Oculomotor palsy with cyclic spasms.
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Blepharoptosis↗
Biomedical subjects
D I Friedman
Publications and source records attributed to D I Friedman.
Effects of rifampicin resistant rpoB mutations on antitermination and interaction with nusA in Escherichia coli.
Rifampicin resistant (Rifr mutations map in the rpoB gene encoding the beta subunit of Escherichia coli RNA polymerase. We have used our collection of 17 sequenced Rifr mutations to investigate the involvement of E. coli RNA polymerase in the antitermination systems enhancing expression of delayed early lambda genes or stable RNA. We have found that Rifr mutations affect both lambda N-mediated antitermination and the cellular antitermination system involved in synthesis of stable RNA. Because NusA is involved in antitermination and termination, we also investigated the interaction of NusA and RNA polymerase by determining whether Rifr mutations alter NusA-dependent termination or antitermination in cells with defective nusA alleles. We have shown that Rifr mutations can either enhance or suppress the phenotypes of defective nusA alleles. Most Rifr mutations alter the temperature range over which the nusA1 allele supports lambda N-mediated antitermination. In addition, a number of Rifr alleles restore termination to the nusA10(Cs) and the nusA11(Ts) mutants defective in this process. Our results indicate that the region of the rpoB gene defined by the Rifr mutations is involved in the antitermination process and affects the activity of the NusA protein directly or indirectly.
Alleles↗
Integration host factor: a protein for all reasons.
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Bacterial Proteins↗
A point mutation in the Nul gene of bacteriophage lambda facilitates phage growth in Escherichia coli with himA and gyrB mutations.
A mutant of lambda was isolated that grows in the Escherichia coli himA delta/gyrB-him320(Ts) double mutant at 42 degrees C; conditions which are non-permissive for wild-type lambda growth. The responsible mutation, ohm1, alters the 40th codon of the Nul reading frame. The Nul and A gene products comprise the terminase protein which cleaves concatameric DNA into unit-length phage genomes during DNA packaging. The Nul-ohm1 gene product acts in trans to support lambda growth in the double himA/gyrB mutant, and lambda cos154 growth in the single himA mutant. The observation that an alteration in Nul suppresses the inhibition of growth in the double himA/gyrB mutant implicates DNA gyrase, as well as integration host factor, in the DNA:protein interactions that occur at the initiation of packaging.
Bacteriophage lambda↗
Analysis of transcription termination signals in the nin region of bacteriophage lambda: the roc deletion.
Deletions in the region, nin, between the P and Q genes of phage lambda remove a portion of the phage genome that includes signals for termination of transcription. These deletions were selected because they permit growth of lambda derivatives defective in the N-mediated transcription antitermination system; i.e., the deletions confer N independence (nin). Thus nin phages (e.g., lambda nin5) grow in most Escherichia coli nus mutants. The nus genes encode functions necessary for N action. We report the isolation of a deletion in the nin region delta roc that confers a partially N-independent phenotype; lambda derivatives with delta roc can grow under normally nonpermissive conditions, 32 degrees C, in a host with the rpoB-nusC60 mutation. The roc deletion also partially suppresses the inhibitory effects of other nus mutations at higher temperatures. Delta roc, which extends from base pairs 41883 to 43825, overlaps the nin5 deletion, which extend from base pairs 40501 to 43306. Unlike the nin5 deletion, the sequences deleted by delta roc do not include a stem-loop structure, tR2, previously shown to have terminator activity. Using promoter and terminator testor vectors, we demonstrate that a 2,400-base-pair fragment that includes the wild-type roc region but excludes tR2 has terminator activity. Thus, delta roc permits a functional division of the transcription termination signals in the nin region.
Bacteriophage lambda↗
lambda N antitermination system: functional analysis of phage interactions with the host NusA protein.
Coliphage lambda gene expression is regulated temporally by systems of termination and antitermination of transcription. The lambda-encoded N protein (pN) acting with host factors (Nus) at sites (nut) located downstream from early promoters is the first of these systems to operate during phage development. We report observations on some of the components of this complex system that, in part, address the way in which these elements interact to render RNA polymerase termination-resistant. (1) The isolation of a conditionally lethal cold-sensitive nusA mutation demonstrates that NusA is essential for bacterial growth. (2) The effect on lambda growth in a host in which the Salmonella NusA protein is overproduced suggests that NusA is essential for N-mediated antitermination in phage lambda. (3) A truncated NusA product, representing only the amino two-thirds of the native protein, is active for both bacterial growth and pN action, indicating that the carboxy end of the molecule may not be a functionally important region. (4) lambda pN can function with the heterologous nut region from Salmonella typhimurium phage P22 when lambda pN is overproduced, demonstrating that lambda pN can function with the nut regions of other lambdoid phages. (5) A single base-pair change in the lambda nutR boxA sequence that was selected to permit a lambda derivative to utilize the Salmonella NusA protein restores lambda growth in the Escherichia coli nusA1 host.
Bacterial Proteins↗
Integration host factor is required for the DNA inversion that controls phase variation in Escherichia coli.
The on-and-off expression (phase variation) of type 1 fimbriae, encoded by fimA, in Escherichia coli is controlled by the inversion of a promoter-containing 314-base-pair DNA element. This element is flanked on each side by a 9-base-pair inverted, repeat sequence and requires closely linked genes for inversion. Homology analysis of the products of these genes, fimB and fimE, reveals a strong similarity with the proposed DNA binding domain of lambda integrase, which mediates site-specific recombination in the presence of integration host factor. Integration host factor, encoded by himA and hip/himD, binds to the sequence 5' TNYAANNNRTTGAT 3', where Y = pyrimidine and R = purine, in mediating integration-excision. In analyzing the DNA flanking the fim 314-base-pair inversion sequence, we found the adjacent sequence 5' TTTAACTTATTGAT 3', which corresponds perfectly with the consensus integration host factor binding site. To characterize the role of himA in phase variation, we transduced either a deletion of himA or an insertionally inactivated hip/himD gene into an E. coli strain with a fimA-lacZ operon fusion. We found the rate of phase variation decreases sharply from 10(-3) to less than 10(-5) per cell per generation. Southern hybridization analysis demonstrates that the himA mutation results in a failure of the switch-generated genetic rearrangement. When the transductant was transformed with a himA+ plasmid, normal switching returned. Thus integration host factor is required for normal type 1 fimbriae phase variation in E. coli.
Bacterial Adhesion↗
RNA 3' end formation in the control of gene expression.
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Bacteria↗
Identification and characterization of mutations in Escherichia coli that selectively influence the growth of hybrid lambda bacteriophages carrying the immunity region of bacteriophage P22.
Mutations in two Escherichia coli genes, sipA and sipB, result in a specific inhibition of the growth of certain hybrid lambdoid bacteriophages, lambda immP22, that have the early regulatory regions and adjacent genes from bacteriophage P22. The sipB391 mutation maps near minute 56 and exerts the strongest inhibitory effect on the growth of the hybrid phages. The sipA1 mutation maps near minute 72 and plays an auxiliary role: enhancing the action of sipB391. Such a role is not limited to sipA1, since there is a similar enhancement by the nusA1 and nusE71 mutations. The Sip-imposed restriction on the growth of lambda immP22 phages is not observed if the phage carries a mutation in the c1 gene. Perhaps this reflects the fact that the c1 product regulates phage DNA replication and is a major determinant in the decision governing whether the phage takes the lytic or lysogenic pathway. Consistent with this idea is the observation that lambda immP22 DNA replication is severely inhibited in bacteria carrying the sipB391 mutation. It is suggested that sip mutations exaggerate the normal role of c1 in limiting lytic growth. This causes a failure in the expression of sufficient amounts of some or all of the lytic gene products required for phage growth.
Bacteriophage lambda↗
Arteriovenous malformation presenting as hemidystonia.
Two men, aged 25 and 33 years, had progressive hemidystonia and an arteriovenous malformation (AVM) in the contralateral cerebral hemisphere. One patient with an AVM in the posterior basal ganglia of the right hemisphere had an older brother with severe generalized dystonia. The second patient had an AVM in the left cortical and subcortical parietal area with no obvious lesion in the basal ganglia. Unlike generalized dystonia, a focal lesion is commonly found in patients with unilateral dystonia. The association of AVM-induced hemidystonia and family history of dystonia suggests that genetic predisposition may be important in some patients with hemidystonia.
Adult↗
A phi 80 function inhibitory for growth of lambdoid phage in him mutants of Escherichia coli deficient in integration host factor. I. Genetic analysis of the Rha phenotype.
Bacteriophage phi 80 and lambda-phi 80 hybrid phage of the type lambda (QSR)80, in which the rightmost 10% of the lambda genome is replaced by corresponding phi 80 material, are unable to grow lytically in himA and hip/himD mutants of Escherichia coli K12 at 32 degrees. The genetic element responsible for the growth defect, rha, has been mapped to the (QSR)80 region and was located more precisely by restriction enzyme and DNA heteroduplex analysis of mutations that result in loss of the Rha phenotype. Such an Rha mutant carrying a 1.5-kb deletion beginning 0.58 kb from the right end of the chromosome and extending leftward locates the rha locus at least in part within this region of (QSR)80. In addition, a substitution derivative of lambda (QSR)80 was isolated which does not exhibit the Rha phenotype. In this phage, lambda-80hy95, the right half of the (QSR)80 region is replaced by DNA homologous to the 95-100% segment of lambda. In mixed infections in the himA42 host at 32 degrees, lambda + does not complement lambda (QSR)80 for growth and the burst size of the coinfecting lambda + is reduced in comparison to that in a single infection. Deletion mutants of lambda (QSR)80 that grow normally in himA42 at 32 degrees in single infections are inhibited for growth in mixed infections with lambda (QSR)80. These results suggest the existence of a trans-acting function which inhibits phage growth in the absence of HimA or Hip/HimD function. It is likely that the rha gene either encodes that function or indirectly controls its action.
Bacteriophage lambda↗
A phi 80 function inhibitory for growth of lambdoid phage in him mutants of Escherichia coli deficient in integration host factor. II. Physiological analysis of the abortive infection.
Derivatives of phage lambda with the rightmost 3% of the genome (the QSR region) from the related phage phi 80 fail to grow at low temperatures (e.g., 32 degrees) in Escherichia coli hosts deficient in either protein component of IHF (integration host factor), the products of the himA and hip/himD genes. The abortive infection of lambda (QSR)80 in mutants defective for IHF was studied in detail. This infection is characterized by a lack of cell lysis and an inhibition of phage DNA replication after an initial period of normal synthesis. An inhibition of host DNA replication also occurs after a similar period of apparently normal synthesis, and the abortive lambda (QSR)80 infection is lethal to the host. An assay of beta-galactosidase activity in lambda (QSR)80-infected cells provided indirect evidence that RNA and protein synthesis continue late into the abortive infection. The defective growth is imposed by the product of the rha gene located in the (QSR)80 genetic material. Two-dimensional electrophoretic analysis of phage proteins produced in ultraviolet (uv)-irradiated phage-infected host cells has demonstrated the existence of a protein that is encoded or whose synthesis is regulated by the rha locus. Based on these findings, possible roles for a HimA-Hip/HimD-controlled rha product in a late stage of phi 80 development are discussed.
Bacterial Proteins↗
The nusA recognition site. Alteration in its sequence or position relative to upstream translation interferes with the action of the N antitermination function of phage lambda.
The phage lambda transcription antitermination protein, pN, acts with host factors, Nus, at sites on the phage genome, nut, to render RNA polymerase resistant to subsequent downstream termination signals. The NusA protein appears to recognize a seven to eight base-pair consensus sequence (5'Py-G-C-T-C-T-T(T)3') called boxA that is found in the promoter-proximal part of the nut region. Two types of change within or near the boxA sequence in the nutR region are shown to interfere with pN-mediated antitermination of transcription that has initiated at the upstream pR promoter. (1) A change of one base-pair (from G to T at the second position) in the boxA sequence significantly reduces pN action. (2) We prove that a frameshift mutation, cro delta 62, at the end of the gene promoter-proximal to the lambda nutR region, interferes with the pN antitermination reaction by allowing translation to proceed beyond cro into the nutR region. Using a series of plasmid constructions, we now show that the inhibition of antitermination caused by the cro delta 62 mutation can be suppressed when translation is terminated upstream from this mutation.
Bacterial Proteins↗
Synergistic effect of himA and gyrB mutations: evidence that him functions control expression of ilv and xyl genes.
We have constructed Escherichia coli strains containing mutations at two different loci, both originally selected for failure to support lambda site-specific recombination: himA and gyrB-him(Ts). Although the gyrB-him(Ts) mutations by themselves reduce supercoiling at high temperature, the double mutants show a far greater effect on supercoiling. Our studies show that growth of phage lambda is severely inhibited and that maintenance of plasmid pBR322 is extremely unstable in the double mutants. Physiological studies also reveal that the double mutants are isoleucine auxotrophs at 42 degrees C. The fact that himA mutants are isoleucine auxotrophs at 42 degrees C in the presence of leucine suggests that a significant component of the isoleucine auxotrophy of the double mutants is a result of the himA mutation. The himA gene encodes the alpha subunit of a protein called the integration host factor. Since mutations in the hip or himD gene encoding beta, the other subunit of the integration host factor, also result in isoleucine auxotrophy in the presence of leucine, we suggest that the integration host factor regulates the synthesis of at least one of the enzymes in the ilv pathway, acetohydroxyacid synthase I, which is encoded by the ilvB gene. Studies of the utilization of various sugars as the sole carbon source suggest that the integration host factor controls expression of some gene(s) involved in the utilization of xylose.
Bacteriophage lambda↗
Mutations in the DNA gyrB gene that are temperature sensitive for lambda site-specific recombination, Mu growth, and plasmid maintenance.
We report the isolation of two mutations in the gyrB gene of Escherichia coli K12 obtained from an initial selection for resistance to coumermycin A1 and a subsequent screening for bacteria that fail to support site-specific recombination of phage lambda, i.e., Him-. These two mutations have a temperature-sensitive Him- phenotype, supporting site-specific recombination efficiently at low temperature, but inefficiently at high temperatures. Like other Him mutants, the gyrB-him mutants fail to plate phage Mu; again this defect is observed only at high temperatures. Additional thermally sensitive characteristics have also been observed; growth of lambda as well as maintenance of the plasmids pBR322 and F' gal are reduced at high temperature. Restriction of foreign DNA imposed by a P1 prophage is also reduced in these mutants. The temperature-sensitive phenotypic characteristics imposed by both the gyrB-him-230(Ts) and gyrB-him-231(Ts) mutations correlate with in vitro studies that show decreased gyrase activity, especially at higher temperatures, and in vivo studies showing reduced supercoiling of lambda DNA in the mutants at high temperature.
Bacteriophage lambda↗
An accessory role for Escherichia coli integration host factor: characterization of a lambda mutant dependent upon integration host factor for DNA packaging.
Bacteriophage lambda grows lytically on Escherichia coli defective for integration host factor, a protein involved in lambda site-specific recombination and the regulation of gene expression. We report the characterization of a mutant, lambda cos154, that, unlike wild-type lambda, is defective for growth in integration host factor-defective E. coli. The cis-dominant mutation in lambda cos154 is a single base pair change in a region of hyphenated dyad symmetry close to the lambda left cohesive end; this mutation prevents DNA packaging. We propose the following two alternative roles for this site in lambda DNA packaging: (i) to bind an E. coli accessory protein required in the absence of integration host factor or (ii) to bind the phage-encoded terminase protein that is essential for DNA packaging.
Bacterial Proteins↗
Interactions of bacteriophage and host macromolecules in the growth of bacteriophage lambda.
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Bacteriophage lambda↗
Evidence that a nucleotide sequence, "boxA," is involved in the action of the NusA protein.
We report the isolation of a mutation, boxA1, in the nutR region of the phage lambda genome. The nutR region, located downstream of the pR promoter, includes the site nutR where the lambda N protein is thought to act to render subsequent transcription termination-resistant. We have previously suggested that the boxA sequence, 5'CGCTCTTA3' (or its RNA analog), located 8 bp promoter-proximal to nutR, might be the recognition site for the E. coli host factor, NusA, which has been shown to be necessary for N action. The boxA1 mutation, an A:T to T:A transversion, results in a changed boxA sequence upstream of nutR, CGCTCTTT. This change is necessary for lambda to effectively use the NusA of Salmonella typhimurium, a NusA function not normally active with the N product of lambda. Other lambdoid phages with unique N functions and nut sites that are normally active with the NusA of Salmonella have boxA sequences with the terminal three Ts. Moreover, sequences closely resembling boxA have been found near transcription termination sequences in E. coli operons where NusA has been shown to be involved in termination. These findings identify boxA as an important recognition signal for the NusA protein.
Bacterial Proteins↗