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Biomedical subjects

M Mayford

Publications and source records attributed to M Mayford.

25 records · Page 2Linked to original sources
PubMed1995

Transgenic approaches to cognition.

Reverse genetic techniques, including gene 'knockouts' and transgenesis, allow defined mutations to be introduced into the mouse genome. The application of these techniques to neurobiology is beginning to provide a bridge between genes and cognition. Specifically, genetically altered mice make it possible to explore molecular mechanisms underlying implicit and explicit forms of learning, short-term and long-term memory, and emotional behaviors. The analysis of these mutant mice has begun to link specific behavioral deficits to defined changes in synaptic physiology.

Animals↗
PubMed1992

Modulation of an NCAM-related adhesion molecule with long-term synaptic plasticity in Aplysia.

A form of learning in the marine mollusk Aplysia, long-term sensitization of the gill- and siphon-withdrawal reflex, results in the formation of new synaptic connections between the presynaptic siphon sensory neurons and their target cells. These structural changes can be mimicked, when the cells are maintained in culture, by application of serotonin, an endogenous facilitating neurotransmitter in Aplysia. A group of cell surface proteins, designated Aplysia cell adhesion molecules (apCAM's) was down-regulated in the sensory neurons in response to serotonin. The deduced amino acid sequence obtained from complementary DNA clones indicated that the apCAM's are a family of proteins that seem to arise from a single gene. The apCAM's are members of the immunoglobulin class of cell adhesion molecules and resemble two neural cell adhesion molecules, NCAM and fasciclin II. In addition to regulating newly synthesized apCAM, serotonin also altered the amount of preexisting apCAM on the cell surface of the presynaptic sensory neurons. By contrast, the apCAM on the surface of the postsynaptic motor neuron was not modulated by serotonin. This rapid, transmitter-mediated down-regulation of a cell adhesion molecule in the sensory neurons may be one of the early molecular changes in long-term synaptic facilitation.

Amino Acid Sequence↗
PubMed1990

The ermC leader peptide: amino acid alterations leading to differential efficiency of induction by macrolide-lincosamide-streptogramin B antibiotics.

The inducibility of ermC by erythromycin, megalomicin, and celesticetin was tested with both wild-type ermC and several regulatory mutants altered in the 19-amino-acid-residue leader peptide, MGIFSIFVISTVHYQP NKK. In the model test system that was used, the ErmC methylase was translationally fused to beta-galactosidase. Mutational alterations that mapped in the interval encoding Phe-4 through Ile-9 of the leader peptide not only affected induction by individual antibiotics, but did so differentially. The subset of mutations that affected inducibility by the two macrolides erythromycin and megalomicin overlapped and were distinct from the subset of mutations that affected induction by celesticetin. These studies provide a model system for experimentally varying the relative efficiencies with which different antibiotics induce the expression of ermC. The possibility that antibiotics with inducing activity interact directly with the nascent leader peptide was tested by using a chemically synthesized decapeptide, MGIFSIFVIS--, attached at its C-terminus to a solid-phase support. This peptide, however, failed to bind erythromycin in vitro.

Amino Acid Sequence↗
PubMed1989

Conformational alterations in the ermC transcript in vivo during induction.

ermC is an inducible antibiotic resistance gene from Staphylococcus aureus, one of several whose expression is regulated at the level of mRNA secondary structure. During induction of ermC, the inhibition of a ribosome active in translation of a short leader peptide by low levels of antibiotic belonging to the macrolide-lincosamide-streptogramin b family is believed to cause a rearrangement in mRNA secondary structure. The resultant conformational isomerization unmasks the methylase ribosome binding site and initiator Met codon, causing increased translation of the ermC transcript. Expression of ermC can also be demonstrated in Bacillus subtilis carrying plasmid pE194. To probe the ermC transcript in vivo during induction, ermC was transferred to B. subtilis by transformation and the resultant transformants were treated with dimethyl sulfate which reacts with N-1 of adenine and N-3 of cytosine residues in a manner that is sensitive to secondary structure. The bases modified in vivo were detected by primer extension with reverse transcriptase using total cellular RNA as template and a complementary ermC-specific oligonucleotide as primer. Physical evidence was obtained for the secondary structural rearrangements predicted by the ermC regulatory model. Additionally, physical evidence was obtained demonstrating that during induction, the stalled ribosome protects codons 9 and 10 of the leader peptide from modification by dimethyl sulfate, in agreement with genetic data obtained previously that identified the integrity of codons 5-9 as critical for induction of ermC by erythromycin.

Amino Acid Sequence↗
PubMed1989

ermC leader peptide. Amino acid sequence critical for induction by translational attenuation.

The ermC mRNA leader segment, which encodes a 19 amino acid leader peptide, MGIFSIFVISTVHYQPNKK, plays a key role in regulating expression of the ErmC methylase. The contribution of specific leader peptide amino acid residues to induction of ermC was studied using a model system in which the ErmC methylase was translationally fused to Escherichia coli beta-galactosidase as indicator gene. Codons of the ermC leader peptide were altered systematically by replacement of leader DNA segments with double-stranded DNA constructed from chemically synthesized oligonucleotides. Missense mutations that resulted in reduced efficiency of induction involved codons for amino acid residues 5 to 9 (-SIFVI-). Nonsense mutations causing termination of the leader peptide at codons 10 (-S-) or 12 (-V-) remained inducible. These findings suggest that the codons for residues 5 to 9 of the leader peptide comprise the critical region in which ribosomes stall in the presence of erythromycin.

Amino Acid Sequence↗
PubMed1985

Messenger RNA from Staphylococcus aureus that specifies macrolide-lincosamide-streptogramin resistance. Demonstration of its conformations and of the leader peptide it encodes.

The +1 site for transcription initiation of the inducible 23 S rRNA adenine methylase encoded by plasmid pE194 was determined experimentally by nuclease S1 mapping of mRNA synthesized in vivo, and by nuclease T1 mapping of (5'-gamma-32P)-end-labeled transcripts synthesized in vitro. By partial digestion of the in vitro transcripts using S1 and cobra venom nuclease as probes of mRNA conformation, the analysis was extended to reveal single-stranded and double-stranded regions, respectively, which correspond to the critical stems and loops postulated for active and inactive conformations of the nascent mRNA. According to the model for induction, the transition from inactive to active conformation involves disruption of mRNA secondary structure which, in turn, is predicated on protracted occupancy by ribosomes complexed with erythromycin of one of the critical stem sequences. Ribosome occupancy of the critical stem sequence is due to the presence of an open reading frame that encodes part of a 19 amino acid residue "leader" peptide. The existence of this peptide, deduced from the nucleotide sequence of the control region upstream from the methylase structural gene, was demonstrated in vivo as part of a translational fusion with Escherichia coli beta-galactosidase in which the first four amino acid residues of the N-terminal sequence of the fusion protein, analyzed directly by the microsequencing method, were found to comprise N-terminal amino acids 2 through 5, Gly-Ile-Phe-Ser, predicted for the leader peptide.

Amino Acid Sequence↗