Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P50583 (asymmetrical)
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Listeria monocytogenes and Shigella flexneri are two unrelated facultative intracellular pathogens which spread from cell to cell by using a similar mode of intracellular movement based on continuous actin assembly at one pole of the bacterium. This process requires the asymmetrical expression of the ActA surface protein in L. monocytogenes and the IcsA (VirG) surface protein in S. flexneri. ActA and IcsA share no sequence homology. To assess the role of the two proteins in the generation of actin-based movement, we expressed them in the genetic context of two non-actin polymerizing, non-pathogenic bacterial species, Listeria innocua and Escherichia coli. In the absence of any additional bacterial pathogenicity determinants, both proteins induced actin assembly and propulsion of the bacteria in cytoplasmic extracts from Xenopus eggs, as visualized by the formation of characteristic actin comet tails. E. coli expressing IcsA moved about two times faster than Listeria and displayed longer actin tails. However, actin dynamics (actin filament distribution and filament half-lives) were similar in IcsA- and ActA-induced actin tails suggesting that by using unrelated surface molecules, L. monocytogenes and S. flexneri move intracellularly by interacting with the same host cytoskeleton components or by interfering with the same host cell signal transduction pathway.
Mol Microbiol 1995 Nov
PMID:The unrelated surface proteins ActA of Listeria monocytogenes and IcsA of Shigella flexneri are sufficient to confer actin-based motility on Listeria innocua and Escherichia coli respectively. 874 26

To understand the origins of the fragile X syndrome and factors predisposing alleles to instability and hyperexpansion, we have compared the haplotype (using markers FRAXAC1, FRAXAC2, and DXS548) and AGG interspersion patterns of the FMR1 CGG repeat for 214 normal and 16 premutation chromosomes. Association testing between interspersion pattern and haplotype reveals a highly significant (P < 0.002) non-random distribution, indicating that all three markers are useful in phylogenetic reconstruction of mutational change. Parsimony analysis of the FMR1 CGG repeat substructure predicts that loss of AGG interruptions has occurred independently on many haplotypes associated with the fragile X syndrome, partially explaining the haplotype diversity of this disease. Among haplotypes found in linkage disequilibrium with the fragile X mutation, two different modes of mutation and predisposition to instability have been identified. One pathway has involved the frequent and recurrent loss of AGG interruptions from rare asymmetrical ancestral array structures. Intergenerational transmission studies suggest that these predisposed chromosomes progress relatively rapidly to the disease state. In contrast, the second mutational pathway involves a single haplotype which has maintained two AGG interruptions. Parsimony analysis of CGG repeat substructure within this haplotype suggests that larger alleles have been generated by gradual increments of CGG repeats distal to the most 3' interruption. Pedigree analysis of the intergenerational stability of alleles of this haplotype confirms a gradual progression toward instability thresholds. As a result, a large reservoir of chromosomes carrying large repeats on this haplotype exists. These chromosomes are predisposed to disease. The present data support a model in which there are at least two different mutational pathways predisposing alleles to instability and hyperexpansion associated with the fragile X syndrome.
Hum Mol Genet 1996 Mar
PMID:Haplotype and interspersion analysis of the FMR1 CGG repeat identifies two different mutational pathways for the origin of the fragile X syndrome. 885 55

Exposure of cultured neonatal rat heart cells to simulated ischaemia results in a cessation of the spontaneous contractile activity and changes at both the level of sarcolemmal phospholipid topology and the ultrastructural level. Reperfusion at a timepoint before irreversible cell damage develops leads to a recovery of contractile activity. Furthermore, the shift in transbilayer distribution of sarcolemmal phosphatidylethanolamine in favour of the outer membrane leaflet, due to the ischaemic period, is reversed during subsequent reperfusion. Also the morphological changes (mitochondrial oedema, reorganization of the mitochondrial cristae and the formation of extrusions at the sarcolemma) are reversible. At the same time total intracellular ATP levels are restored to 80% of control. The role of cellular ATP content on sarcolemmal phospholipid topology was further studied by the use of the calcium antagonist verapamil (10 microM), which preserved cellular ATP content by inhibiting cell contractility before the onset of ischaemia. After 120 min of ischaemia, cell ATP content was still 63% of control in the presence of verapamil, versus 20% of control in untreated cells. Verapamil treatment also prevented the loss of the asymmetrical distribution of phosphatidylethanolamine and sarcolemmal disruption, the latter occurring during 120 min of ischaemia in untreated cells. It is proposed that maintenance of phospholipid asymmetry of the sarcolemma of the myocytes depends on the cellular ATP concentrations, indicating the involvement of an ATP dependent aminophospholipid translocase.
Mol Membr Biol
PMID:Sarcolemmal phosphatidylethanolamine reorganization during simulated ischaemia and reperfusion: reversibility and ATP dependency. 890 44

PI-SceI, a double-stranded DNA endonuclease from Saccharomyces cerevisiae, is generated by protein splicing of an intein, which is an internal polypeptide within a larger precursor protein. The enzyme initiates the mobility of the intein by cleaving at inteinless alleles of the VMA1 gene. Genetic and biochemical studies reveal that the enzyme makes numerous base-specific and phosphate backbone contacts with its 31 bp asymmetrical recognition site. This site can be divided into two regions, both of which contain nucleotides that are essential for cleavage by PI-SceI. Region I contains the PI-SceI cleavage site while Region II includes an adjacent sequence that covers two helical turns. Mutational, interference and DNA mobility shift analyses demonstrate that Region II is sufficient for high-affinity PI-SceI binding. Within this region, PI-SceI uses primarily phosphate backbone and some major groove interactions to contact the DNA, while within Region I, protein binding involves predominantly major groove interactions that overlap and lie proximal to the cleavage site. Interestingly, DNA binding by PI-SceI induces DNA conformational changes within Region II that are entirely exclusive of Region I sequences. Furthermore, additional distortion occurs when PI-SceI binds to Region I in conjunction with Region II. The importance of this latter distortion in the cleavage pathway is underscored by substrate mutations at or near the cleavage site that reduce or eliminate both Region I DNA bending and substrate cleavage. Based on these findings, we propose a model in which sequence-specific contacts made by PI-SceI contribute to its localization to the cleavage site and to its stabilization of a DNA conformation that is required for catalysis. Finally, we discuss how the recognition characteristics of PI-SceI may have allowed the evolution of other endonucleases with altered, but similar, specificities.
J Mol Biol 1996 Oct 25
PMID:Substrate recognition and induced DNA distortion by the PI-SceI endonuclease, an enzyme generated by protein splicing. 891 99

Immunofluorescence microscopy was used to visualize the FtsZ band that marks the site of septation in Sporosarcina ureae. Image analysis indicated that the vegetative division was symmetrically located with respect to the ends of the cells. Fusions of lacZ to the sporulation loci, spollA and cotE, of Bacillus subtilis were introduced into S. ureae by mobilization of plasmids containing the fusions from Escherichia coli. The fusions showed similar patterns of sporulation-associated expression in S. ureae to those observed in B. subtilis. Formation of beta-galactosidase encoded by the spollA-lacZ fusion made it possible to identify early sporulating cells by immunofluorescence microscopy. Analysis of the position of FtsZ bands in cells expressing spollA-lacZ indicated that the location of sporulation division was symmetrical with respect to the ends of the cells, in sharp contrast to the asymmetrical location of septation in sporulating Bacilli. It is inferred that asymmetry of location of the sporulation division is not essential for the compartmentalization of gene expression that follows the division.
Mol Microbiol 1997 Sep
PMID:The division during bacterial sporulation is symmetrically located in Sporosarcina ureae. 935 Aug 65

The bacteriophage Mu C gene encodes a 16.5 kDa site-specific DNA binding protein that is a transcriptional activator of the four "late" promoters, Pmom, Plys, PI and PP. A symmetrical consensus C recognition sequence, TTAT[N5-6]ATAA, containing an inverted tetrad repeat separated by a spacer of five to six G+C-rich nucleotides, has been proposed. To investigate this, we used oligonucleotide mutagenesis to introduce random substitutions within and flanking the proposed C-target region; each variant site was tested for C recognition by an in vivo functional transactivation assay. We observed that all single mutations, in either tetrad, reduced C activation. Although two out of ten substitutions within the spacer reduced activation, the spacer region does not appear to make specific contact with C. We also used in vitro chemical-protection and -interference to study C contacts with Pmom. The results indicate that C contacts Pmom DNA on only one face of the helix through interactions within two adjacent major grooves; this conclusion was supported by gel shift analyses using synthetic oligonucleotide duplexes containing I.C or other base-pair substitutions. Evidence is also presented that C-Pmom contacts are asymmetrical, and that they extend two nucleotides 3' to the promoter-proximal tetrad. We also show that C binding induces a deformation, possibly a bend, in Pmom DNA.
J Mol Biol 1997 Nov 07
PMID:Interaction of the bacteriophage Mu transcriptional activator protein, C, with its target site in the mom promoter. 936 69

In a previous study, samples of the grain aphid Sitobion avenae (F.) were collected from wheat and adjacent cocksfoot hosts in a population thought to be primarily parthenogenetic, and DNA from individual aphids was analysed with a multilocus technique. Here we have applied single-locus microsatellites and a mitochondrial DNA marker to a subset of the same DNA extracts, and have made several additional inferences about important genetic and population processes in S. avenae. Microsatellite analysis indicated very high levels of genic and genotypic variation. S. avenae fell into three genotypic groups inferred to be almost noninterbreeding, while analysis of linkage and Hardy-Weinberg equilibria suggested high levels of sexual recombination within each genotypic group. Host specialization was evident: one lineage was found only on wheat, and one (bearing many alleles inferred to be introgressed from the blackberry-grass aphid S. fragariae (Walker)) was found only on cocksfoot. The third group of interrelated genotypes was found commonly on both hosts. Although most genotypes were found only once, some were much more numerous in the sample than expected from the frequency of the alleles they contained. This, and rapid temporal changes in genotypic composition of samples, indicates strong selective differences between genotypes and lineages. In the major genotypic group, the commonest genotypes were significantly more homozygous than were rare ones: thus these data may help to explain the frequent observation of homozygous excess in aphid allozymes. The genotype group showing S. avenae-like as well as S. fragariae-like alleles also carried S. fragariae-like mitochondrial DNA in at least 25/31 cases, indicating gender-asymmetrical hybridization.
Mol Ecol 1997 Nov
PMID:Genetic structure of an aphid studied using microsatellites: cyclic parthenogenesis, differentiated lineages and host specialization. 939 64

The pituitary cell-specific transcription factor Pit-1 has been show to trans-activate expression of the prolactin (PRL) promoter in non-pituitary cells. However, the cyclic AMP response element (CRE)-binding protein CREB is known to play a major role in cell-specific expression of hepatocyte-specific genes. Since the PRL promoter contains an asymmetrical form of a cyclic AMP response element (termed the CLE), we investigated whether CREB could also induce PRL promoter activity in non-pituitary cells. Transient expression in rat glial C6 cells of a constitutively active CREB-VP16 fusion protein strongly trans-activated expression of a co-transfected rat PRL promoter construct, (-187)PRL-CAT. Analysis by 5'-deletion showed that this response requires PRL promoter sequences between positions -113/-75. CREB-VP16 did not stimulate expression in C6 cells of any of three control promoter-CAT constructs, implying that the strong response of the PRL promoter to activated CREB is both promoter-specific, and is not due to non-specific transcriptional effects of the potent VP16 moiety of CREB-VP16. Surprisingly, mutations in the CLE only slightly reduced activation by CREB-VP16 of construct (-204)PRL-CAT, implying that the major action of CREB-VP16 on the PRL promoter does not involve a direct interaction with the CLE. CREB-VP16 stimulated PRL-CAT activity in C6 cells as strongly as, and synergistically with, Pit-1. These results imply that CREB can strongly and specifically activate expression of the PRL promoter in non-pituitary cells, via a mechanism different from that employed by Pit-1.
Mol Cell Endocrinol 1994 May
PMID:A constitutively active form of CREB can activate expression of the rat prolactin promoter in non-pituitary cells. 939 71

The objectives of this research were: 1) to investigate the time course of the cytogenetic defects induced by acrylamide (AA) treatment (5 x 50 mg/kg) of male germ cells in first-cleavage zygote metaphases using PAINT/DAPI analysis, and 2) to characterize the correlation between chromosomal aberrations at first cleavage, dominant lethality, and heritable translocations. PAINT/DAPI analysis employs multicolor fluorescence in situ hybridization painting plus DAPI staining to detect both stable and unstable chromosomal aberrations at first-cleavage metaphase of the zygote. High levels of chromosomally defective zygotes were detected after mating at all postmeiotic stages (20-190-fold, P < 0.001). Early spermatozoa (6.5 d post-treatment) were the most sensitive, with 76% of the zygotes carrying cytogenetic defects. A significant 10-fold increase was also detected 27.5 d post-treatment, indicating that AA had a cytogenetic effect on meiotic stages. PAINT/DAPI analysis revealed that: 1) AA-induced chromosomal breaks occurred at random, and 2) the frequencies of symmetrical and asymmetrical exchanges were similar at all mating days, except 9.5 d after AA treatment, where significantly (P < 0.02) more asymmetrical aberrations were found. Furthermore, the proportions of zygotes carrying unstable and stable chromosomal aberrations followed a similar post-treatment time course as the proportions of dominant lethality among embryos and heritable translocations among offspring. These findings indicate that PAINT/DAPI analysis of zygotic metaphases is a promising method for detecting male germ cell mutagens capable of inducing chromosomal aberrations and for evaluating the associated risks for embryonic loss and balanced translocations at birth.
Environ Mol Mutagen 1997
PMID:Induction of chromosomal aberrations in mouse zygotes by acrylamide treatment of male germ cells and their correlation with dominant lethality and heritable translocations. 943 82

ApoB RNA-editing enzyme (APOBEC-1) is a cytidine deaminase. Molecular modeling and mutagenesis show that APOBEC-1 is related in quaternary and tertiary structure to Escherichia coli cytidine deaminase (ECCDA). Both enzymes form a homodimer with composite active sites constructed with contributions from each monomer. Significant gaps are present in the APOBEC-1 sequence, compared to ECCDA. The combined mass of the gaps (10 kDa) matches that for the minimal RNA substrate. Their location in ECCDA suggests how APOBEC-1 can be reshaped to accommodate an RNA substrate. In this model, the asymmetrical binding to one active site of a downstream U (equivalent to the deamination product) helps target the other active site for deamination of the upstream C substrate.
J Mol Biol 1998 Jan 30
PMID:Escherichia coli cytidine deaminase provides a molecular model for ApoB RNA editing and a mechanism for RNA substrate recognition. 946 41


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