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Biomedical subjects

Christopher Lee

Publications and source records attributed to Christopher Lee.

At least 19 recordsLinked to original sources
PubMed2006

The HIV positive selection mutation database.

The HIV positive selection mutation database is a large-scale database available at http://www.bioinformatics.ucla.edu/HIV/ that provides detailed selection pressure maps of HIV protease and reverse transcriptase, both of which are molecular targets of antiretroviral therapy. This database makes available for the first time a very large HIV sequence dataset (sequences from approximately 50 000 clinical AIDS samples, generously contributed by Specialty Laboratories, Inc.), which makes possible high-resolution selection pressure mapping. It provides information about not only the selection pressure on individual sites but also how selection pressure at one site is affected by mutations on other sites. It also includes datasets from other public databases, namely the Stanford HIV database [S. Y. Rhee, M. J. Gonzales, R. Kantor, B. J. Betts, J. Ravela and R. W. Shafer (2003) Nucleic Acids Res., 31, 298-303]. Comparison between these datasets in the database enables cross-validation with independent datasets and also specific evaluation of the effect of drug treatment.

Acquired Immunodeficiency Syndrome↗
PubMed2006

The ASAP II database: analysis and comparative genomics of alternative splicing in 15 animal species.

We have greatly expanded the Alternative Splicing Annotation Project (ASAP) database: (i) its human alternative splicing data are expanded approximately 3-fold over the previous ASAP database, to nearly 90,000 distinct alternative splicing events; (ii) it now provides genome-wide alternative splicing analyses for 15 vertebrate, insect and other animal species; (iii) it provides comprehensive comparative genomics information for comparing alternative splicing and splice site conservation across 17 aligned genomes, based on UCSC multigenome alignments; (iv) it provides an approximately 2- to 3-fold expansion in detection of tissue-specific alternative splicing events, and of cancer versus normal specific alternative splicing events. We have also constructed a novel database linking orthologous exons and orthologous introns between genomes, based on multigenome alignment of 17 animal species. It can be a valuable resource for studies of gene structure evolution. ASAP II provides a new web interface enabling more detailed exploration of the data, and integrating comparative genomics information with alternative splicing data. We provide a set of tools for advanced data-mining of ASAP II with Pygr (the Python Graph Database Framework for Bioinformatics) including powerful features such as graph query, multigenome alignment query, etc. ASAP II is available at http://www.bioinformatics.ucla.edu/ASAP2.

Alternative Splicing↗
PubMed2006

Alternative splicing and RNA selection pressure--evolutionary consequences for eukaryotic genomes.

Genome-wide analyses of alternative splicing have established its nearly ubiquitous role in gene regulation in many organisms. Genome sequencing and comparative genomics have made it possible to look in detail at the evolutionary history of specific alternative exons or splice sites, resulting in a flurry of publications in recent years. Here, we consider how alternative splicing has contributed to the evolution of modern genomes, and discuss constraints on evolution associated with alternative splicing that might have important medical implications.

Alternative Splicing↗
PubMed2006

An expectation-maximization algorithm for probabilistic reconstructions of full-length isoforms from splice graphs.

Reconstructing full-length transcript isoforms from sequence fragments (such as ESTs) is a major interest and challenge for bioinformatic analysis of pre-mRNA alternative splicing. This problem has been formulated as finding traversals across the splice graph, which is a directed acyclic graph (DAG) representation of gene structure and alternative splicing. In this manuscript we introduce a probabilistic formulation of the isoform reconstruction problem, and provide an expectation-maximization (EM) algorithm for its maximum likelihood solution. Using a series of simulated data and expressed sequences from real human genes, we demonstrate that our EM algorithm can correctly handle various situations of fragmentation and coupling in the input data. Our work establishes a general probabilistic framework for splice graph-based reconstructions of full-length isoforms.

Algorithms↗
PubMed2006

Distinguishing HIV-1 drug resistance, accessory, and viral fitness mutations using conditional selection pressure analysis of treated versus untreated patient samples.

BACKGROUND: HIV can evolve drug resistance rapidly in response to new drug treatments, often through a combination of multiple mutations 123. It would be useful to develop automated analyses of HIV sequence polymorphism that are able to predict drug resistance mutations, and to distinguish different types of functional roles among such mutations, for example, those that directly cause drug resistance, versus those that play an accessory role. Detecting functional interactions between mutations is essential for this classification. We have adapted a well-known measure of evolutionary selection pressure (Ka/Ks) and developed a conditional Ka/Ks approach to detect important interactions. RESULTS: We have applied this analysis to four independent HIV protease sequencing datasets: 50,000 clinical samples sequenced by Specialty Laboratories, Inc.; 1800 samples from patients treated with protease inhibitors; 2600 samples from untreated patients; 400 samples from untreated African patients. We have identified 428 mutation interactions in Specialty dataset with statistical significance and we were able to distinguish primary vs. accessory mutations for many well-studied examples. Amino acid interactions identified by conditional Ka/Ks matched 80 of 92 pair wise interactions found by a completely independent study of HIV protease (p-value for this match is significant: 10-70). Furthermore, Ka/Ks selection pressure results were highly reproducible among these independent datasets, both qualitatively and quantitatively, suggesting that they are detecting real drug-resistance and viral fitness mutations in the wild HIV-1 population. CONCLUSION: Conditional Ka/Ks analysis can detect mutation interactions and distinguish primary vs. accessory mutations in HIV-1. Ka/Ks analysis of treated vs. untreated patient data can distinguish drug-resistance vs. viral fitness mutations. Verification of these results would require longitudinal studies. The result provides a valuable resource for AIDS research and will be available for open access upon publication at http://www.bioinformatics.ucla.edu/HIV.

Journal Article↗
PubMed2006

Evolutionary divergence of exon flanks: a dissection of mutability and selection.

The intronic sequences flanking exon-intron junctions (i.e., exon flanks) are important for splice site recognition and pre-mRNA splicing. Recent studies show a higher degree of sequence conservation at flanks of alternative exons, compared to flanks of constitutive exons. In this article we performed a detailed analysis on the evolutionary divergence of exon flanks between human and chimpanzee, aiming to dissect the impact of mutability and selection on their evolution. Inside exon flanks, sites that might reside in ancestral CpG dinucleotides evolved significantly faster than sites outside of ancestral CpG dinucleotides. This result reflects a systematic variation of mutation rates (mutability) at exon flanks, depending on the local CpG contexts. Remarkably, we observed a significant reduction of the nucleotide substitution rate in flanks of alternatively spliced exons, independent of the site-by-site variation in mutability due to different CpG contexts. Our data provide concrete evidence for increased purifying selection at exon flanks associated with regulation of alternative splicing.

Alternative Splicing↗
PubMed2006

Can RNA selection pressure distort the measurement of Ka/Ks?

Recently, an interesting question has emerged in the evolutionary interpretation of sequence substitution data as evidence of amino acid selection pressure. Specifically, the Ka/Ks metric was designed to measure selection pressure on amino acid substitutions, assuming that the synonymous substitution rate Ks reflects the neutral nucleotide substitution rate. However, there is increasing evidence for selection pressure at silent sites due to constraints of RNA splicing. Is Ka/Ks an appropriate metric for selection pressure on amino acid substitutions, in the presence of other selection pressures acting only at the RNA level (such as selection for exonic splicing enhancers)? Or can the resulting decreases in Ks from such selection pressures introduce bias into the Ka/Ks metric, so that it no longer gives an accurate measure of amino acid level selection pressure? In this review, we present both mathematical models and empirical evidence for these divergent points of view.

Amino Acid Substitution↗
PubMed2005

Yarn design for functional tissue engineering.

Tissue engineering requires the ability to design scaffolds with mechanical properties similar to those of the native tissue. Here, B. mori silk yarns are used as a model system to demonstrate the potential benefits and drawbacks of several textile methods used to fabricate tissue engineering scaffolds. Fibers are plied, twisted, cabled, braided, and/or textured to form several geometries with a wide range of mechanical outcomes. Predictable changes in ultimate tensile strength and stiffness are demonstrated following processing and as a function of test environment. The mechanical effects of increasing turns per inch and combining groups of fibers into higher-order yarn structures are demonstrated. Braids, one of the most commonly used textile structures, are shown to be limited by a change in stiffness following the locking-angle and therefore, potentially not the ideal structure for tissue engineering. Cabled yarns appear to allow the most flexibility in mechanical outcomes with a highly organized geometry. Twisted yarns, while more economical than cabled yarns, result in a higher stiffness and lower percent elongation at break than cabled yarns.

Animals↗
PubMed2005

Inferring protein domain interactions from databases of interacting proteins.

We describe domain pair exclusion analysis (DPEA), a method for inferring domain interactions from databases of interacting proteins. DPEA features a log odds score, Eij, reflecting confidence that domains i and j interact. We analyzed 177,233 potential domain interactions underlying 26,032 protein interactions. In total, 3,005 high-confidence domain interactions were inferred, and were evaluated using known domain interactions in the Protein Data Bank. DPEA may prove useful in guiding experiment-based discovery of previously unrecognized domain interactions.

Animals↗
PubMed2005

Evidence of functional selection pressure for alternative splicing events that accelerate evolution of protein subsequences.

Recently, it was proposed that alternative splicing may act as a mechanism for opening accelerated paths of evolution, by reducing negative selection pressure, but there has been little evidence so far that this mechanism could produce adaptive benefit. Here, we use metrics of very different types of selection pressures [e.g., against amino acid mutations (Ka/Ks), against mutations at synonymous sites (Ks), and for protein reading-frame preservation] to address this question by genomewide analyses of human, chimpanzee, mouse, and rat. These data show that alternative splicing relaxes Ka/Ks selection pressure up to 7-fold, but intriguingly this effect is accompanied by a strong increase in selection pressure against synonymous mutations, which propagates into the adjacent intron, and correlates strongly with the alternative splicing level observed for each exon. These effects are highly local to the alternatively spliced exon. Comparisons of these four genomes consistently show an increase in the density of amino acid mutations (Ka) in alternatively spliced exons and a decrease in the density of synonymous mutations (Ks). This selection pressure against synonymous mutations in alternatively spliced exons was accompanied in all four genomes by a striking increase in selection pressure for protein reading-frame preservation, and both increased markedly with increasing evolutionary age. Restricting our analysis to a subset of exons with strong evidence for biologically functional alternative splicing produced identical results. Thus alternative splicing apparently can create evolutionary "hotspots" within a protein sequence, and these events have evidently been selected for during mammalian evolution.

Alternative Splicing↗
PubMed2005

Evidence that public database records for many cancer-associated genes reflect a splice form found in tumors and lack normal splice forms.

Alternative splicing is widespread in the human genome, and it appears that many genes display different splice forms in cancerous tissue than in normal human tissues. However, since cDNAs for many cancer-associated genes were originally cloned from tumor samples, it is important to ask whether this repertoire of cDNAs provides a complete or representative picture of the transcript isoforms found in normal tissues. To answer this, we used bioinformatics and RT-PCR to identify novel splice forms, focusing on in-frame exonskips, for a panel of 50 cancer-associated genes in normal tissue samples. These data show that in nearly two-thirds of the genes, normal tissues expressed previously unknown splice forms, of which 40% were normally a dominant splice form. Surprisingly, the tumor-associated splice forms were twice as likely to be represented in GenBank than their normal tissue-associated splice forms, most likely because 70% of the mRNAs in GenBank for these genes were cloned from tumor samples. As an example, we describe a novel normal splice form of IKBbeta, an important regulator of the NFkappaB pathway. Our data suggest that systematic re-evaluation of cancer genes' splice forms in normal tissue will yield insights into their distinct functions in normal tissues and in cancer. Our database contains 1308 novel normal splice forms, including many known cancer genes.

Alternative Splicing↗
PubMed2005

Assessing the application of Ka/Ks ratio test to alternatively spliced exons.

SUMMARY: Recently, the Ka/Ks ratio test, which assesses the protein-coding potentials of genomic regions based on their non-synonymous to synonymous divergence rates, has been proposed and successfully used in genome annotations of eukaryotes. We systematically performed the Ka/Ks ratio test on 925 transcript-confirmed alternatively spliced exons in the human genome, which we describe in this manuscript. We found that 22.3% of evolutionarily conserved alternatively spliced exons cannot pass the Ka/Ks ratio test, compared with 9.8% for constitutive exons. The false negative rate was the highest (85.7%) for exons with low frequencies of transcript inclusion. Analyses of alternatively spliced exons supported by full-length mRNA sequences yielded similar results, and nearly half of exons involved in ancestral alternative splicing events could not pass this test. Our analysis suggests a future direction to incorporate comparative genomics-based alternative splicing predictions with the Ka/Ks ratio test in higher eukaryotes with extensive RNA alternative splicing.

Algorithms↗
PubMed2005

MPP+-induced COX-2 activation and subsequent dopaminergic neurodegeneration.

The importance of cyclooxygenase-2 (COX-2) in mediating Parkinson's disease (PD) was suggested in reports, indicating that COX-2 selective inhibitors or genetic knockout reduce 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopaminergic (DA) neurotoxicity in a mouse model of PD. However, cell types and mechanisms underlying the activation of COX-2 have not been clearly elucidated in these animal studies. Using primary neuron-glia cultures, we aimed to determine 1) whether microglia participate in 1-methyl-4-phenylpryridinium (MPP)-induced COX-2 activation and 2) whether the activation of COX-2 contributes to subsequent neurotoxicity. MPP, in a concentration-dependent manner, increased prostaglandin E2 (PGE2) production in mixed neuron-microglia cultures but not in enriched neuron, microglia, or astroglia cultures nor in mixed neuron-astroglia cultures. MPP-induced PGE2 increase was completely abolished by treatment with DuP697, a COX-2 selective inhibitor. DuP697 also significantly reduced MPP-induced DA neurotoxicity as determined by DA uptake assay. Immunocytochemistry and confocal microscopy studies showed enhanced COX-2 expression in both microglia and neurons after MPP treatment. However, neuronal increase in COX-2 expression was not totally dependent on the production of PGE2 from microglia, since microglia deficient in COX-2 only attenuated, but did not completely block, MPP-increased PGE2 production in mixed neuron-microglia cultures, suggesting that part of PGE2 production was originated from neurons. Together, these results indicate that MPP-induced COX-2 expression and subsequent PGE2 production depend on interactions between neurons and microglia. Microgliosis may also be responsible for the COX-2 activation in neurons, leading to the enhanced DA neurotoxicity, which, in turn, reinforces microgliosis. Thus inhibition of microgliosis and COX-2 activity may stop this vicious circle and be valuable strategies in PD therapy.

1-Methyl-4-phenylpyridinium↗
PubMed2005

Targeting alternatively spliced sequence features for cancer diagnosis and therapeutics.

Alternative splicing is emerging as a major new mechanism of functional regulation in mammals, and there is increasing evidence that human cancers often involve significant changes in alternative splicing. In some cases, these changes contribute functionally to the maintenance of the transformed state and could be useful as novel targets for anticancer therapy. In other cases, they reflect changes due to tumorigenesis and could be useful for diagnostic purposes. Fundamentally, alternative splicing offers a novel opportunity to target individual subregions of a gene product that are preferentially expressed in tumors and which are not found in isoforms of the same gene found preferentially in normal tissues.

Alternative Splicing↗
PubMed2005

Bioinformatics analysis of alternative splicing.

Over the past few years, the analysis of alternative splicing using bioinformatics has emerged as an important new field, and has significantly changed our view of genome function. One exciting front has been the analysis of microarray data to measure alternative splicing genome-wide. Pioneering studies of both human and mouse data have produced algorithms for discerning evidence of alternative splicing and clustering genes and samples by their alternative splicing patterns. Moreover, these data indicate the presence of alternative splice forms in up to 80 per cent of human genes. Comparative genomics studies in both mammals and insects have demonstrated that alternative splicing can in some cases be predicted directly from comparisons of genome sequences, based on heightened sequence conservation and exon length. Such studies have also provided new insights into the connection between alternative splicing and a variety of evolutionary processes such as Alu-based exonisation, exon creation and loss. A number of groups have used a combination of bioinformatics, comparative genomics and experimental validation to identify new motifs for splice regulatory factors, analyse the balance of factors that regulate alternative splicing, and propose a new mechanism for regulation based on the interaction of alternative splicing and nonsense-mediated decay. Bioinformatics studies of the functional impact of alternative splicing have revealed a wide range of regulatory mechanisms, from NAGNAG sites that add a single amino acid; to short peptide segments that can play surprisingly complex roles in switching protein conformation and function (as in the Piccolo C2A domain); to events that entirely remove a specific protein interaction domain or membrane anchoring domain. Common to many bioinformatics studies is a new emphasis on graph representations of alternative splicing structures, which have many advantages for analysis.

Algorithms↗
PubMed2005

Cortical adaptation following partial glossectomy with primary closure: implications for reconstruction of the oral tongue.

OBJECTIVES: The considerable variability in functional outcomes for speech and swallowing with different reconstruction techniques following partial glossectomy may reflect the ability of patients to adapt to altered tongue structure. The purpose of this study was to determine mechanisms of cortical adaptation in swallowing to partial glossectomy reconstructed with primary closure. METHODS: Four patients treated with partial glossectomy and primary closure underwent functional magnetic resonance imaging at a mean of 6 months after operation, and the data were compared to those from 8 healthy controls. RESULTS: Statistically significant increases in activity occurred predominately in the parietal cortices and the cerebellum. The volume of the resection was most highly correlated with activity in the premotor and parietal cortices and cerebellum. CONCLUSIONS: The adaptive changes in the cortex following partial glossectomy with primary closure reflect adaptation to the biomechanics of tongue movement during swallowing, and not altered sensation in the tongue.

Adaptation, Physiological↗