Showing papers by "Laboratory of Molecular Biology published in 1981"
09 Apr 1981
TL;DR: The complete sequence of the 16,569-base pair human mitochondrial genome is presented and shows extreme economy in that the genes have none or only a few noncoding bases between them, and in many cases the termination codons are not coded in the DNA but are created post-transcriptionally by polyadenylation of the mRNAs.
Abstract: The complete sequence of the 16,569-base pair human mitochondrial genome is presented. The genes for the 12S and 16S rRNAs, 22 tRNAs, cytochrome c oxidase subunits I, II and III, ATPase subunit 6, cytochrome b and eight other predicted protein coding genes have been located. The sequence shows extreme economy in that the genes have none or only a few noncoding bases between them, and in many cases the termination codons are not coded in the DNA but are created post-transcriptionally by polyadenylation of the mRNAs.
8,783 citations
TL;DR: Because it is likely that the authors are approaching saturation of genes affecting primarily the microtubule cells, there appear to be relatively few genes that affect the growth and function of this class of cells and no others.
640 citations
TL;DR: A family of plasmid cloning vectors has been constructed that make use of the leftward promoter of phage λ to provide for efficient expression of cloned genes in Escherichia coli.
632 citations
TL;DR: A change in the position of the distal tip cell in the gonad at an early point in development is correlated with achange in the axial polarity of the germ line tissue in the intact animal.
593 citations
TL;DR: It is suggested that the wild-type unc-86 and lin-4 genes act to modify latent reiterative cell lineages, which are revealed when the activity of one of these genes is eliminated.
526 citations
TL;DR: To be able to sequence genetic material a new approach was desirable and a relatively rapid small-scale method for the fractionation of P-labelled oligonucleotides became the basis for most subsequent studies of RNA sequences.
Abstract: In spite of the important role played by DNA sequences in living matter, it is only relatively recently that general methods for their determination have been developed. This is mainly because of the very large size of DNA molecules, the smallest being those of the simple bacteriophages such as qXl74 (which contains about 5,000 nucleotides). It was therefore difficult to develop methods with such complicated systems. There are however some relatively small RNA molecules notably the transfer RNAs of about 75 nucleotides, and these were used for the early studies on nucleic acid sequences (1). Following my work on amino acid sequences in proteins (2) I turned my attention to RNA and, with G.G. Brownlee and B.G. Barrell, developed a relatively rapid small-scale method for the fractionation of P-labelled oligonucleotides (3). This became the basis for most subsequent studies of RNA sequences. The general approach used in these studies, and in those on proteins, depended on the principle of partial degradation. The large molecules were broken down, usually by suitable enzymes, to give smaller products which were then separated from each other and their sequence determined. When sufficient results had been obtained they were fitted together by a process of deduction to give the complete sequence. This approach was necessarily rather slow and tedious, often involving successive digestions and fractionations, and it was not easy to apply it to the larger DNA molecules. When we first studied DNA some significant sequences of about 50 nucleotides in length were obtained with this method (4,5), but it seemed that to be able to sequence genetic material a new approach was desirable and we turned our attention to the use of copying procedures.
522 citations
TL;DR: In this paper, experiments are reported in which laser microsurgery is used to kill individual cells in the developing somatic gonad; a change observed in the expected behavior of these cells suggests that extrinsic cues may normally play a role in controlling that behavior.
505 citations
TL;DR: Fibroblasts contain a specific internalization pathway that carries hormones as well as some proteins and viruses from the cell surface to the cell interior through the bristle-coated pits of the membrane.
Abstract: Fibroblasts contain a specific internalization pathway that carries hormones as well as some proteins and viruses from the cell surface to the cell interior. Initially, the ligands bind to mobile receptors that are randomly distributed on the cell surface. Next the ligand-receptor complexes are trapped and concentrated in specialized regions of the membrane termed bristle-coated pits. From the pit a smooth-walled vesicle containing the ligand forms and carries the ligand to the cell interior. Because of its role in receptor-mediated endocytosis, this vesicle has been termed a "receptosome."
474 citations
TL;DR: It is reported here that the imidazole stabilizes bound molecular oxygen with a hydrogen bond, as revealed by neutron diffraction analysis.
Abstract: Myoglobin (Mb) reversibly binds molecular oxygen in vertebrate muscle and consists of a polypeptide chain of 153 residues and one haem, which closely resembles one subunit of a haemoglobin (Hb) tetramer. In oxygenated myglobin (oxyMb) the iron atom is coordinated by four porphyrin nitrogen atoms, Ne of the invariant ‘proximal’ histidine (F8), and molecular oxygen1. The oxygen molecule lies in a tight pocket, bounded by two hydrophobic groups (Phe CD1 Val E11) and the side chain of the ‘distal’ histidine (E7). This histidine is present in Hb and Mb of many different organisms, with substitution by glutamine or leucine found in only a few cases. The function of the residue is not clear, although it does present steric hindrance to linear ligands such as carbon monoxide and favours ‘bent’ ones, such as O2. We report here that the imidazole stabilizes bound molecular oxygen with a hydrogen bond, as revealed by neutron diffraction analysis.
427 citations
TL;DR: A DNA segment close to the 5' end of the chicken adult beta-globin gene contains a hypersensitive site for nuclease action, which is in fact an accessible region which extends from approximately 60 to approximately 260 base pairs 5' from the start of mRNA transcription.
362 citations
TL;DR: The properties of mutations of the homoeotic gene extra sex combs indicate that the product of this gene is required early in development for correctly initiating segmental determination.
Abstract: The properties of mutations of the homoeotic gene extra sex combs (esc) indicate that the product of this gene is required early in development for correctly initiating segmental determination. Specifically, the esc+ gene product appears to be a necessary component of the process by which other homoeotic genes, such as those of the bithorax complex, are selectively turned on, or off, in particular segmental primordia. The resulting combinations of active and inactive genes established at this time then maintain the specific pathways of development followed subsequently by the different segments.
TL;DR: The phenotypes of cell lineage mutants of the nematode Caenorhabditis elegans are described, which define 14 genes, which differ in the breadth and nature of their phenotypic defects.
TL;DR: This approach has been extended to study the periodicity of defined-sequence DNAs using another enzyme, micrococcal nuclease, as well as DNase I, and finds the helical repeat of poly(dA-dT) to be 10.5±<0.1 base pairs, a value very close to that found earlier by us for random-sequence DNA.
Abstract: We have recently described a new approach to measuring the helical periodicity of random sequence DNA in solution1. It consists of binding short, stiff pieces of DNA to various flat surfaces and using DNase I to probe the most accessible phosphodiester bonds. The differences in length of the fragments produced are measured with high precision to give the helical repeat directly. This approach has now been extended to study the periodicity of defined-sequence DNAs using another enzyme, micrococcal nuclease, as well as DNase I. We find the helical repeat of poly(dA-dT) to be 10.5±<0.1 base pairs (bp), a value very close to that found earlier by us for random-sequence DNA, 10.6±0.1 bp, and confirmed here. Poly(dA).poly(dT) exhibits a distinctly different helical repeat, 10.0±0.1 bp.
TL;DR: The isolation and characterization of apparent null alleles of the Antp locus lead to a homoeotic phenotype which is the reverse of the dominant Antennapedia phenotype, namely, they result in the transformation of the second leg into an antenna but do not alter the development of the normal antenna itself.
Abstract: Insects are thought to have evolved from millipede-like ancestors composed largely of a series of identical, leg-bearing segments. This view of insect evolution is supported by the existence of homoeotic mutations which transform particular abdominal and head segments into thoracic segments1–3. Such transformations are described as atavistic3, because they return specialized segments to a more primitive condition. In Drosophila, several dominant mutations of the homoeotic locus Antennapedia (Antp) lead to a transformation of the antenna to the second leg4–8. Here, I describe the isolation and characterization of apparent null alleles of the Antp locus. These mutations lead to a homoeotic phenotype which is the reverse of the dominant Antennapedia phenotype, namely, they result in the transformation of the second leg into an antenna but do not alter the development of the normal antenna itself. This result indicates that (1) the product of the wild-type Antp gene is normally active in the mesothorax where it promotes a mesothoracic pathway of development instead of an antennal pathway, (2) the Antp+ gene product is normally absent or inactive in the antenna, and (3) dominant mutations of the locus result in the inappropriate activity of the wild-type gene product in the antenna, and hence in the transformation of antenna to leg. Thus, unlike most other homoeotic gene products, the product of the Antp+ gene seems to promote, not to repress or modify, an atavistic condition.
TL;DR: Comparison with haemagglutinin molecules shows that the H1 and H2 subtypes are more homologous to one another than to the other subtypes, which suggests that they have diverged only recently from one to another.
Abstract: Influenza remains a serious cause of disease in man and of death in the aged. Vaccination is only transiently effective in providing immunity1, because the surface haemagglutinin molecule of the virus can evolve rapidly, allowing the virus to escape neutralization by immunizing antibody. Of the three influenza A subtypes2 known to cause infection in man, the H1 subtype is of particular interest as it has proved to be the dominant human subtype this century and is now co-circulating with variants of the H3 subtype. Here we present the sequence of the haemagglutinin gene of an early H1 subtype (strain A/PR/8/34) as determined by recombinant DNA methods and dideoxy sequencing. From the deduced amino acid sequence of the haemagglutinin we define an antigenic site at amino acid residue 160. Comparison with haemagglutinin molecules3–9 shows that the H1 and H2 subtypes are more homologous to one another than to the other subtypes, which suggests that they have diverged only recently from one to another.
TL;DR: Using complement from mouse strains carrying the H‐2k haplotype it was found that, under conditions of complement limitation and saturating antibody, a fixed amount of complement could lyse about 3 to 4 times as many IgG‐coated sheep red cells as IgM‐ coated red cells.
Abstract: The ability of monoclonal mouse IgM, IgD and IgG anti-(4-hydroxy-3-nitrophenyl) acetyl (NP) antibodies to activate mouse complement was studied using a hemolytic assay. Efficient hemolysis was obtained with IgG2a, IgG2b and IgG3 antibodies but no lysis was observed using a monoclonal IgD. Of several IgG1 antibodies tested, three gave no detectable hemolysis, although weak but significant hemolysis was obtained with two other IgG1. IgM was found to be powerfully hemolytic in that it was effective at lower concentrations than were IgG. However, using complement from mouse strains carrying the H-2k haplotype it was found that, under conditions of complement limitation and saturating antibody, a fixed amount of complement could lyse about 3 to 4 times as many IgG-coated sheep red cells as IgM-coated red cells. This discrimination between IgM and IgG as regards the efficiency of complement utilization is controlled by a gene in the S region of H-2 and is not apparent with complement from mice carrying the b, d or s allele at that locus.
TL;DR: These results identify for the first time native proteins whose stability is affected by the Lon proteolytic pathway and indicate that the Lon system may be important in regulating gene expression in E. coli.
TL;DR: The nucleotide sequence of the promoter distal region of the atp (or unc) operon of Escherichia coli has been determined and it encodes the gamma, beta and epsilon subunits of the ATP-synthase complex and includes a noncoding sequence in which transcription of the operon probably terminates.
Abstract: The nucleotide sequence of the promoter distal region of the atp (or unc) operon of Escherichia coli has been determined. It encodes the gamma, beta and epsilon subunits of the ATP-synthase complex and includes a noncoding sequence in which transcription of the operon probably terminates. This work completes the nucleotide sequence of the operon which contains nine genes: eight encode structural proteins of the ATP-synthase complex; a ninth, the first in the operon, may be a pilot for assembly. The genes for the alpha and beta subunits have evolved from a common ancestor.
TL;DR: Granaticin, an antibiotic which prevents the charging of leucyl-transfer ribonucleic acid, was used to show that the stringent response inhibited the formation of xanthosine monophosphate from inosine Monophosphates in Bacillus subtilis cultures.
Abstract: We have investigated the changes in the guanosine 5′-triphosphate (GTP) and P-ribosyl-PP pools in stringent and relaxed strains of Bacillus subtilis under conditions frequently used to initiate sporulation. After a shift-down from a Casamino Acids-glutamate to a glutamate medium (Sterlini-Mandelstam shift-down), the pools of adenosine 5′-triphosphate and P-ribosyl-PP increased in both strains; in the stringent strain, ppGpp and pppGpp increased and GTP decreased rapidly, whereas in the relaxed strain, ppGpp and pppGpp increased only slightly and GTP decreased only slowly and less extensively. The stringent strain sporulated well, whereas the relaxed strain sporulated late and poorly. Addition of decoyinine, an inhibitor of guanosine 5′-monophosphate synthetase, caused a further decrease of GTP and initiated good sporulation of the relaxed strain. After a shift-down from a glucose-lactate to a lactate medium (Ramaley-Burden shift-down) the pool of P-ribosyl-PP (and GTP) decreased in both strains, indicating a shortage of purine precursors. This shift-down also caused a stringent response which prevented the consumption of nucleotides, as shown by the maintenance of adenosine 5′-triphosphate at a high concentration in the stringent strain but not in the relaxed strain. After a delay, the relaxed strain, in which GTP decreased as fast as in the stringent strain, sporulated also as efficiently. In nutrient sporulation medium the stringent strain and, less effectively, the relaxed strain accumulated ppGpp and pppGpp transiently towards the end of exponential growth. Eventually, the P-ribosyl-PP pool decreased drastically in both strains. In all cases the initiation of sporulation was correlated with a significant decrease of GTP. Granaticin, an antibiotic which prevents the charging of leucyl-transfer ribonucleic acid, was used to show that the stringent response inhibited the formation of xanthosine monophosphate from inosine monophosphate. It prevented the accumulation of xanthosine monophosphate in decoyinine-treated cultures of the stringent strain but not in those of the relaxed strain.
TL;DR: The complete structure of the neuraminidase gene in influenza A/PR/8/34 has been determined by cloning into the bacteriophage M13 and sequencing with dideoxynucleotide chain terminators.
Abstract: The complete structure of the neuraminidase gene in influenza A/PR/8/34 has been determined by cloning into the bacteriophage M13 and sequencing with dideoxynucleotide chain terminators. The gene is 1,413 nucleotides long, codes for a protein of 454 amino acids and has five potential glycosylation sites. We suggest that the neuraminidase, unlike the influenza haemagglutinin, is oriented with its N-terminus buried in the viral membrane.
TL;DR: The results obtained with the alternating copolymers support the suggestion that regions of double-stranded DNA containing alternating purine-pyrimidine sequences may exist as structural variants of the classical B-form under physiological salt conditions.
TL;DR: The results show that the tRNA Tyr base modifications occur in a sequential order: the early 104 precursor already has five base modifications on it, including all the modifications in the TΨ loop; 11 additional modifications only become apparent in the 92 precursor; and two base modifications are introduced in the anticodon loop only after splicing of the intervening sequence.
TL;DR: It is shown that discrete veins will form provided that the transport permeability increases rapidly enough with flux, and given that the movement of auxin is not too polar, in the sense that there is a substantial amount of diffusive movement of Auxin in addition to polar transport.
Abstract: The hormone auxin is transported through many plant tissues with a definite velocity. It is thought that certain channels, or pumps, located at the basal ends of cells, are responsible for the hormone's transport. It is also known that auxin will induce veins when applied to suitable tissues. T. Sachs has suggested that it is the flow of the hormone that induces vessels. He suggests that discrete strands form because the transport capacity of a pathway increases with the flux that that pathway carries, leading to a canalization of flow. I cast this in the form of a more specific hypothesis: I suppose the permeability for the transport of auxin through the basal plasmalemma of a cell (by means of whatever kind of pump or channel) to increase with flux. I then show that discrete veins will form provided that the transport permeability increases rapidly enough with flux, and provided that the movement of auxin is not too polar, in the sense that there is a substantial amount of diffusive movement of auxin in addition to polar transport. The same hypothesis offers an explanation for the loops of veins found under certain conditions.
TL;DR: It is concluded that the duck/Ukraine/63 hemagglutinin sequence fully corroborates its previous identification by immunological and other methods as belonging to the H3 subtype and provides support at the molecular level for the general idea that the wide variety of influenza viruses known to be present in animals can serve as a gene reservoir for human influenza A viruses.
TL;DR: It is concluded that the polarity that has previously been assumed for the thin filament is incorrect and a revised steric blocking model to be constructed, involving a reversal of thin filament polarity combined with the change in the geometry of myosin subfragment-1-actin interaction.
TL;DR: The optical and circular dichroism spectra show that the heme environment in HbI and HbII resembles that of vertebrate hemoglobins, although distinctive features are present in the deoxygenated derivative.
TL;DR: The catalytic triad involving Asp102, His57 and Ser195 agree well with the results found for trypsin and Streptomyces griseus protease A and the results are discussed in terms of the slow, pH-dependent interconversion of α- and γ-chymotrypsin.
TL;DR: The stimulation of adrenergic differentiation of neural crest cells is the first evidence for a positive regulatory role of fibronectin in differentiation.
TL;DR: Specific changes observed in tissue on methylation of reiterated DNA sequences on aging, leukosis and hormone treatment support the idea that DNA methylation is associated with cellular differentiation or transformation and may be one of the possible mechanisms for regulation of transcription and replication in eukaryotes.