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Results for “insertion, inversion”

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PubMed2005

Argonaute protein PIWI controls mobilization of retrotransposons in the Drosophila male germline.

Proteins of the Argonaute family have been identified as key components of RNA interference (RNAi) pathway. RNAi-related mechanisms are implicated in the regulation of gene expression and repression of transposable elements in eukaryotes. The piwi gene encoding protein of the Drosophila Argonaute family was shown to be required for the germ stem cells maintenance. Here, we show that piwi is involved in silencing of LTR retrotransposons in testes. piwi mutations led to derepression of endogenous retrotransposon copia as well as to upregulation of the reporter gene driven by copia LTR. piwi mutation causes accumulation of retrotransposon mdg1 transcripts at the apical tip of testes, including germinal proliferative center where PIWI protein was shown to be expressed. We applied inverse PCR approach to detect the newly arisen insertions of the mdg1 retrotransposon in the progeny of individual piwi mutant males. Owing to piwi mutation a high rate of mdg1 transpositions was revealed. Thus, piwi is involved in the silencing of retrotransposons in the precursors of male gametes. Our results provide the first evidence that protein of the Argonaute family prevents retrotranspositions. It is supposed that the disturbance of RNA silencing system in germinal cells might cause transposition burst.

Animals↗

PubMed1984

Spontaneous deletion of citrate-utilizing ability promoted by insertion sequences.

The citrate utilization (Cit+) transposon Tn3411 was shown to be flanked by directly repeated sequences (IS3411L and IS3411R) by restriction enzyme analysis and electron microscope observation. Cit- deletion mutants were frequently found to be generated in pBR322::Tn3411 by intramolecular recombination between the two copies of IS3411. The flanking IS3411 elements of Tn3411 were shown to be functional insertion sequences by Tn3411-mediated direct and inverse transposition. Tn3411-mediated inverse transposition from pBR322::Tn3411 to the F-plasmid derivative pED100 occurred more efficiently than that of direct transposition of the Cit+ determinant. This was thought to be due to the differential transposability of IS3411L and IS3411R in the transposition process. The frequency of transposition of IS3411 marked with a chloramphenicol resistance determinant was much higher than IS3411-mediated cointegrate formation, suggesting that replicon fusions are not essential intermediates in the transposition process of Tn3411 or IS3411. Spontaneous deletions occurred with high frequency in recA hosts. The spontaneous deletion promoted by homologous recombination between two IS3411 elements in Tn3411 was examined with deletion mutants.

Chromosome Deletion↗

PubMed2003

Involvement of the adc operon and manganese homeostasis in Streptococcus gordonii biofilm formation.

Pioneer oral bacteria, including Streptococcus gordonii, initiate the formation of oral biofilms on tooth surfaces, which requires differential expression of genes that recognize unique environmental cues. An S. gordonii::Tn917-lac biofilm-defective mutant was isolated by using an in vitro biofilm formation assay. Subsequent inverse PCR and sequence analyses identified the transposon insertion to be near the 3' end of an open reading frame (ORF) encoding a protein homologous to a Streptococcus pneumoniae repressor, AdcR. The S. gordonii adc operon, consisting of the four ORFs adcR, adcC, adcB, and adcA, is homologous to the adc operon of S. pneumoniae, which plays a role in zinc and/or manganese transport and genetic competence in S. pneumoniae. AdcR is a metal-dependent repressor protein containing a putative metal-binding site, AdcC contains a consensus-binding site for ATP, AdcB is a hydrophobic protein with seven hydrophobic membrane-spanning regions, and AdcA is a lipoprotein permease with a putative metal-binding site. The three proteins (AdcC through -A) are similar to those of the binding-lipoprotein-dependent transport system of gram-positive bacteria. Reverse transcriptase PCR confirmed that adcRCBA are cotranscribed as an operon in S. gordonii and that the transposon insertion in S. gordonii adcR::Tn917-lac had resulted in a polar mutation. Expression of adcR, measured by the beta-galactosidase activity of the adcR::Tn917-lac mutant, was growth phase dependent and increased when the mutant was grown in media with high levels of manganese (>1 mM) and to a lesser extent in media with zinc, indicating that AdcR may be a regulator at high levels of extracellular manganese. A nonpolar inactivation of adcR generated by allelic replacement resulted in a biofilm- and competence-defective phenotype. The biofilm-defective phenotype observed suggests that AdcR is an active repressor when synthesized and acts at a distant site(s) on the chromosome. Thus, the adc operon is involved in manganese acquisition in S. gordonii and manganese homeostasis and appears to modulate sessile growth in this bacterium.

ATP-Binding Cassette Transporters↗

PubMed2003

Application of chimeric feline foamy virus-based retroviral vectors for the induction of antiviral immunity in cats.

In order to define the potential and applicability of replication-competent foamy virus-based vaccine vectors, recombinant feline foamy virus (FFV) vectors encoding defined segments of the feline calicivirus (FCV) capsid protein E domain were constructed. In cell cultures, these FFV-FCV vectors efficiently transduced and expressed a hybrid fusion protein consisting of the essential FFV Bet protein and the attached FCV E domains. The stability of the vectors in vitro was inversely correlated to the size of the heterologous insert. The deletion of a part of the FFV U3 sequence in these FFV-FCV vectors did not interfere with replication and titer in cell cultures but increased the genetic stability of the hybrid vectors. Selected chimeric vectors were injected into immunocompetent cats and persisted in the transduced host concomitant with a strong and specific humoral immune response against vector components. In a substantial number of cats, antibodies directed against the FCV E domain were induced by the FFV-FCV vectors, but no FCV-neutralizing activities were detectable in vitro. When the vaccinated cats were challenged with a high-titer FCV dose, sterile immunity was not induced by any of the hybrid FFV-FCV vectors. However, the FFV-FCV vector with a truncated U3 region of the long terminal repeat promoter significantly reduced the duration of FCV shedding after challenge and suppressed the appearance of FCV-specific ulcers. Possible mechanisms contributing to the partial protection will be discussed.

Amino Acid Sequence↗

PubMed2003

COP9 signalosome subunit 3 is essential for maintenance of cell proliferation in the mouse embryonic epiblast.

Csn3 (Cops3) maps to the mouse chromosome 11 region syntenic to the common deletion interval for the Smith-Magenis syndrome, a contiguous gene deletion syndrome. It encodes the third subunit of an eight-subunit protein complex, the COP9 signalosome (CSN), which controls a wide variety of molecules of different functions. Mutants of this complex caused lethality at early development of both plants and Drosophila melanogaster. CSN function in vivo in mammals is unknown. We disrupted the murine Csn3 gene in three independent ways with insertional vectors, including constructing a approximately 3-Mb inversion chromosome. The heterozygous mice appeared normal, although the protein level was reduced. Csn3(-/-) embryos arrested after 5.5 days postcoitum (dpc) and resorbed by 8.5 dpc. Mutant embryos form an abnormal egg cylinder which does not gastrulate. They have reduced numbers of epiblast cells, mainly due to increased cell death. In the Csn3(-/-) mice, subunit 8 of the COP9 complex was not detected by immunohistochemical techniques, suggesting that the absence of Csn3 may disrupt the entire COP9 complex. Therefore, Csn3 is important for maintaining the integrity of the COP9 signalosome and is crucial to maintain the survival of epiblast cells and thus the development of the postimplantation embryo in mice.

Animals↗

PubMed1994

A single up-stream element confers responsiveness to 1,25-dihydroxyvitamin D3 and tumor necrosis factor-alpha in the rat osteocalcin gene.

Tumor necrosis factor-alpha (TNF alpha) is one of several autocrine/paracrine factors known to exert potent inhibitory effects on bone. We have shown that TNF alpha inhibition of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]-stimulated synthesis of the bone-specific protein osteocalcin (OC) occurs by decreasing steady state levels of OC mRNA, suggesting a pretranslational mechanism. In many genes, TNF alpha action is mediated by the transcription factor NF kappa B. Analysis of OC 5'-flanking DNA revealed a sequence structurally homologous to the previously described NF kappa B-binding site and, thus, a potential TNF alpha response element. Deletion analysis was performed to identify the sequences mediating the response to TNF alpha in osteoblastic ROS 17/2.8 cells by transient transfection with reporter constructs containing rat OC 5'-flanking DNA [chloramphenicol acetyltransferase (CAT)] that retained or deleted homologous NF kappa B sites or a previously defined 1,25-(OH)2D3 response element (VDRE). Transfection with all reporter constructs resulted in low basal CAT activity, measured 72 h after transfection. 1,25-(OH)2D3 stimulated CAT activity 2.8- to 4.5-fold in cells transfected with constructs that included the VDRE. TNF alpha inhibited 1,25-(OH)2D3-stimulated, but not basal, CAT activity. Deletion analysis localized the effect of TNF alpha to a sequence between -522 and -306 relative to the OC transcription start site, an area that included the VDRE but deleted a homologous NF kappa B element. Transfection of cells with a heterologous reporter containing one copy of the OC VDRE inserted in correct orientation or two copies in inverse orientation was sufficient to confer a response to TNF alpha. Gel mobility shift analysis of DNA-nuclear protein interaction revealed that 1,25-(OH)2D3 stimulated an increase in binding of nuclear proteins to an OC 32P-VDRE probe. Preincubation of nuclear extract with specific monoclonal antibodies confirmed that the proteins binding the VDRE included the vitamin D receptor and retinoid-X receptor. TNF alpha treatment of cells inhibited the 1,25-(OH)2D3-stimulated increase in nuclear protein binding to the VDRE. These results suggest 1) the VDRE is sufficient to confer a response to the inhibitory effect of TNF alpha on 1,25-(OH)2D3-stimulated rat OC gene transcription; 2) the action of TNF alpha does not require homologous NF kappa B response elements; and 3) the mechanism of TNF alpha inhibition of 1,25-(OH)2D3-stimulated OC gene expression includes modulation of binding of the vitamin D receptor/retinoid-X receptor heterodimer to the VDRE.

Animals↗

PubMed1996

Direct noninvasive measurement of tear oxygen tension beneath gas-permeable contact lenses in rabbits.

PURPOSE: To develop a noninvasive measurement of tear oxygen tension and to determine tear oxygen tension and calculated oxygen flux rates during wear of contact lenses of varying oxygen transmissibility (Dk/L). METHODS: The basis of the tear oxygen tension (PO2) measurement is phosphorescence quenching of Pd-meso-tetra-(4-carboxyphenyl) porphine by oxygen. A fiber-optic bundle, positioned approximately 2 mm from a cuvette or from the corneal surface, delivers an excitation flash (< 4 microseconds, 539 +/- 23 nm) and collects the phosphorescence emission (> 645 nm) of the dye at 1 MHz for 2 ms. Porphine (500 microM) + 1% albumin is instilled directly into the tears of a sedated rabbit, and phosphorescence is measured before and after contact lens insertion. RESULTS: Phosphorescence intensity decay lifetimes are related inversely to PO2 and follow the Stern-Volmer relationship. Calibration control experiments revealed a temperature effect on lifetime of 1% per degree, dye concentration independence over a 100-fold dilution, and no significant pH effect between 6.8 and 7.9. Phosphorescence lifetimes indicated that tear PO2 beneath contact lenses reached steady state within 3 to 8 minutes. Mean steady state tear PO2 (mm Hg +/- SE; N > 7) beneath contact lenses was: polymethylmethacrylate (Dk/L = 0), tear PO2 = 0.58 +/- 0.22; Oxyflow f30 (Dk/L = 15.3), tear PO2 = 27.7 +/- 3.4; Fluorex 700 Dk/L = 30), tear PO2 = 52.5 +/- 3.8; Oxyflow 151 (Dk/L = 51.9), tear PO2 = 78.7 +/- 23.9. Steady state tear PO2 beneath an Oxyflow f30 after application of topical anesthetic and 10 minutes of lens wear was 41 +/- 1.2 mm Hg, significantly higher than without anesthetic. CONCLUSIONS: Phosphorescence-based measurements allow the direct, sensitive, and noninvasive assessment of oxygen availability beneath a contact lens. Oxygen levels beneath permeable contact lenses found by direct measurement are significantly lower than what mathematical models predict. The measured values, however, are close to PO2 estimates using the equivalent oxygen percentage (EOP) technique. Tear PO2 measurements may be used to examine metabolic disturbances as shown by reduced oxygen consumption rates after application of topical anesthetic.

Anesthetics, Local↗

PubMed1998

The snapped inferior rectus.

PURPOSE: To evaluate the management and possible aetiology of the snapped inferior rectus muscle in strabismus surgery. METHODS: Three patients are described whose inferior rectus muscle broke across its width some 8-10mm behind the insertion while being held on a squint hook without excessive force during strabismus surgery. The proximal part of the muscle was not found. The distal part of the snapped muscle was excised for pathological examination. Transposition of the inferior halves of the adjacent horizontal muscles to the insertion of the inferior rectus (a modified inverse-Knapp procedure) was performed in all cases. RESULTS: After the transposition surgery, one patient was orthotropic in the primary position, one patient required a prism correction to produce a range of single vision and the third patient was orthotropic after a further operation. There was good depression in one case and the other two had a limitation of depression. In all cases, horizontal movements remained intact and there were no signs of anterior segment ischaemia. CONCLUSIONS: The unique relations of the inferior rectus to the surrounding tissues may be a factor in causing the breaking of this muscle. Two of the patients were elderly and this may be a factor also. Transposition surgery is the appropriate management when the proximal part of the snapped muscle cannot be located and has satisfactory but imperfect results.

Adult↗

PubMed1994

Transcription activation mediated by chromosomal inversion in rat cells.

To investigate the relationship between the local configuration of a gene and its level of expression, we constructed a rat cell line, Hy5, carrying a mutant polyomavirus middle T oncogene (pmt) whose overexpression converted the cells to the transformed state. The structure of the transgene was such that pmt was able to undergo chromosomal inversion at a relatively high rate by a cross-over in flanking pBR322 sequences. Hy5 cells became spontaneously transformed at a rate of 10(-5) per cell generation and all of the transformants analysed had sustained pmt inversion. CpG sequences were partially methylated in the Hy5 insert but appeared demethylated in transformants. In two subclones derived from untransformed Hy5 cells, the pmt insert was densely methylated, transcriptionally inactive and unable to undergo homologous recombination. Our results suggest that DNA repair associated with recombinational events leads to a heritable hypomethylation of the locus which is responsible for its activation.

Animals↗

PubMed1997

Analysis of DNA inversions in the shufflon of plasmid R64.

The shufflon, a multiple DNA inversion system in the plasmid R64, consists of four DNA segments flanked and separated by seven 19-bp repeat sequences. Site-specific recombinations mediated by the rci product occur between each inverted repeat sequence, resulting in inversions of the four segments independently or in groups. The seven 19-bp repeat sequences are classified into four types (repeat-a, -b, -c, and -d), according to their 3-bp variable sequences. We individually cloned A, B, and C segments of the R64 shufflon and determined the in vivo inversion frequency of each segment. The inversion frequencies of three segments differed greatly. The inversion frequency declined in the following order: segments A, B, and C. Synthetic 19-mer oligonucleotides corresponding to both strands of repeat-a, -b, -c, and -d sequences were inserted into appropriate sites of pBR322. The rci-mediated DNA inversion occurred between two synthetic inverted repeats, indicating that the 19-bp inverted repeat sequences are the sole elements required in cis for the shufflon system. The inversion frequencies of DNA segments flanked by various sequences indicate that the four types of repeat sequences determine the inversion frequency of the four DNA segments of the R64 shufflon. Deletion of a DNA segment flanked by direct repeat sequences could not be detected.

DNA Nucleotidyltransferases↗

PubMed1993

Vaccinia virus recombinants co-expressing hepatitis B virus surface and core antigens.

Using the Praha strain of vaccinia virus (VV) two double recombinant VVs expressing the surface and capsid HBV proteins (HBsAg and HBcAg) under the control of the P7.5 promoter were constructed. In the first construct the gene coding for HBsAg was inserted into the HindIII J fragment (TK gene) and the gene coding for HBcAg was inserted into the HindIII M fragment (host range, K1L gene) of the VV genome. To test whether the expression of the foreign genes was influenced by the insertion site, in the second construct their locations were inversely changed. When compared with single VV-HBV recombinants expressing either HBsAg or HBcAg, the double recombinants expressed in vitro approximately the same amounts of the respective antigens. The particles formed by either HBsAg or HBcAg expressed by recombinant viruses, were isolated and examined by electron microscopy. Particles composed of both HBsAg and HBcAg were not detected in cultures infected with one of the double recombinants. The residual virulence in 3-week-old mice of the single recombinants was not markedly altered by the insertion of the second gene. The immunogenicity in mice of both the single and double recombinants was comparable and was not influenced by the location of the HBV genes in VV genome, as revealed by antibodies developed against the respective antigens.

Animals↗

PubMed1993

Molecular analysis of T-cell receptor transcripts in a human T-cell leukemia bearing a t(1;14) and an inv(7); cell surface expression of a TCR-beta chain in the absence of alpha chain.

The polymerase chain reaction (PCR) was used to amplify cDNA in order to characterize normal and hybrid T-cell receptor (TCR) gene rearrangements derived from a T-cell acute lymphoblastic leukemia (T-ALL) bearing a chromosome 7 inversion. The nucleotide sequence analysis of the amplified product showed the presence of an out-of-frame V beta/J gamma/C gamma transcript and an in-frame V gamma/J gamma/C beta transcript which result from an interlocus recombination between the TCR-beta and gamma loci and the transcription of the reciprocal hybrid TCR gene. The sequence analysis of the reciprocal DNA segment directly involved in the breakpoint of the inversion showed a recombination between a J gamma-sequence heptamer signal and a coding J beta gene segment. The exonuclease nibbling of each DNA extremity and the non-templated nucleotide insertion observed at both coding and reciprocal joints demonstrate that the inversion is mediated by the lymphocyte recombinase complex. During T-cell differentiation, TCR-beta genes are rearranged prior to TCR-alpha genes. In the present case of T-ALL, we have shown that TCR-delta genes are rearranged at both loci, excluding productive rearrangements of TCR-alpha genes. The molecular analysis of the normal TCR genes derived from the leukemic cells revealed the presence of productively rearranged TCR-beta, gamma, and delta genes. Cell surface staining of these cells showed the presence of CD3 molecules and of a TCR-beta chain corresponding to the normal beta allele expressed in a disulfide-linked complex with a protein which is not TCR-alpha, gamma, or delta. This could represent the cell surface expression of the hybrid TCR-V gamma/C beta protein or the expression of a TCR-beta homodimer.

Adult↗

PubMed1980

DNA rearrangements associated with a transposable element in yeast.

The his4-912 mutation results from insertion of a 6200 bp transposable element into the his4 gene of yeast. In order to clone the his4-912 mutation, the plasmid pBR322 was integrated into the his4 gene by means of yeast transformation, and then the vector sequences and the his4-912 insertion element were excised as a single restriction fragment. This his 4-912 insertion element is homologous to Ty1, a family of repetitive yeast DNA sequences. His+ revertants derived from the his4-912 mutant carry a number of chromosomal aberrations including deletions, translocations, a transposition and an inversion. The majority of His+ revertants result from deletions which have both endpoints within the element and which leave behind only 300 bp of the insertion element. Other derivatives of the his4-912 mutant carry deletions which have one endpoint in the insertion element and one endpoint in the his4 coding sequence. In two His+ revertants carrying reciprocal translocations, the chromosome III translocation breakpoints occur within the his4-912 insertion element. A His+ revertant carrying an inversion of most of the left arm of chromosome III may be an intermediate in transposition of the his4-912 insertion element to a new site on chromosome III.

Base Sequence↗

PubMed2002

Functional genomics in the mouse.

The mouse is the premier genetic model organism for the study of human disease and development. With the recent advances in sequencing of the human and mouse genomes, there is strong interest now in large-scale approaches to decipher the function of mouse genes using various mutagenesis technologies. This review discusses what tools are currently available for manipulating and mutagenizing the mouse genome, such as ethylnitrosourea and gene trap mutagenesis, engineered inversions and deletions using the cre-lox system, and proviral insertional mutagenesis in somatic cells, and how these are being used to uncover gene function.

Alkylating Agents↗

PubMed1998

Testing the charge difference hypothesis for the assembly of a eucaryotic multispanning membrane protein.

The Glut1 glucose transporter is a glycoprotein whose membrane topology has been verified by a number of experimental observations, all of which are consistent with a 12-transmembrane helix model originally based on hydrophobicity analysis. We used Glut1 as a model multispanning membrane protein to test the Charge Difference Hypothesis (Hartmann, E., Rapoport, T. A., and Lodish, H. F. (1989) Proc. Natl. Acad. Sci. U. S. A. 86, 5786-5790), which asserts that the topology of a eucaryotic multispanning membrane protein is determined solely by the amino acid charge difference across the first transmembrane segment. The charge difference across the first transmembrane segment of Glut1 was progressively inverted in two independent series of mutants, one series in which only the number of positively charged amino acid residues in the two flanking domains was altered and the other in which only the number of negatively charged residues in the two flanking domains was changed. The results indicate that the charge difference across the first transmembrane segment does affect the topology of the protein, but that contrary to the hypothesis, it only dictates the orientation of the first transmembrane segment and the disposition of the amino terminus and the first linker domain. Charge inversion resulted in the formation of aberrant molecules in which either the first or second transmembrane segment failed to insert into the membrane. The topology of downstream regions of Glut1 was unaffected by charge inversion across the first transmembrane segment, indicating that downstream sequences are important in determining the local topological disposition of the molecule.

Amino Acid Sequence↗

PubMed2005

Investigations of the effect of DNA size in transient transfection assay using dual luciferase system.

In transient transfection assays deletions, insertions, or truncations are broadly used to define cis regulatory elements, such as promoters, enhancers, and silencers. This application is based on the assumption that size changes of a test fragment have little or no effect on the apparent activity of the reporter genes. While it is known that unusually large size of a construct, such as PAC DNA, substantially reduces transfection efficiency, the size effect of a DNA fragment ranging from a couple of hundred basepairs to a few thousand basepairs, within which most transient transfection assays are performed, has not been rigorously investigated. Here we report a systematic study on the effect of plasmid size on the measurement of promoter/enhancer activity in transient transfection assay based on the dual luciferase reporter system. By inserting various lengths of stuffer DNA into the luciferase parental plasmids, we found an inverse correlation between the relative luciferase activity and the construct size. Furthermore, this size effect can be only partially corrected by inserting the same size of stuffer DNA into both experimental and control plasmids. The results suggest that the effect of DNA size should be taken into consideration for evaluation of transcriptional elements using transient transfection analysis.

DNA↗

PubMed1997

Nonrandom location of IS1 elements in the genomes of natural isolates of Escherichia coli.

We have studied the spatial distribution of IS1 elements in the genomes of natural isolates comprising the ECOR reference collection of Escherichia coli. We find evidence for nonrandomness at three levels. Many pairs of IS1 elements are in much closer proximity (< 10 kb) than can be accounted for by chance. IS1 elements in close proximity were identified by long-range PCR amplification of the genomic sequence between them. Each amplified region was sequenced and its map location determined by database screening of DNA hybridization. Among the ECOR strains with at least two IS1 elements, 54% had one or more pairs of elements separated by < 10 kb. We propose that this type of clustering is a result of "local hopping," in which we assume that a significant proportion of tranposition events leads to the insertion of a daughter IS element in the vicinity of the parental element. A second level of nonrandomness is found in strains with a modest number of IS1 elements that are mapped through the use of inverse PCR to amplify flanking genomic sequences: in these strains, the insertion sites tend to be clustered over a smaller region of chromosome than would be expected by chance. A third level of nonrandomness is observed in the composite distribution of IS elements across strains: among 20 mapped IS1 elements, none were found in the region of 48-77 minutes, a significant gap. One region of the E. coli chromosome, at 98 min, had a cluster of IS1 elements in seven ECOR strains of diverse phylogenetic origin. We deduce from sequence analysis that this pattern of distribution is a result of initial insertion in the most recent common ancestor of these strains and therefore not a hot spot of insertion. Analysis using long-range PCR with primers for IS2 and IS3 also yielded pairs of elements in close proximity, suggesting that these elements may also occasionally transpose by local hopping.

Chromosome Mapping↗

PubMed1994

Salmonella typhimurium loci involved in survival within macrophages.

A set of Tn10 mutants of Salmonella typhimurium which have a diminished capacity to survive in murine macrophages and decreased virulence in mice has been described previously. In this study, we characterized 30 of these mutants and determined map locations of Tn10 insertions for 23 of these strains. In addition, short fragments of transposon-flanking DNA were cloned, and the nucleotide sequence was determined for 23 mutants. Seven mutants carried transposon insertions in known genes, representing six loci: htrA, prc, purD, fliD, nagA, and smpB. The possible roles of these genes in Salmonella virulence are discussed. One insertion was found to be in an unknown gene which shared homology with the open reading frames Bv' and Bv located in the pin inversion system of Shigella boydii. In one mutant, Tn10 was found to be inserted in a gene with significant homology to adhE of Escherichia coli and Clostridium acetobutylicum. The map location and degree of homology indicate that the Salmonella gene encodes a related, but different, dehydrogenase. In 14 of the mutants analyzed, Tn10 was inserted into genes which had no significant homologies to entries in the DNA and protein data bases. In conclusion, 16 insertions define loci, termed ims for impaired macrophage survival, which have not yet been described in S. typhimurium but have been shown previously to be necessary for full virulence in mice. Although most ims loci are distributed randomly throughout the genome, a cluster was found between 75 and 78 min on the Salmonella chromosome.

Amino Acid Sequence↗