U1-mediated exon definition interactions can coordinate the activities of major and minor spliceosomes.     

Genomic organization and evolution of alternative exons in a Drosophila calcium channel gene

The genomic organization of a gene coding for an alpha 1 subunit of a voltage-gated calcium channel of Drosophila melanogaster (Dmca IA) was determined. Thirty-four exons, distributed over 45 kb of genomic sequence, have been identified and mapped, including exons in three regions involved in alternative splicing and new sites potentially involved in RNA editing. The comparison of the intron/exon boundaries of this channel with a mammalian counterpart shows that the genomic structure of these two genes has remained fairly similar during evolution, with more than half of the Drosophila intron positions being perfectly conserved compared to the human channel. Phylogenetic analysis of the mutually exclusive alternative exons revealed that they have diverged considerably. It is suggested that this divergence, rather than reflecting evolutionary age, is the likely result of accelerated rates of evolution following duplication.     

Expression of CD44 variant exons by primary and metastatic oral squamous carcinomas

Abnormal CD44 expression in many neoplasms correlates with behaviour, but reports on its role in oral squamous carcinoma are contradictory. CD44 expression was characterised in a closely matched series of oral carcinomas with and without metastases in both frozen and formalin-fixed tissue and correlated with behaviour and histological grading parameters. Eleven primary oral squamous carcinomas without metastases and nine primary carcinomas with 19 matched metastases were stained immunocytochemically for CD44H and products of variant exons v3, v4/5, v6 and v9. Patterns of staining in frozen and formalin-fixed tissue were correlated with invasive front grading and behaviour using exact inferential statistics. Most primary carcinomas stained for all exons tested but some showed loss of expression of v4/5. Loss of expression was more marked in metastases, but there was no correlation between expression and behaviour or grade. Stromal surfaces of epithelial cells often expressed variant exon products reflecting loss of polarity. This, together with selective loss of v4 and v5 in primary carcinomas and their more frequent loss in metastases, suggests that CD44 may play a role in metastasis of some oral squamous carcinomas.     

Treasures amongst the trash: functional RNAs within introns

Health care professionals are interested in practical definitions that provide solutions to problems. There are many advantages to defining chemical dependence as a disease. This paper gives a health care professional clear directions regarding identifying the problem of chemical dependence and specifying a specific treatment plan for patients. We review the issues of personal responsibility, the moral model of addictions, and the neuropharmacology of the disease process, and advocate for the disease model as an effective, practical method for treating alcoholism and drug dependence.