Characterization of DNAs fromCoprinus lagopus andMucor azygospora

Resumé Les rapports G+C des ADN deCoprinus lagopus etMucor azygospora ont été étudiés. Le profil de fusion indique que l'ADN duC. lagopus est composé de deux fractions, une principale (90%) de rapport G+C 52 moles pourcent, une mineure (10%) de G+C 32 moles pourcent. Par contre l'ADN deM. azygospora contient une fraction unique de G+C 38 moles pourcent. L'étude de la cinétique de réassociation DNA:DNA montre que la dimension génomique («genome size») deC. lagopus est de 2×10¹² et qu'il y a moin de 10% de DNA à séquences répétées de nucléotides.

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This research was supported in part by the U.S. Atomic Energy Commission Contract No. AT (40-1) 4182 and the Research Corporation, New York, to S.K.D. We are grateful to ProfessorG. Turian, University of Geneva, for making possibleM. Ojha's participation in this research.     

Monoclonal antibodies to the acetylcholine receptor by a normally functioning auto-anti-idiotypic mechanism

Recently we described a procedure for preparing antibodies to the acetylcholine receptor (AChR) based on immunoglobulin idiotypes and on the hypothesis that, regardless of functional differences, macromolecules of the same specificity will show structural homologies in their binding sites. Antibodies were prepared in rabbits to a structurally constrained agonist of AChR, trans-3,3'-bis[alpha-(trimethylammonio)methyl]azobenzene bromide (BisQ). These antibodies mimicked the binding specificity of AChR in its activated state--agonists were bound with affinities that were in accord with their biological activities and antagonists were bound poorly. Rabbits were then immunized with a specifically purified preparation of anti-BisQ to elicit a population of antibodies specific for the binding sites of anti-BisQ. A portion of the anti-idiotypic antibodies produced in the second set of rabbits cross-reacted with determinants on AChR preparations from Torpedo californica, Electrophorus electricus and rat muscle. Moreover, several of the rabbits showed signs of experimental myasthenia gravis, in which circulating AChR antibodies are typically found. To devise a more direct route to monoclonal anti-receptor antibodies we based our strategy on acceptance of the concept of the anti-idiotypic network theory of Jerne. According to this theory, injection of an antigen elicits, in addition to antibodies to the antigen, other populations that include anti-idiotypic antibodies directed at the combining sites of the antigen-specific antibodies. If the antigen-specific antibodies recognize a ligand of a receptor, then the anti-idiotypic antibodies should bind receptor. Thus, when a mouse is immunized with a bovine serum albumin conjugate of BisQ (BisQ-BSA), it should be possible to expand populations of spleen cells that secrete antibodies which bind anti-BisQ and AChR, in addition to populations specific for BisQ. Fusion of the spleen cells with an appropriate myeloma line should yield monoclonal anti-AChR antibodies. Here we report the success of this approach and its implications.     

A candidate prostate cancer susceptibility gene at chromosome 17p

It is difficult to identify genes that predispose to prostate cancer due to late age at diagnosis, presence of phenocopies within high-risk pedigrees and genetic complexity. A genome-wide scan of large, high-risk pedigrees from Utah has provided evidence for linkage to a locus on chromosome 17p. We carried out positional cloning and mutation screening within the refined interval, identifying a gene, ELAC2, harboring mutations (including a frameshift and a nonconservative missense change) that segregate with prostate cancer in two pedigrees. In addition, two common missense variants in the gene are associated with the occurrence of prostate cancer. ELAC2 is a member of an uncharacterized gene family predicted to encode a metal-dependent hydrolase domain that is conserved among eukaryotes, archaebacteria and eubacteria. The gene product bears amino acid sequence similarity to two better understood protein families, namely the PSO2 (SNM1) DNA interstrand crosslink repair proteins and the 73-kD subunit of mRNA 3' end cleavage and polyadenylation specificity factor (CPSF73).     

Neuronal ensemble control of prosthetic devices by a human with tetraplegia

Neuromotor prostheses (NMPs) aim to replace or restore lost motor functions in paralysed humans by routeing movement-related signals from the brain, around damaged parts of the nervous system, to external effectors. To translate preclinical results from intact animals to a clinically useful NMP, movement signals must persist in cortex after spinal cord injury and be engaged by movement intent when sensory inputs and limb movement are long absent. Furthermore, NMPs would require that intention-driven neuronal activity be converted into a control signal that enables useful tasks. Here we show initial results for a tetraplegic human (MN) using a pilot NMP. Neuronal ensemble activity recorded through a 96-microelectrode array implanted in primary motor cortex demonstrated that intended hand motion modulates cortical spiking patterns three years after spinal cord injury. Decoders were created, providing a 'neural cursor' with which MN opened simulated e-mail and operated devices such as a television, even while conversing. Furthermore, MN used neural control to open and close a prosthetic hand, and perform rudimentary actions with a multi-jointed robotic arm. These early results suggest that NMPs based upon intracortical neuronal ensemble spiking activity could provide a valuable new neurotechnology to restore independence for humans with paralysis.     

Implications of epidermal-dermal relations following injury for understanding association of carcinogenesis with hair regeneration or scars

Zusammenfassung Während des normalen Heilungsvorganges in Hautverletzungen wurde nachgewiesen, dass das Epithel die unterliegende traumatisierte Cutis « überfällt ». Normalerweise werden solche « überfallenden » Epithelsporen entweder durch Fremdkörperreaktion des Bindegewebes eliminert, oder aber eine « kompetente » Cutis bewirkt die Bildung von Talgdrüsen oder neuer Haarfollikel. In Anbetracht der engen Korrelation zwischen Haarbildung, Narben und Carcinogenese wird auf Grund der histologischen Befunde der Autoren und aus einer Übersicht der einschlägigen embryologischen Forschungsliteratur gefolgert, dass das endgültig massgebliche Verhalten solcher « überfallenden » pluripoten tialen Epithelsporen während des Wundheilungsvorganges teilweise von der Natur der epikutanen Überfälle abhängig ist, und vornehmlich von der Reaktivität der unterliegenden Cutis.Epikutane Neoplasie könnte infolgedessen hauptsächlich als Ergebnis des Versagens einer beschädigten Cutis betrachtet werden, die weder die überfallende Epidermis eliminiert, noch die Haarpapillen differenziert, in ihrer Reaktion gegen überfallende, epidermale Sporen oder neue Haar-Anlagen, die während normalem Heilungsvorgang gebildet wurden; dies gilt für jedwede Verletzung, die chemisch oder physisch verursacht wurde. Überfallende epidermale Sporen könnten zuerst verhornte Zysten bilden, in welchen späterhin neoplastische Veränderungen in Erscheinung treten.     

Mining the human genome using microarrays of open reading frames

To test the hypothesis that the human genome project will uncover many genes not previously discovered by sequencing of expressed sequence tags (ESTs), we designed and produced a set of microarrays using probes based on open reading frames (ORFs) in 350 Mb of finished and draft human sequence. Our approach aims to identify all genes directly from genomic sequence by querying gene expression. We analysed genomic sequence with a suite of ORF prediction programs, selected approximately one ORF per gene, amplified the ORFs from genomic DNA and arrayed the amplicons onto treated glass slides. Of the first 10,000 arrayed ORFs, 31% are completely novel and 29% are similar, but not identical, to sequences in public databases. Approximately one-half of these are expressed in the tissues we queried by microarray. Subsequent verification by other techniques confirmed expression of several of the novel genes. Expressed sequence tags (ESTs) have yielded vast amounts of data, but our results indicate that many genes in the human genome will only be found by genomic sequencing.     

CTCF-binding sites flank CTG/CAG repeats and form a methylation-sensitive insulator at the DM1 locus

An expansion of a CTG repeat at the DM1 locus causes myotonic dystrophy (DM) by altering the expression of the two adjacent genes, DMPK and SIX5, and through a toxic effect of the repeat-containing RNA. Here we identify two CTCF-binding sites that flank the CTG repeat and form an insulator element between DMPK and SIX5. Methylation of these sites prevents binding of CTCF, indicating that the DM1 locus methylation in congenital DM would disrupt insulator function. Furthermore, CTCF-binding sites are associated with CTG/CAG repeats at several other loci. We suggest a general role for CTG/CAG repeats as components of insulator elements at multiple sites in the human genome.