摘要:

SummaryPhosphorylation of the α subunit of eukaryotic initiation factor 2 (elF‐2α) is one of the best‐characterized mechanisms for down‐regulating total protein synthesis in mammalian cells in response to various stress conditions. Recent work indicates that regulation of theGCN4gene ofSaccharomyces cerevisiaeby amino acid availability represents a gene‐specific case of translational control by phosphorylation of elF‐2α, Four short open reading frames in the leader ofGCN4mRNA (uORFs) restrict the flow of scanning ribosomes from the cap site to theGCN4initiation codon. When amino acids are abundant, ribosomes translate the first uORF and reinitiate at one of the remaining uORFs in the leader, after which they dissociate from the mRNA. Under conditions of amino acid starvation, many ribosomes which have translated uORFI fail to reinitiate at uORFs 2‐4 and utilize theGCN4start codon instead. Failure to reinitiate at uORFs 2‐4 in starved cells results from a reduction in the GTP‐bound form of elF‐2 that delivers charged initiator tRNAiMetto the ribosome. When the levels of elF‐2·GTP·Met‐tRNAiMetternary complexes are low, many ribosomes will not rebind this critical initiation factor following translation of uORF1 until after scanning past uORF4, but before reachingGCN4.Phosphorylation of elF‐2 by the protein kinase GCN2 decreases the concentration of elF‐2·GTP·Met‐tRNAiMetcomplexes by inhibiting the guanine nucleotide exchange factor for elF‐2, which is the same mechanism utilized in mammalian cells to inhibit total protein s

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01947.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryLysyl‐tRNA synthetases are synthesized from two distinct genes inEscherichia coli, lysS(constitutively) andlysU(inducibly); however, the physiological significance and the differential control mechanism of these two genes have been a long‐standing puzzle. Recent studies have successfully uncovered a significant control mechanism oflysUexpression, which involves the leucine‐responsive regulatory protein (Lrp) and a translational enhancer element called‘downstream box'. Moreover, it is likely that there is a mechanism underlying co‐ordinate expression oflysUwith other genes outside the leucine‐Lrp regulon under harsh conditions such as low pH and anaerobiosis. A possible mechanism of lysyl‐tRNA synthetase expression and functio

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01948.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryThePseudomonas aeruginosagenespilB‐DandpilQare necessary for the assembly of type 4 fimbriae. Homologues of these genes and of the subunit (pilin) gene have been described in various different bacterial species, but not always in association with type 4 fimbrial biosynthesis and function. Pil‐like proteins are also involved in protein secretion, DNA transfer by conjugation and transformation, and morphogenesis of filamentous bacteriophages. It seems likely that the Pil homologues function in the processing and export of proteins resembling type 4 fimbrial sub‐units, and in their organization into fimbrial‐like structures. These may either be true type 4 fimbriae, or components of protein complexes which act in the transport of macromolecules (DNA or protein) into or out of the cell. Some PilB‐like and PilQ‐like proteins are apparently also involved in the assembly of non‐type 4 polymeric structures (filamentous phage virions and conjugative pili). The diverse studies summarized in this review are providing insight into an extensive infrastructural system which appears to be utilized in the formation of a variety of cell surface‐assoc

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01949.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryGalactose transport and metabolism inEscherichia coliinvolves a multicomponent amphibolic pathway. Galactose transport is accomplished by two different galactose‐specific transport systems. At least four of the genes and operons involved in galactose transport and metabolism have promoters containing similar regulatory sequences. These sequences are recognized by at least three regulators, Gal repressor (GalR), Gal isorepressor (GalS) and cAMP receptor protein (CRP), which modulate transcription from these promoters. The negative regulators, GalR and GalS, discriminate between utilization of the high‐affinity (regulated by GalS) and low‐affinity (regulated by GalR) transport systems, and modulate the expression of genes for galactose metabolism in an overlapping fashion. GalS is itself autogenously regulated and CRP dependent, while the gene for GalR is constitutive. Thegaloperon encoding the enzymes for galactose metabolism has two promoters regulated by CRP in opposite ways; one (P1) is stimulated and the other (P2) inhibited by CRP. Both promoters are strongly repressed by GalR but weakly by GalS. All but one of the constituent promoters of thegalregulon have two operators. Thegalregulon has the potential to coordinate galactose metabolism and transport in a highly efficient manner, under a wide variety of conditions of galactose availab

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01950.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryExposure of yeast cells to high osmolarities leads to dehydration, collapse of ion gradients over the plasma membrane and decrease in cell viability. The response of yeast cells to high external osmolarities is designated osmostress response. It is likely that both osmoregulatory and general stress reactions are involved in this so far poorly understood process. Part of the response aims at raising the internal osmotic potential, i.e. the production of osmolytes such as glycerol, and exclusion of toxic solutes. In addition, heat‐shock proteins and trehalose are synthesized, probably to protect cellular components and to facilitate repair and recovery. Recent analyses of osmosensitive yeast mutants strongly suggest the involvement of protein kinase‐mediated signal‐transduction pathways in the maintenance of the osmotic integrity of the cell. This has stimulated interesting hypotheses as to the actual osmosensing mech

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01951.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryTheBacillus subtilis ccpAgene has previously been shown to be involved in repression ofamyEexpression when cells are grown in excess glucose. The region of theB. subtilischromosome downstream fromccpAwas characterized to determine if additional genes involved in carbohydrate metabolism were present. Two open reading frames that exhibited sequence similarity to theEscherichia coliandB. subtilis motAandmotBmotility genes were found immediately downstream fromccpA; disruption of this region had no effect on growth, sporulation or motility. Two divergent transcriptional units containing theacsAandacuABCgenes were also found in this region. TheacsAgene encodes acetyl‐CoA synthetase, and inactivation of this gene resulted in loss of the ability to utilize acetate as a carbon source for growth or sporulation. Disruption of theacuABCgenes resulted in poor growth or sporulation on acetoin or butanediol. TheacsAandacuABCpromoter sequences were identified by primer extension, and are in close proximity. Two sequences resembling theamyOregulatory target site necessary for glucose repression ofamyEwere identified in theacsA‐acuABCpromoter regi

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01952.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryExpression of a promoterlesscatgene fused to a DNA fragment of approximately 400 bp, beginning at –313 ofEscherichia coli hns, was significantly repressed inE. coliandSalmonella typhimuriumstrains with wild‐typehnsbut not in mutants carryinghnsalleles. CAT expression from fusions containing a shorter (110 bp) segment ofhnswas essentially unaffected in the same genetic backgrounds. The stage of growth was found to influence the extent of repression which was maximum (approximately 75%) in mid‐log cultures and negligible in cells entering the stationary phase. The level of repression in early‐log phase was lower than in mid‐log phase cultures, probably because of the presence of high levels of Fis protein, which counteracts the H‐NS inhibition by stimulatinghnstranscription. The effects observedin vivowere mirrored by similar results obtainedin vitroupon addition of purified H‐NS and Fis protein to transcriptional systems programmed with the samehnscaf fusions. Electrophoretic gel shift assays, DNase I footprinting and cyclic permutation get analyses revealed that H‐NS binds preferentially to the upstream region of its own gene recognizing two rather extended segments of DNA on both sides of a bend centred around –150. When these sites are filled by H‐NS, an additional site between approximately –20 and –65, which partly overlaps the promoter, is also occupied. Binding of H‐NS to this site is probably the ultimate cause of transc

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01953.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryTheoprFgene, expressingPseudomonas aeruginosamajor outer membrane protein OprF, was subjected to semi‐random linker mutagenesis by insertion of a 1.3 kbHincllkanamycin‐resistance fragment from plasmid pUC4KAPA into multiple blunt‐ended restriction sites in theoprFgene. The kanamycin‐resistance gene was then removed byPstldigestion, which left a 12 nucleotide pair linker residue. Nine unique clones were identified that contained such linkers at different locations within theoprFgene and were permissive for the production of full‐length OprF variants. In addition, one permissive site‐directed insertion, one non‐permissive insertion and one carboxy‐terminal insertion leading to proteolytic truncation were also identified. These mutants were characterized by DNA sequencing and reactivity of the OprF variants with a bank of 10 OprF‐specific monoclonal antibodies. Permissive clones produced OprF variants that were shown to be reactive with the majority of these monoclonal antibodies, except where the insertion was suspected of interrupting the epitope for the specific monoclonal antibody. In addition, these variants were shown to be 2‐mercaptoethanol modifiable, to be resistant to trypsin cleavage in intact cells and partly cleaved to a high‐molecular‐weight core fragment in outer membranes and, where studied, to be accessible to indirect immunofluorescenee labelling in intact cells by monoclonal antibodies specific for surface epitopes. Based on these data, a revised structural mod

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01954.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryIn this paper we show that theEscherichia coliprotein Fis has a regulatory function in Mu transposition in the presence of Mu repressor. Fis can lower the transposition frequency of a mini‐Mu 3–80‐fold, but only if the Mu repressor is expressed simultaneously. In this novel type of regulation of transposition by the concerted action of Fis and repressor, the IAS, the internal activating sequence, is also involved as deletion of this site leads to the loss of the Fis effect. As the IAS contains strong repressor binding sites these are probably the target for the repressor in the observed negative regulation by Fis and repressor. However, the role of Fis and repressor is not only to inactivate the IAS, since a 4bp insertion in the IAS, which changes the spacing of the repressor‐binding site, abolishes the enhancing function of the IAS but leaves the repressor‐Fis effect intact. A likely target for Fis in this regulation is a strong Fis‐binding site, which is located adjacent to the L2 transposase‐binding site. However, when this Fis‐binding sequence was substituted by a random sequence and Fis no longer showed specific binding to this site, the Fis effect was still observed. Although it is still possible that Fis can function by binding to this non‐specific site in a particular complex, it seems more likely that Fis is directly or indirectly involved in determining the level

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01955.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryUsing thetetM‐containing conjugative transposon Tn916 as a mutagenesis tool, we identified two distinct classes of transposon insertions in the meningococcal chromosome. Class I Insertions have an intact copy of Tn916 that appears to have transposed by a novel recombinational mechanism, similar to the transposition of conjugative transposons in Gram‐positive bacteria. Class II insertions were characterized by deletions of Tn916 but preservation of thetetMdeterminant. In addition, we identified Class II Tn916‐like insertions in the naturally occurring 25.2 MDatetM‐containing plasmids of bothNeisseria meningitidisandNeisseria gonorrhoeae.The truncated Tn916 like insertions appeared to be present in the same site in these two plasmids; however, the deletions of the transposon were different. Plasmid sequence adjacent to the truncated transposon in the 25.2 MDa plasmids was found in a tetracycline‐sensitiveN. gonorrhoeae24.5 MDa conjugative plasmid. These data suggest that the 25.2 MDa plasmids are the result of one or a series of Class II Tn916‐like insertions into 24.5 MDa conjugative plasmids. Class II insertions of Tn916‐like transposons are implicated in the dissemination oftetMresistance in pathog

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01956.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY