Biomedical subjects

C P Ponting

Publications and source records attributed to C P Ponting.

At least 19 recordsLinked to original sources

PubMed2001

A new sequence motif linking lissencephaly, Treacher Collins and oral-facial-digital type 1 syndromes, microtubule dynamics and cell migration.

A previously unidentified sequence motif has been identified in the products of genes mutated in Miller-Dieker lissencephaly, Treacher Collins, oral-facial-digital type 1 and contiguous syndrome ocular albinism with late onset sensorineural deafness syndromes. An additional homologous motif was detected in a gene product fused to the fibroblast growth factor receptor type 1 in patients with an atypical stem cell myeloproliferative disorder. In total, over 100 eukaryotic intracellular proteins are shown to possess a LIS1 homology (LisH) motif, including several katanin p60 subunits, muskelin, tonneau, LEUNIG, Nopp140, aimless and numerous WD repeat-containing beta-propeller proteins. It is suggested that LisH motifs contribute to the regulation of microtubule dynamics, either by mediating dimerization, or else by binding cytoplasmic dynein heavy chain or microtubules directly. The predicted secondary structure of LisH motifs, and their occurrence in homologues of Gbeta beta-propeller subunits, suggests that they are analogues of Ggamma subunits, and might associate with the periphery of beta-propeller domains. The finding of LisH motifs in both treacle and Nopp140 reinforces previous observations of functional similarities between these nucleolar proteins. Uncharacterized LisH motif-containing proteins represent candidates for other diseases associated with aberrant microtubule dynamics and defects of cell migration, nucleokinesis or chromosome segregation.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗

PubMed2001

Mutations in the fukutin-related protein gene (FKRP) cause a form of congenital muscular dystrophy with secondary laminin alpha2 deficiency and abnormal glycosylation of alpha-dystroglycan.

The congenital muscular dystrophies (CMD) are a heterogeneous group of autosomal recessive disorders presenting in infancy with muscle weakness, contractures, and dystrophic changes on skeletal-muscle biopsy. Structural brain defects, with or without mental retardation, are additional features of several CMD syndromes. Approximately 40% of patients with CMD have a primary deficiency (MDC1A) of the laminin alpha2 chain of merosin (laminin-2) due to mutations in the LAMA2 gene. In addition, a secondary deficiency of laminin alpha2 is apparent in some CMD syndromes, including MDC1B, which is mapped to chromosome 1q42, and both muscle-eye-brain disease (MEB) and Fukuyama CMD (FCMD), two forms with severe brain involvement. The FCMD gene encodes a protein of unknown function, fukutin, though sequence analysis predicts it to be a phosphoryl-ligand transferase. Here we identify the gene for a new member of the fukutin protein family (fukutin related protein [FKRP]), mapping to human chromosome 19q13.3. We report the genomic organization of the FKRP gene and its pattern of tissue expression. Mutations in the FKRP gene have been identified in seven families with CMD characterized by disease onset in the first weeks of life and a severe phenotype with inability to walk, muscle hypertrophy, marked elevation of serum creatine kinase, and normal brain structure and function. Affected individuals had a secondary deficiency of laminin alpha2 expression. In addition, they had both a marked decrease in immunostaining of muscle alpha-dystroglycan and a reduction in its molecular weight on western blot analysis. We suggest these abnormalities of alpha-dystroglycan are caused by its defective glycosylation and are integral to the pathology seen in MDC1C.

Adult↗

PubMed2001

Sequence variation and disease in the wake of the draft human genome.

The sequencing phase of the human genome project will soon be over. In its wake, repertoires of sequence polymorphisms among the human population are being sampled and a battery of functional genomics projects, from gene and protein expression studies to whole proteome interaction experiments, are generating vast quantities of data. Now that the data, or the means to generate data, are available it is the application of this information in enhancing our understanding of biology that represents the next formidable challenge. Two prominent issues should be considered. First, existing data must be analysed using the best methods available. The prediction of enzymatic activity for bestrophin, whose gene is mutated in Best macular dystrophy, is described in this review. This is an example of the experimentally testable hypotheses that can result from such detailed and exhaustive analyses. Secondly, the torrents of data from high-throughput studies will need to be made more accessible to all using web-based resources that integrate and digest complementary data types. The internet sites that showcase the human genome sequence are blazing a new trail. Ultimately, the success of genome sequencing and functional genomics will be measured not by the quantity and accuracy of raw data generated, but how rapidly they can be harnessed to span the divide between genotype and phenotype.

Alleles↗

PubMed2001

Domain homologues of dopamine beta-hydroxylase and ferric reductase: roles for iron metabolism in neurodegenerative disorders?

One of the defining characteristics of neurodegenerative diseases, including Parkinson's, Alzheimer's and Huntington's diseases, is abnormal accumulations of iron, specifically in affected areas. Following injection of iron in rat brains, a relatively selective lesion of dopamine neurons, similar to parkinsonism, occurs. These observations indicate that Fe(II)-mediated generation of free radical species, by the Fenton reaction, might contribute to the pathoetiology of these diseases. Iron is known to possess multiple roles in the biosynthesis of catecholamines in dopaminergic neurons. These include, as Fe(II), facilitating the production of dopamine from phenylalanine by tyrosine hydroxylase, and as heme, assisting the recycling of ascorbate by cytochrome b-561 required for the generation of norepinephrine from dopamine by dopamine beta-hydroxylase. In this study, it is demonstrated that a human and mouse gene product, stromal cell-derived receptor 2, is a homologue of cytochrome b-561 and duodenal cytochrome b, and is thus predicted to be active as a ferric reductase. Moreover, this protein also contains a domain homologous to the N-terminal regulatory region of dopamine beta-hydroxylase. These findings from sequence analysis lead to a prediction that stromal cell-derived receptor 2 is a catecholamine-regulated ferric reductase active in the brain. Dysfunction of cytochrome b-561 or stromal cell-derived receptor 2, therefore, might predispose individuals to abnormal accumulation of Fe(III) and/or generation of cytotoxic free radicals as a consequence of a rapid cycling between Fe(III) and Fe(II). The hypothesis that aberrant ferric reductase activities are involved in the progression of neurodegenerative diseases should open up new avenues of research, and possibly therapy, for these devastating diseases.

Amino Acid Sequence↗

PubMed2001

The kyphoscoliosis (ky) mouse is deficient in hypertrophic responses and is caused by a mutation in a novel muscle-specific protein.

The ky mouse mutant exhibits a primary degenerative myopathy preceding chronic thoraco-lumbar kyphoscoliosis. The histopathology of the ky mutant suggests that Ky protein activity is crucial for normal muscle growth and function as well as the maturation and stabilization of the neuromuscular junction. Muscle hypertrophy in response to increasing demand is deficient in the ky mutant, whereas adaptive fibre type shifts take place. The ky locus has previously been localized to a small region of mouse chromosome 9 and we have now identified the gene and the mutation underlying the kyphoscoliotic mouse. The ky transcript encodes a novel protein that is detected only in skeletal muscle and heart. The identification of the ky gene will allow detailed analysis of the impact of primary myopathy on idiopathic scoliosis in mice and man.

Amino Acid Sequence↗

PubMed2001

Plagiarized bacterial genes in the human book of life.

The initial analysis of the human genome draft sequence reveals that our 'book of life' is multi-authored. A small but significant proportion of our genes owes their heritage not to antecedent eukaryotes but instead to bacteria. The publicly funded Human Genome Project study indicates that about 0.5% of all human genes were copied into the genome from bacterial sources. Detailed sequence analyses point to these 'horizontal gene transfer' events having occurred relatively recently. So how did the human 'book of life' evolve to be a chimaera, part animal and part bacterium? And what was the probable evolutionary impact of such gene plagiarism?

Bacteria↗

PubMed2001

JmjC: cupin metalloenzyme-like domains in jumonji, hairless and phospholipase A2beta.

On the basis of significant sequence similarity, we have identified JmjC domains in more than 100 eukaryotic and bacterial sequences. These include human hairless, mutated in individuals with alopecia universalis, retinoblastoma-binding protein 2 and several putative chromatin-associated proteins. JmjC domains are predicted to be metalloenzymes that adopt the cupin fold, and are candidates for enzymes that regulate chromatin remodelling.

Amino Acid Motifs↗

PubMed2001

Issues in predicting protein function from sequence.

Identifying homologues, defined as genes that arose from a common evolutionary ancestor, is often a relatively straightforward task, thanks to recent advances made in estimating the statistical significance of sequence similarities found from database searches. The extent by which homologues possess similarities in function, however, is less amenable to statistical analysis. Consequently, predicting function by homology is a qualitative, rather than quantitative, process and requires particular care to be taken. This review focuses on the various approaches that have been developed to predict function from the scale of the atom to that of the organism. Similarities in homologues' functions differ considerably at each of these different scales and also vary for different domain families. It is argued that due attention should be paid to all available clues to function, including orthologue identification, conservation of particular residue types, and the co-occurrence of domains in proteins. Pitfalls in database searching methods arising from amino acid compositional bias and database size effects are also discussed.

Amino Acids↗

PubMed2001

CHROMA: consensus-based colouring of multiple alignments for publication.

CHROMA annotates multiple protein sequence alignments by consensus to produce formatted and coloured text suitable for incorporation into other documents for publication. The package is designed to be flexible and reliable, and has a simple-to-use graphical user interface running under Microsoft Windows. Both the executables and source code for CHROMA running under Windows and Linux (portable command-line only) are freely available at http://www.lg.ndirect.co.uk/chroma. Software enquiries should be directed to CHROMA@lg.ndirect.co.uk.

Amino Acid Sequence↗

PubMed2001

Novel protein domains and repeats in Drosophila melanogaster: insights into structure, function, and evolution.

Sequence database searching methods such as BLAST, are invaluable for predicting molecular function on the basis of sequence similarities among single regions of proteins. Searches of whole databases however, are not optimized to detect multiple homologous regions within a single polypeptide. Here we have used the prospero algorithm to perform self-comparisons of all predicted Drosophila melanogaster gene products. Predicted repeats, and their homologs from all species, were analyzed further to detect hitherto unappreciated evolutionary relationships. Results included the identification of novel tandem repeats in the human X-linked retinitis pigmentosa type-2 gene product, repeated segments in cystinosin, associated with a defect in cystine transport, and 'nested' homologous domains in dysferlin, whose gene is mutated in limb girdle muscular dystrophy. Novel signaling domain families were found that may regulate the microtubule-based cytoskeleton and ubiquitin-mediated proteolysis, respectively. Two families of glycosyl hydrolases were shown to contain internal repetitions that hint at their evolution via a piecemeal, modular approach. In addition, three examples of fruit fly genes were detected with tandem exons that appear to have arisen via internal duplication. These findings demonstrate how completely sequenced genomes can be exploited to further understand the relationships between molecular structure, function, and evolution.

Amino Acid Sequence↗

PubMed2001

Sialidase-like Asp-boxes: sequence-similar structures within different protein folds.

Sequence similarity is the most common measure currently used to infer homology between proteins. Typically, homologous protein domains show sequence similarity over their entire lengths. Here we identify Asp box motifs, initially found as repeats in sialidases and neuraminidases, in new structural and sequence contexts. These motifs represent significantly similar sequences, localized to beta hairpins within proteins that are otherwise different in sequence and three-dimensional structure. By performing a combined sequence- and structure-based analysis we detect Asp boxes in more than nine protein families, including bacterial ribonucleases, sulfite oxidases, reelin, netrins, some lipoprotein receptors, and a variety of glycosyl hydrolases. Although the function common to each of these proteins, if any, remains unclear, we discuss possible functions of Asp boxes on the basis of previously determined experimental results and discuss different evolutionary scenarios for the origin of Asp-box containing proteins.

Acetylglucosaminidase↗

PubMed2001

Genome cartography through domain annotation.

The evolutionary history of eukaryotic proteins involves rapid sequence divergence, addition and deletion of domains, and fusion and fission of genes. Although the protein repertoires of distantly related species differ greatly, their domain repertoires do not. To account for the great diversity of domain contexts and an unexpected paucity of ortholog conservation, we must categorize the coding regions of completely sequenced genomes into domain families, as well as protein families.

Animals↗

PubMed2000

Syncoilin, a novel member of the intermediate filament superfamily that interacts with alpha-dystrobrevin in skeletal muscle.

Dystrophin coordinates the assembly of a complex of structural and signaling proteins that are required for normal muscle function. A key component of the dystrophin protein complex is alpha-dystrobrevin, a dystrophin-associated protein whose absence results in neuromuscular junction defects and muscular dystrophy. To gain further insights into the role of alpha-dystrobrevin in skeletal muscle, we used the yeast two-hybrid system to identify a novel alpha-dystrobrevin-binding partner called syncoilin. Syncoilin is a new member of the intermediate filament superfamily and is highly expressed in skeletal and cardiac muscle. In normal skeletal muscle, syncoilin is concentrated at the neuromuscular junction, where it colocalizes and coimmunoprecipitates with alpha-dystrobrevin-1. Expression studies in mammalian cells demonstrate that, while alpha-dystrobrevin and syncoilin associate directly, overexpression of syncoilin does not result in the self-assembly of intermediate filaments. Finally, unlike many components of the dystrophin protein complex, we show that syncoilin expression is up-regulated in dystrophin-deficient muscle. These data suggest that alpha-dystrobrevin provides a link between the dystrophin protein complex and the intermediate filament network at the neuromuscular junction, which may be important for the maintenance and maturation of the synapse.

Amino Acid Sequence↗

PubMed2000

Alternative splicing of dystrobrevin regulates the stoichiometry of syntrophin binding to the dystrophin protein complex.

Dystrophin coordinates the assembly of a complex of structural and signalling proteins that is required for normal muscle function. A key component of the dystrophin-associated protein complex (DPC) is alpha-dystrobrevin, a dystrophin-related and -associated protein whose absence results in muscular dystrophy and neuromuscular junction defects [1,2]. The current model of the DPC predicts that dystrophin and dystrobrevin each bind a single syntrophin molecule [3]. The syntrophins are PDZ-domain-containing proteins that facilitate the recruitment of signalling proteins such as nNOS (neuronal nitric oxide synthase) to the DPC [4]. Here we show, using yeast two-hybrid analysis and biochemical binding studies, that alpha-dystrobrevin in fact contains two independent syntrophin-binding sites in tandem. The previously undescribed binding site is situated within an alternatively spliced exon of alpha-dystrobrevin, termed the variable region-3 (vr3) sequence, which is specifically expressed in skeletal and cardiac muscle [5,6]. Analysis of the syntrophin-binding region of dystrobrevin reveals a tandem pair of predicted alpha helices with significant sequence similarity. These alpha helices, each termed a syntrophin-binding motif, are also highly conserved in dystrophin and utrophin. Together these data show that there are four potential syntrophin-binding sites per dystrophin complex in skeletal muscle: two on dystrobrevin and two on dystrophin or utrophin. Furthermore, alternative splicing of dystrobrevin provides a mechanism for regulating the stoichiometry of syntrophin association with the DPC. This is likely to have important consequences for the recruitment of specific signalling molecules to the DPC and ultimately for its function.

Alternative Splicing↗

PubMed2000

Proteins of the endoplasmic-reticulum-associated degradation pathway: domain detection and function prediction.

Sequence database searches, using iterative-profile and Hidden-Markov-model approaches, were used to detect hitherto-undetected homologues of proteins that regulate the endoplasmic reticulum (ER)-associated degradation pathway. The translocon-associated subunit Sec63p (Sec=secretory) was shown to contain a domain of unknown function found twice in several Brr2p-like RNA helicases (Brr2=bad response to refrigeration 2). Additionally, Cue1p (Cue=coupling of ubiquitin conjugation to ER degradation), a yeast protein that recruits the ubiquitin-conjugating (UBC) enzyme Ubc7p to an ER-associated complex, was found to be one of a large family of putative scaffolding-domain-containing proteins that include the autocrine motility factor receptor and fungal Vps9p (Vps=vacuolar protein sorting). Two other yeast translocon-associated molecules, Sec72p and Hrd3p (Hrd=3-hydroxy-3-methylglutaryl-CoA reductase degradation), were shown to contain multiple tetratricopeptide-repeat-like sequences. From this observation it is suggested that Sec72p associates with a heat-shock protein, Hsp70, in a manner analogous to that known for Hop (Hsp70/Hsp90 organizing protein). Finally, the luminal portion of Ire1p (Ire=high inositol-requiring), thought to convey the sensing function of this transmembrane kinase and endoribonuclease, was shown to contain repeats similar to those in beta-propeller proteins. This finding hints at the mechanism by which Ire1p may sense extended unfolded proteins at the expense of compact folded molecules.

Amino Acid Sequence↗

PubMed2000

Identification of distant homologues of fibroblast growth factors suggests a common ancestor for all beta-trefoil proteins.

Determination of the structures of fibroblast growth factors and interleukin-1s has previously revealed that they both adopt a beta-trefoil fold, similar to those found in Kunitz soybean trypsin inhibitors, ricin-like toxins, plant agglutinins and hisactophilin. These families possess distinct functions and occur in different subcellular localisations, and they appear to lack significant similarities in their sequences, ligands and modes of ligand binding. We have analysed the significance of sequence identities observed after structure alignment and provide statistical evidence that these beta-trefoil proteins are all homologues, having arisen from a common ancestor. In addition, we have explored the sequence space of all beta-trefoil proteins and have determined that the actin-binding proteins fascins, and other proteins of unknown function, are beta-trefoil family homologues. Unlike other beta-trefoil proteins, the triplicated repeats in each of the four beta-trefoil domains of fascins are significantly similar in sequence. This hints at how the beta-trefoil fold arose from the duplication of an ancestral gene encoding a homotrimeric single-repeat protein. The combined analysis of structure and sequence databases for detecting significant similarities is suggested as a highly sensitive approach to determining the common ancestry of extremely divergent homologues.

Amino Acid Sequence↗

PubMed2000

Regulation of chromatin structure by site-specific histone H3 methyltransferases.

The organization of chromatin into higher-order structures influences chromosome function and epigenetic gene regulation. Higher-order chromatin has been proposed to be nucleated by the covalent modification of histone tails and the subsequent establishment of chromosomal subdomains by non-histone modifier factors. Here we show that human SUV39H1 and murine Suv39h1--mammalian homologues of Drosophila Su(var)3-9 and of Schizosaccharomyces pombe clr4--encode histone H3-specific methyltransferases that selectively methylate lysine 9 of the amino terminus of histone H3 in vitro. We mapped the catalytic motif to the evolutionarily conserved SET domain, which requires adjacent cysteine-rich regions to confer histone methyltransferase activity. Methylation of lysine 9 interferes with phosphorylation of serine 10, but is also influenced by pre-existing modifications in the amino terminus of H3. In vivo, deregulated SUV39H1 or disrupted Suv39h activity modulate H3 serine 10 phosphorylation in native chromatin and induce aberrant mitotic divisions. Our data reveal a functional interdependence of site-specific H3 tail modifications and suggest a dynamic mechanism for the regulation of higher-order chromatin.

Amino Acid Sequence↗

PubMed2000

Homology-based method for identification of protein repeats using statistical significance estimates.

Short protein repeats, frequently with a length between 20 and 40 residues, represent a significant fraction of known proteins. Many repeats appear to possess high amino acid substitution rates and thus recognition of repeat homologues is highly problematic. Even if the presence of a certain repeat family is known, the exact locations and the number of repetitive units often cannot be determined using current methods. We have devised an iterative algorithm based on optimal and sub-optimal score distributions from profile analysis that estimates the significance of all repeats that are detected in a single sequence. This procedure allows the identification of homologues at alignment scores lower than the highest optimal alignment score for non-homologous sequences. The method has been used to investigate the occurrence of eleven families of repeats in Saccharomyces cerevisiae, Caenorhabditis elegans and Homo sapiens accounting for 1055, 2205 and 2320 repeats, respectively. For these examples, the method is both more sensitive and more selective than conventional homology search procedures. The method allowed the detection in the SwissProt database of more than 2000 previously unrecognised repeats belonging to the 11 families. In addition, the method was used to merge several repeat families that previously were supposed to be distinct, indicating common phylogenetic origins for these families.

Algorithms↗