The haplo-spliceo-transcriptome: common variations in alternative splicing in the human population

Numerous inherited human genetic disorders are caused by defects in pre-mRNA splicing. Two recent studies have added a new twist to the link between genetic variation and pre-mRNA splicing by identifying SNPs that correlate with heritable changes in alternative splicing but do not cause disease. This suggests that allele-specific alternative splicing is a mechanism that accounts for individual variation in the human population.     

Fingerprint profile of Ginkgo biloba nutritional supplements by LC/ESI-MS/MS

Ginkgo biloba is one of the most popular herb nutrition supplements, with terpene lactones and flavonoids being the two major active components. A fingerprint profile method was developed using a capillary HPLC/MS method which can identify more than 70 components from the G. biloba product. The method allows the flavonoids and terpene lactones to be detected simultaneously and information of both the parent ion and its fragmentation can be obtained in just one HPLC/MS run. Targeted post-acquisition analysis allows mass spectrometric information regarding the identification of flavonoid components to be easily distinguished from other data, however the same approach for terpene lactones was less successful due to dimer formation and requires further development. The fingerprint profiles of five commercial G. biloba nutritional supplements were obtained and compared; variation of some components among the samples was observed and fortification could be detected. In the quality control analysis of the G. biloba product this method could be viewed as complementary to specific quantitative analysis of some bioactive components of the herb.     

Stability analysis of higher-order neural networks for combinatorial optimization

Recurrent neural networks with higher order connections, from here on referred to as higher-order neural networks (HONNs), may be used for the solution of combinatorial optimization problems. In Ref. 5 a mapping of the traveling salesman problem (TSP) onto a HONN of arbitrary order was developed, thereby creating a family of related networks that can be used to solve the TSP. In this paper, we explore the trade-off between network complexity and quality of solution that is made available by the HONN mapping of the TSP. The trade-off is investigated by undertaking an analysis of the stability of valid solutions to the TSP in a HONN of arbitrary order. The techniques used to perform the stability analysis are not new, but have been widely used elsewhere in the literature. The original contribution in this paper is the application of these techniques to a HONN of arbitrary order used to solve the TSP. The results of the stability analysis show that the quality of solution is improved by increasing the network complexity, as measured by the order of the network. Furthermore, it is shown that the Hopfield network, as the simplest network in the family of higher-order networks, is expected to produce the poorest quality of solution.     

Exon-specific RNAi: a tool for dissecting the functional relevance of alternative splicing

The goal of functional genomics is to determine the function of each protein encoded by an organism. Typically, this is done by inactivating individual genes and, subsequently, analyzing the phenotype of the modified organisms. In higher eukaryotes, where a tremendous amount of alternative splicing occurs, such approaches are not feasible because they have the potential to simultaneously affect multiple proteins that could have quite distinct and important functions. Thus, it is necessary to develop techniques that inactivate only a subset of proteins synthesized from genes encoding alternatively spliced mRNAs. Here we demonstrate that RNA interference (RNAi) can be used to selectively degrade specific alternatively spliced mRNA isoforms in cultured Drosophila cells. This is achieved by treating the cells with double-stranded RNA corresponding to an alternatively spliced exon. This technique may prove to be a powerful tool to assess the function of proteins synthesized from alternatively spliced mRNAs. In addition, these results have implications regarding the mechanism of RNAi in Drosophila.     

Analysis of the human neurexin genes: alternative splicing and the generation of protein diversity

The neurexins are neuronal proteins that function as cell adhesion molecules during synaptogenesis and in intercellular signaling. Although mammalian genomes contain only three neurexin genes, thousands of neurexin isoforms may be expressed through the use of two alternative promoters and alternative splicing at up to five different positions in the pre-mRNA. To begin understanding how the expression of the neurexin genes is regulated, we have determined the complete nucleotide sequence of all three human neurexin genes: NRXN1, NRXN2, and NRXN3. Unexpectedly, two of these, NRXN1 ( approximately 1.1 Mb) and NRXN3 ( approximately 1.7 Mb), are among the largest known human genes. In addition, we have identified several conserved intronic sequence elements that may participate in the regulation of alternative splicing. The sequences of these genes provide insight into the mechanisms used to generate the diversity of neurexin protein isoforms and raise several interesting questions regarding the expression mechanism of large genes.     

CF45-1, a secreted protein which participates in Dictyostelium group size regulation

Developing Dictyostelium cells aggregate to form fruiting bodies containing typically 2 × 10⁴ cells. To prevent the formation of an excessively large fruiting body, streams of aggregating cells break up into groups if there are too many cells. The breakup is regulated by a secreted complex of polypeptides called counting factor (CF). Countin and CF50 are two of the components of CF. Disrupting the expression of either of these proteins results in cells secreting very little detectable CF activity, and as a result, aggregation streams remain intact and form large fruiting bodies, which invariably collapse. We find that disrupting the gene encoding a third protein present in crude CF, CF45-1, also results in the formation of large groups when cells are grown with bacteria on agar plates and then starve. However, unlike countin⁻ and cf50⁻ cells, cf45-1⁻ cells sometimes form smaller groups than wild-type cells when the cells are starved on filter pads. The predicted amino acid sequence of CF45-1 has some similarity to that of lysozyme, but recombinant CF45-1 has no detectable lysozyme activity. In the exudates from starved cells, CF45-1 is present in a ~450-kDa fraction that also contains countin and CF50, suggesting that it is part of a complex. Recombinant CF45-1 decreases group size in colonies of cf45-1⁻ cells with a 50% effective concentration (EC₅₀) of ~8 ng/ml and in colonies of wild-type and cf50⁻ cells with an EC₅₀ of ~40 ng/ml. Like countin⁻ and cf50⁻ cells, cf45-1⁻ cells have high levels of cytosolic glucose, high cell-cell adhesion, and low cell motility. Together, the data suggest that CF45-1 participates in group size regulation in Dictyostelium.