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Biomedical subjects

S Brenner

Publications and source records attributed to S Brenner.

At least 163 records · Page 9Linked to original sources
PubMed1996

Surface display of proteins on bacteriophage lambda heads.

We have developed plasmid and phage vectors for the display of foreign proteins on the surface of bacteriophage lambda capsid by modifying the D gene which encodes the major head protein gpD. The vectors have multiple cloning sites, and permit colour selection and conditional chain termination for recombinants. Displayed proteins can be fused to either the N or C terminus of gpD by a peptide linker. The conditional chain termination scheme, via a host Escherichia coli suppressor activity, allows the fusion and assembly of homomultimeric proteins as well as control of the number of fusion proteins per phage particle. We have successfully displayed beta-lactamase, IgG-binding domains of the Staphylococcus aureus protein A, and beta-galactosidase by cloning the genes into the vector. The constructs express functionally active proteins fused to gpD that assemble into phage particles. These results suggest that gpD may be fused to many other peptides and proteins at their N or C terminus and the fusion products may be accessible on the surface of bacteriophage lambda particles.

Amino Acid Sequence↗
PubMed1996

The conserved role of Krox-20 in directing Hox gene expression during vertebrate hindbrain segmentation.

Transient segmentation in the hindbrain is a fundamental morphogenetic phenomenon in the vertebrate embryo, and the restricted expression of subsets of Hox genes in the developing rhombomeric units and their derivatives is linked with regional specification. Here we show that patterning of the vertebrate hindbrain involves the direct upregulation of the chicken and pufferfish group 2 paralogous genes, Hoxb-2 and Hoxa-2, in rhombomeres 3 and 5 (r3 and r5) by the zinc finger gene Krox-20. We identified evolutionarily conserved r3/r5 enhancers that contain high affinity Krox-20. binding sites capable of mediating transactivation by Krox-20. In addition to conservation of binding sites critical for Krox-20 activity in the chicken Hoxa-2 and pufferfish Hoxb-2 genes, the r3/r5 enhancers are also characterized by the presence of a number of identical motifs likely to be involved in cooperative interactions with Krox-20 during the process of hindbrain patterning in vertebrates.

Animals↗
PubMed1996

A functional role for some Fugu introns larger than the typical short ones: the example of the gene coding for ribosomal protein S7 and snoRNA U17.

The compact genome of Fugu rubripes, with its very small introns, appears to be particularly suitable to study intron-encoded functions. We have analyzed the Fugu gene for ribosomal protein S7 (formerly S8, see Note), whose Xenopus homolog contains in its introns the coding sequences for the small nucleolar RNA U17. Except for intron length, the organization of the Fugu S7 gene is very similar to that of the Xenopus counterpart. The total length of the Fugu S7 gene is 3930 bp, compared with 12691 bp for Xenopus. This length difference is uniquely due to smaller introns. Although short, the six introns are longer than the approximately 100 bp size of most Fugu introns, as they host U17 RNA coding sequences. While four of the six U17 sequences are 'canonical', the remaining two represent diverged U17 pseudocopies. In fact, microinjection in Xenopus oocytes of in vitro synthesized Fugu transcripts containing the 'canonical' U17f sequence results in efficient production of mature U17 RNA, while injection of a transcript containing the U17 psi b sequence does not.

Animals↗
PubMed1996

Translocation events in the evolution of aminoacyl-tRNA synthetases.

We have characterized hisS, the gene encoding the histidyl-tRNA synthetase (HisRS) from the tetraodontoid fish Fugu rubripes. The hisS gene is about 3.5 kbp long and contains 13 exons and 12 introns of 172 bp, on average. The Fugu hisS gene encodes a putative protein of 519 amino acids with the three motifs identified as signatures of class 2 aminoacyl-tRNA synthetases. A model for the shifting of intron 8 between Fugu and hamster is proposed based on the successive appearance of a cryptic splicing site followed by an insertion mutation that created a new acceptor site. In addition, sequence comparisons suggest that the hisS gene has undergone a translocation through the first intron. As a result, the Fugu HisRS has an N-terminal sequence markedly different from that in the human and hamster enzymes. We propose that similar events have been responsible for variations at the N-terminal end of other aminoacyl-tRNA synthetases. Our analysis suggests that this involves exchanges through introns of two exons encoding an ancestral 32-amino acid motif.

Amino Acid Sequence↗
PubMed1996

Molecular cloning of two cannabinoid type 1-like receptor genes from the puffer fish Fugu rubripes.

The puffer fish, Fugu rubripes (Fugu), has been proposed as a model vertebrate genome. We have characterized two putative G-protein-coupled receptor encoding genes, FCB1A and FCB1B, obtained by degenerate PCR and low-stringency hybridization of a Fugu genomic library. These two genes show high homology to the human cannabinoid receptor type 1 (HCB1), but very low homology to the type 2 receptor. The amino acid sequences of the FCB1A and FCB1B genes are 66.2% identical, and the homology of each gene to HCB1 is 72.2 and 59.0%, respectively. The transcripts of both the FCB1A and the FCB1B receptors are abundant in the brain. No type 2 receptor could be cloned from Fugu. These data suggest that although two cannabinoid receptor-like genes are found in the puffer fish, both show similarity to the type 1 receptor found in human.

Amino Acid Sequence↗
PubMed1996

Interaction between cAMP-dependent protein kinase catalytic subunit and peptide inhibitors analyzed with lambda repressor fusions.

The lambda phage repressor is currently used as a genetic tool to analyze homodimeric interactions in Escherichia coli. We have applied this system to detect the interaction that takes place within an enzyme-protein inhibitor complex. The sequences encoding the catalytic subunit of the cAMP-dependent protein kinase and the active portion of the natural thermostable protein kinase inhibitor have been fused to the carboxy terminus of the repressor DNA binding domain and introduced into compatible plasmids. Co-expression of the two gene fusions in E. coli lead to the formation of heterodimers that confer a high level of protection from lambda phage infection. The level of lambda immunity depends specifically upon the amino acid sequence of the interacting proteins, as a single amino acid substitution in the inhibitor peptide (Phe10-Ala) restores the sensitivity phenotype.

Amino Acid Sequence↗
PubMed1996

Isolation, characterization and evolution of nine pufferfish (Fugu rubripes) actin genes.

The Japanese pufferfish Fugu rubripes (Fugu) has a small genome of about 400 Mb. Nine different actin genes have been isolated and sequenced from a genomic library constructed from this teleost. The six muscle-type actin genes include two alpha-skeletal actins, three alpha-cardiac actins and an alpha-anomalous (testis type) actin, and the three cytoplasmic actins include two beta-cytoplasmic actins and a beta-cytoplasmic (vascular type) actin. The two skeletal muscle actin genes have identical genomic organization, but differ by five amino acid residues. The three cardiac actin genes code for the same protein but differ in their nucleotide sequences and genomic organization. beta-Cytoplasmic actin1 differs by three amino acids from beta-cytoplasmic actin2. The alpha-anomalous (testis type) and beta-cytoplasmic (vascular type) actins are novel vertebrate actins. The amino acid sequence of alpha-anomalous (testis type) actin is the most divergent of all the known vertebrate actins and transcripts of this gene are abundant in the testis. The beta-cytoplasmic (vascular type) actin gene has eight introns, similar to mammalian smooth muscle actins, and is expressed in vascular tissues such as the gills, kidney and skin. Several known regulatory elements are found in the 5' flanking sequences and the first intron of various Fugu actin genes. The intron patterns of the various Fugu actins seem to be the result of loss of certain introns from a common ancestral gene.

Actins↗
PubMed1996

Cloning and functional expression of cDNAs encoding human and rat pancreatic polypeptide receptors.

PCR was used to isolate nucleotide sequences that may encode novel members of the neuropeptide Y receptor family. By use of a PCR product as a hybridization probe, a full-length human cDNA was isolated that encodes a 375-aa protein with a predicted membrane topology identifying it as a member of the G-protein-coupled receptor superfamily. After stable transfection of the cDNA into human embryonic kidney 293 cells, the receptor exhibited high affinity (Kd = 2.8 nM) for 125I-labeled human pancreatic polypeptide (PP). Competition binding studies in whole cells indicated the following rank order of potency: human PP = bovine PP > or = human [Pro34]peptide YY > rat PP > human peptide YY = human neuropeptide Y. Northern blot analysis revealed that human PP receptor mRNA is most abundantly expressed in skeletal muscle and, to a lesser extent, in lung and brain tissue. A rat cDNA clone encoding a high-affinity PP receptor that is 74% identical to the human PP receptor at the amino acid level was also isolated. These receptor clones will be useful in elucidating the functional role of PP and designing selective PP receptor agonists and antagonists.

Amino Acid Sequence↗
PubMed1996

Out of Africa.

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Journal Article↗
PubMed1996

Conservation of synteny between the genome of the pufferfish (Fugu rubripes) and the region on human chromosome 14 (14q24.3) associated with familial Alzheimer disease (AD3 locus)

The genome of the pufferfish (Fugu rubripes) (400 Mb) is approximately 7.5 times smaller than the human genome, but it has a similar gene repertoire to that of man. If regions of the two genomes exhibited conservation of gene order (i.e., were syntenic), it should be possible to reduce dramatically the effort required for identification of candidate genes in human disease loci by sequencing syntenic regions of the compact Fugu genome. We have demonstrated that three genes (dihydrolipoamide succinyltransferase, S31iii125, and S20i15), which are linked to FOS in the familial Alzheimer disease focus (AD3) on human chromosome 14, have homologues in the Fugu genome adjacent to Fugu cFOS. The relative gene order of cFOS, S31iii125, and S20i15 was the same in both genomes, but in Fugu these three genes lay within a 12.4-kb region, compared to >600 kb in the human AD3 locus. These results demonstrate the conservation of synteny between the genomes of Fugu and man and highlight the utility of this approach for sequence-based identification of genes in human disease loci.

Acyltransferases↗
PubMed1996

Loose ends.

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Journal Article↗
PubMed1996

Small is beautiful: comparative genomics with the pufferfish (Fugu rubripes).

As the Human Genome Project advances, it is clear that the emphasis will switch from accumulation of data to their interpretation. Comparative genomics provides a powerful way in which to analyse sequence data. Indeed, there is already a long list of 'model' organisms, which allow comparative analyses in a variety of ways. The very small vertebrate genome of the pufferfish provides a simple and economical way of comparing sequence data from mammals and fish, representing a large evolutionary divergence and so permitting the identification of essential elements that are still present in both species. These elements include genes and the associated machinery that controls their expression; elements that, in many cases, have survived the test of time.

Animals↗
PubMed1996

Penicillamine-induced bullous dermatoses.

The successful therapeutic use of D-penicillamine (DPA) has been hindered by its many adverse effects. Autoimmune bullous syndromes are among the less common adverse DPA reactions; they are not dose dependent and appear late in the treatment of diseases of altered immunity, most often rheumatoid arthritis. The majority of the DPA-induced bullous syndromes belong to the pemphigus spectrum, usually pemphigus foliaceus or erythematosus, have a lower prevalence of demonstrable tissue-fixed or circulating antibodies than spontaneously occurring pemphigus, display abnormal direct immunofluorescent patterns, and have a generally favorable prognosis. However, many cases do exhibit a full-blown chronic disease, unaffected by DPA withdrawal. DPA-induced cicatricial pemphigoid is a severe disease of both mucous and cutaneous involvement with a prognosis similar to the spontaneous disease. Cases of DPA-induced epidermolysis bullosa acquisita and DPA-induced bullous pemphigoid were not sufficiently substantiated by immunofluorescence or immunoprecipitation criteria.

Drug Eruptions↗