Mutations in ATP1A3 cause Alternating Hemiplegia of Childhood (AHC) by disrupting function of the... more Mutations in ATP1A3 cause Alternating Hemiplegia of Childhood (AHC) by disrupting function of the neuronal Na+/K+ ATPase. Published studies to date indicate 2 recurrent mutations, D801N and E815K, and a more severe phenotype in the E815K cohort. We performed mutation analysis and retrospective genotype-phenotype correlations in all eligible patients with AHC enrolled in the US AHC Foundation registry from 1997-2012. Clinical data were abstracted from standardized caregivers' questionnaires and medical records and confirmed by expert clinicians. We identified ATP1A3 mutations by Sanger and whole genome sequencing, and compared phenotypes within and between 4 groups of subjects, those with D801N, E815K, other ATP1A3 or no ATP1A3 mutations. We identified heterozygous ATP1A3 mutations in 154 of 187 (82%) AHC patients. Of 34 unique mutations, 31 (91%) are missense, and 16 (47%) had not been previously reported. Concordant with prior studies, more than 2/3 of all mutations are cluster...
The complex of central problems in data analysis consists of three components: (1) detecting the ... more The complex of central problems in data analysis consists of three components: (1) detecting the dependence of variables using quantitative measures, (2) defining the significance of these dependence measures, and (3) inferring the functional relationships among dependent variables. We have argued previously that an information theory approach allows separation of the detection problem from the inference of functional form problem. We approach here the third component of inferring functional forms based on information encoded in the functions. We present here a direct method for classifying the functional forms of discrete functions of three variables represented in data sets. Discrete variables are frequently encountered in data analysis, both as the result of inherently categorical variables and from the binning of continuous numerical variables into discrete alphabets of values. The fundamental question of how much information is contained in a given function is answered for these discrete functions , and their surprisingly complex relationships are illustrated. The all-important effect of noise on the inference of function classes is found to be highly heterogeneous and reveals some unexpected patterns. We apply this classification approach to an important area of biological data analysis-that of inference of genetic interactions. Genetic analysis provides a rich source of real and complex biological data analysis problems, and our general methods provide an analytical basis and tools for characterizing genetic problems and for analyzing genetic data. We illustrate the functional description and the classes of a number of common genetic interaction modes and also show how different modes vary widely in their sensitivity to noise.
Genomic technologies are best defined as technologies used to manipulate and analyze genomic info... more Genomic technologies are best defined as technologies used to manipulate and analyze genomic information. The evolution of this collective power began in earnest with the invention of DNA cloning in the 1970's and most of the technology derives from the last quarter of the 20th century. The historical impact of these technologies is clearly immense. With the genome sequence becoming available for many organisms, including humans, another new view of biology has recently emerged. This review examines the shape and texture of this recent evolution, with a particular emphasis on new technology: DNA cloning, macromolecular structure analysis (X-ray crystallography and NMR), DNA sequencing, DNA synthesis, amplification by the polymerase chain reaction, and transgenic animals (bacteria through mammals) ORDER FROM www.caister.com UK/Europe: Caister
CITATIONS 27 READS 12 4 authors, including: Some of the authors of this publication are also work... more CITATIONS 27 READS 12 4 authors, including: Some of the authors of this publication are also working on these related projects: Statistical modeling and simulation of short RNA sequencing experiments View project P2X4 receptors and dopamine neurotransmission View project ABSTRACT IS1 is one of the smallest transposable elements found in bacteria (768 bp).
The insertion sequence IS] displays a complex array of open reading frames (ORF). In an attempt t... more The insertion sequence IS] displays a complex array of open reading frames (ORF). In an attempt to identify those which encode polypeptide products, we have systematically placed each ORF under the control of the P1 promoter of phage lambda. In the expression system we used, only the product of the insA gene was present in high enough amounts to be detected by polyacrylamide gel electrophoresis. The production of InsA was further increased in a firt codon hook-up to phage 17 trAnscriptional and translational initiation sinals. Cell extracts from IsA overproducers display a DNA binding activity specific for the ends of IS1. This activity was identified as the IsA protein itself.
The experiments reported here provide strong evidence indicating that the transposition frequency... more The experiments reported here provide strong evidence indicating that the transposition frequency of the bacterial insertion sequence IS] is determined principally by two ISJ-specified proteins. The first, InsA, was previously shown to bind to the ends of the element and to act as a repressor. We present both physical and genetic evidence which reveals that the second, the InsAB' transposase, is a fusion of InsA with the product of a downstream reading frame, InsB'. Synthesis of this protein occurs by a -1 frameshift between the insA and insB' frames. It requires the presence of an intact retroviral-like frameshift signal composed of an A6C motif and a downstream region able to form several alternative secondary structures. In vivo studies show that IS1 transposition activity depends on the relative rather than on the absolute levels of InsA and InsAB'. The ratio is determined primarily at the translational level by frameshifting and appears to be relatively insensitive to large variations in levels of transcription. This novel homeostatic control could therefore protect IS] from activation as a consequence of insertion into active transcription units.
Extracellular RNAs (exRNAs) have been identified in all tested biofluids and have been associated... more Extracellular RNAs (exRNAs) have been identified in all tested biofluids and have been associated with a variety of extracellular vesicles, ribonucleoprotein complexes and lipoprotein complexes. Much of the interest in exRNAs lies in the fact that they may serve as signalling molecules between cells, their potential to serve as biomarkers for prediction and diagnosis of disease and the possibility that exRNAs or the extracellular particles that carry them might be used for therapeutic purposes. Among the most significant bottlenecks to progress in this field is the lack of robust and standardized methods for collection and processing of biofluids, separation of different types of exRNA-containing particles and isolation and analysis of exRNAs. The Sample and Assay Standards Working Group of the Extracellular RNA Communication Consortium is a group of laboratories funded by the U.S. National Institutes of Health to develop such methods. In our first joint endeavour, we held a series ...
The integration host factor of Escherichia coli (IHF) is a small, histone-like protein which part... more The integration host factor of Escherichia coli (IHF) is a small, histone-like protein which participates in the integration of bacteriophage lambda into the E. coli chromosome and in a number of regulatory processes. Our recent footprinting analysis has shown that IHF binds specifically to the ends of the transposable element IS1, as well as to several sites within a short segment of the plasmid pBR322. We have extended our studies of the binding of the IHF molecule to these sites in vitro using a gel retardation assay. We report here that IHF bends the DNA upon binding, as judged from the strong cyclic dependence of the protein-induced mobility shift on the position of the binding site. Using cloned, synthetic ends of IS1 as substrates, we have found that some mutations within the conserved bases of the IHF consensus binding sequence abolish binding, and that alterations of the flanking sequences can greatly reduce IHF binding. The presence of multiple IHF sites on a single DNA fr...
Genomic technologies are best defined as technologies used to manipulate and analyze genomic info... more Genomic technologies are best defined as technologies used to manipulate and analyze genomic information. The evolution of this collective power began in earnest with the invention of DNA cloning in the 1970's and most of the technology derives from the last quarter of the 20th century. The historical impact of these technologies is clearly immense. With the genome sequence becoming available for many organisms, including humans, another new view of biology has recently emerged. This review examines the shape and texture of this recent evolution, with a particular emphasis on new technology: DNA cloning, macromolecular structure analysis (X-ray crystallography and NMR), DNA sequencing, DNA synthesis, amplification by the polymerase chain reaction, and transgenic animals (bacteria through mammals).
A S S phisms (SNPs) [a number comparable to what is already publicly available (21)] Venter et al... more A S S phisms (SNPs) [a number comparable to what is already publicly available (21)] Venter et al. show that these new opportunities-to paraphrase another milestone a dcle-"have not escaped their notice" (22). . 9. The observed 40,000-fold variation in eukaryote haploid DNA content ("Cvalue") is unrelated to organismic complexity or to the numbers of protein-coding genes; see T.
We present evidence that the Escherichia coli DNA binding protein, IHF, plays an important role i... more We present evidence that the Escherichia coli DNA binding protein, IHF, plays an important role in conjugal transfer of the plasmid F. Our results suggest that IHF exerts this effect by positively effecting transcription of the transfer (tra) operon of the plasmid.
Mutations in ATP1A3 cause Alternating Hemiplegia of Childhood (AHC) by disrupting function of the... more Mutations in ATP1A3 cause Alternating Hemiplegia of Childhood (AHC) by disrupting function of the neuronal Na+/K+ ATPase. Published studies to date indicate 2 recurrent mutations, D801N and E815K, and a more severe phenotype in the E815K cohort. We performed mutation analysis and retrospective genotype-phenotype correlations in all eligible patients with AHC enrolled in the US AHC Foundation registry from 1997-2012. Clinical data were abstracted from standardized caregivers' questionnaires and medical records and confirmed by expert clinicians. We identified ATP1A3 mutations by Sanger and whole genome sequencing, and compared phenotypes within and between 4 groups of subjects, those with D801N, E815K, other ATP1A3 or no ATP1A3 mutations. We identified heterozygous ATP1A3 mutations in 154 of 187 (82%) AHC patients. Of 34 unique mutations, 31 (91%) are missense, and 16 (47%) had not been previously reported. Concordant with prior studies, more than 2/3 of all mutations are cluster...
The complex of central problems in data analysis consists of three components: (1) detecting the ... more The complex of central problems in data analysis consists of three components: (1) detecting the dependence of variables using quantitative measures, (2) defining the significance of these dependence measures, and (3) inferring the functional relationships among dependent variables. We have argued previously that an information theory approach allows separation of the detection problem from the inference of functional form problem. We approach here the third component of inferring functional forms based on information encoded in the functions. We present here a direct method for classifying the functional forms of discrete functions of three variables represented in data sets. Discrete variables are frequently encountered in data analysis, both as the result of inherently categorical variables and from the binning of continuous numerical variables into discrete alphabets of values. The fundamental question of how much information is contained in a given function is answered for these discrete functions , and their surprisingly complex relationships are illustrated. The all-important effect of noise on the inference of function classes is found to be highly heterogeneous and reveals some unexpected patterns. We apply this classification approach to an important area of biological data analysis-that of inference of genetic interactions. Genetic analysis provides a rich source of real and complex biological data analysis problems, and our general methods provide an analytical basis and tools for characterizing genetic problems and for analyzing genetic data. We illustrate the functional description and the classes of a number of common genetic interaction modes and also show how different modes vary widely in their sensitivity to noise.
Genomic technologies are best defined as technologies used to manipulate and analyze genomic info... more Genomic technologies are best defined as technologies used to manipulate and analyze genomic information. The evolution of this collective power began in earnest with the invention of DNA cloning in the 1970's and most of the technology derives from the last quarter of the 20th century. The historical impact of these technologies is clearly immense. With the genome sequence becoming available for many organisms, including humans, another new view of biology has recently emerged. This review examines the shape and texture of this recent evolution, with a particular emphasis on new technology: DNA cloning, macromolecular structure analysis (X-ray crystallography and NMR), DNA sequencing, DNA synthesis, amplification by the polymerase chain reaction, and transgenic animals (bacteria through mammals) ORDER FROM www.caister.com UK/Europe: Caister
CITATIONS 27 READS 12 4 authors, including: Some of the authors of this publication are also work... more CITATIONS 27 READS 12 4 authors, including: Some of the authors of this publication are also working on these related projects: Statistical modeling and simulation of short RNA sequencing experiments View project P2X4 receptors and dopamine neurotransmission View project ABSTRACT IS1 is one of the smallest transposable elements found in bacteria (768 bp).
The insertion sequence IS] displays a complex array of open reading frames (ORF). In an attempt t... more The insertion sequence IS] displays a complex array of open reading frames (ORF). In an attempt to identify those which encode polypeptide products, we have systematically placed each ORF under the control of the P1 promoter of phage lambda. In the expression system we used, only the product of the insA gene was present in high enough amounts to be detected by polyacrylamide gel electrophoresis. The production of InsA was further increased in a firt codon hook-up to phage 17 trAnscriptional and translational initiation sinals. Cell extracts from IsA overproducers display a DNA binding activity specific for the ends of IS1. This activity was identified as the IsA protein itself.
The experiments reported here provide strong evidence indicating that the transposition frequency... more The experiments reported here provide strong evidence indicating that the transposition frequency of the bacterial insertion sequence IS] is determined principally by two ISJ-specified proteins. The first, InsA, was previously shown to bind to the ends of the element and to act as a repressor. We present both physical and genetic evidence which reveals that the second, the InsAB' transposase, is a fusion of InsA with the product of a downstream reading frame, InsB'. Synthesis of this protein occurs by a -1 frameshift between the insA and insB' frames. It requires the presence of an intact retroviral-like frameshift signal composed of an A6C motif and a downstream region able to form several alternative secondary structures. In vivo studies show that IS1 transposition activity depends on the relative rather than on the absolute levels of InsA and InsAB'. The ratio is determined primarily at the translational level by frameshifting and appears to be relatively insensitive to large variations in levels of transcription. This novel homeostatic control could therefore protect IS] from activation as a consequence of insertion into active transcription units.
Extracellular RNAs (exRNAs) have been identified in all tested biofluids and have been associated... more Extracellular RNAs (exRNAs) have been identified in all tested biofluids and have been associated with a variety of extracellular vesicles, ribonucleoprotein complexes and lipoprotein complexes. Much of the interest in exRNAs lies in the fact that they may serve as signalling molecules between cells, their potential to serve as biomarkers for prediction and diagnosis of disease and the possibility that exRNAs or the extracellular particles that carry them might be used for therapeutic purposes. Among the most significant bottlenecks to progress in this field is the lack of robust and standardized methods for collection and processing of biofluids, separation of different types of exRNA-containing particles and isolation and analysis of exRNAs. The Sample and Assay Standards Working Group of the Extracellular RNA Communication Consortium is a group of laboratories funded by the U.S. National Institutes of Health to develop such methods. In our first joint endeavour, we held a series ...
The integration host factor of Escherichia coli (IHF) is a small, histone-like protein which part... more The integration host factor of Escherichia coli (IHF) is a small, histone-like protein which participates in the integration of bacteriophage lambda into the E. coli chromosome and in a number of regulatory processes. Our recent footprinting analysis has shown that IHF binds specifically to the ends of the transposable element IS1, as well as to several sites within a short segment of the plasmid pBR322. We have extended our studies of the binding of the IHF molecule to these sites in vitro using a gel retardation assay. We report here that IHF bends the DNA upon binding, as judged from the strong cyclic dependence of the protein-induced mobility shift on the position of the binding site. Using cloned, synthetic ends of IS1 as substrates, we have found that some mutations within the conserved bases of the IHF consensus binding sequence abolish binding, and that alterations of the flanking sequences can greatly reduce IHF binding. The presence of multiple IHF sites on a single DNA fr...
Genomic technologies are best defined as technologies used to manipulate and analyze genomic info... more Genomic technologies are best defined as technologies used to manipulate and analyze genomic information. The evolution of this collective power began in earnest with the invention of DNA cloning in the 1970's and most of the technology derives from the last quarter of the 20th century. The historical impact of these technologies is clearly immense. With the genome sequence becoming available for many organisms, including humans, another new view of biology has recently emerged. This review examines the shape and texture of this recent evolution, with a particular emphasis on new technology: DNA cloning, macromolecular structure analysis (X-ray crystallography and NMR), DNA sequencing, DNA synthesis, amplification by the polymerase chain reaction, and transgenic animals (bacteria through mammals).
A S S phisms (SNPs) [a number comparable to what is already publicly available (21)] Venter et al... more A S S phisms (SNPs) [a number comparable to what is already publicly available (21)] Venter et al. show that these new opportunities-to paraphrase another milestone a dcle-"have not escaped their notice" (22). . 9. The observed 40,000-fold variation in eukaryote haploid DNA content ("Cvalue") is unrelated to organismic complexity or to the numbers of protein-coding genes; see T.
We present evidence that the Escherichia coli DNA binding protein, IHF, plays an important role i... more We present evidence that the Escherichia coli DNA binding protein, IHF, plays an important role in conjugal transfer of the plasmid F. Our results suggest that IHF exerts this effect by positively effecting transcription of the transfer (tra) operon of the plasmid.
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Papers by David Galas