Regulation of resistance to leprosy by chromosome 1 locus in the mouse

Mice of different inbred strains vary in their resistance to intravenous infection with Mycobacterium lepraemurium (MLM). The mean survival time of MLM-infected A/J and DBA/2 mice is significantly longer than that of similarly infected C57BL/6 and BALB/c mice. The typing of AXB/BXA recombinant inbred strains (A = A/J, B = C57BL/6) for the trait of relative resistance/susceptibility to MLM revealed a perfect match with the strain distribution pattern of resistance/susceptibility to Mycobacterium bovis (BCG), the trait which is controlled by the Bcg (Ity, Lsh) locus on chromosome 1. The control, by this gene, of response to MLM was further confirmed by the demonstration that BALB/c-Bcg ^r congenic mice,which carry the DBA/2-derived Bcg ^r (resistant) allele on chromosome 1, are significantly more resistant to MLM infection than their BALB/c (Bcg ^s , susceptible) counterparts.     

Circular dichroism studies on α- and β-ketosides of 5-acetamido-3,5-dideoxy- -glycero- -galacto-nonulopyranosonic acid (N-acetylneuraminic acid) and of some of its derivatives

The circular dichroism spectra of a number of N-acetylneuraminic acid derivatives in aqueous solution were studied. For all compounds, the Cotton effects were found to be in the spectral range of the acetamido and carboxyl chromophores. The c.d. curves of the methyl, ethyl, and allyl α- -ketosides are characterized by a broad, positive band centered at λ ≈ 195 nm with a slight skew towards the higher wavelengths and weak bands between λ 225 and 255 nm, whereas the methyl β- -ketoside and the corresponding methyl ester show only an intense positive band with a broad shoulder in the same spectral range. 5-Acetamido-3,5-dideoxy- -glycero-β- -galacto-nonulopyranose, its methyl β- -ketoside, and 5-acetamido-3,5-dideoxy- -glycero- -galacto-nonulopyranosonamide containing only the acetamido chromophore showed one single positive Cotton effect centered at λ ≈ 192 nm. The c.d. spectrum of 5-acetamido-3,5-dideoxy- -glycero- -galacto-nonulopyranosonic acid confirms the β- configuration of the free acid in aqueous solution, whereas the shape of the c.d. curve of O-(N-acetyl-α- -neuraminopyranosyl)-(2→3)-O-β- -galactopyranosyl-(1→4)- -glucopyranose resembles that of the methyl, ethyl, and allyl α- -ketosides 2-4.     

High-resolution linkage map in the vicinity of the Lp locus

Looptail (Lp) is a mutation that profoundly affects neurulation in mouse and is characterized by craniorachischisis, an open neural tube extending from the midbrain to the tail in embryos homozygous for the mutation. Lp maps to the distal portion of mouse chromosome 1, and as part of a positional cloning approach, we have generated a high-resolution linkage map of the Lp chromosomal region. For this, we have carried out extensive segregation analysis in a total of 706 backcross mice informative for Lp and derived from two crosses, (Lp/ + X SJL/J)F1 X SJL/J and (Lp/ + X SWR/J)F1 X SWR/J. In addition, 269 mice from a (Mus spretus X C57BL/6J)F1 X C57BL/6J interspecific backcross were also used to order marker loci and calculate intergene distances for this region. With these mice, a total of 28 DNA markers corresponding to either cloned genes or anonymous markers of the SSLP or SSCP-types were mapped within a 5-cM interval overlapping the Lp region, with the following locus order and interlocus distances (in cM): centromere-D1Mit110 / Atp1β1 / Cd3ζ / Cd3η / D1Mit145 — D1Hun14 / D1Mit15 — D1Mit111 / D1Mit112 — D1Mit114 — D1Mit148 / D1Mit205/ D1Mit36 / D1Mit146 / D1Mit147 / D1Mit270 / D1Hun13 — Fcgr2 — Mpp — Apoa2/Fcer1γ - Lp - D1Mit149 / Spna1/Fcer1α-Eph1-Hlx1/D1Mit62. These studies have allowed the delineation of a maximum genetic interval for Lp of 0.5 cM, a size amenable to physical mapping techniques.     

Physical and Genetic Maps of thedeafwaddlerRegion on Distal Mouse Chr 6

Thedeafwaddler(dfw) mutation, displaying motor ataxia and profound deafness, arose spontaneously in a C3H/HeJ colony and was mapped previously to distal mouse Chr 6. In this study, a high-resolution genetic map was generated by positioning 10 microsatellite markers and 5 known genes on a 968-meioses intersubspecific backcross segregating fordfw[(CAST/Ei–+/+ × C3HeB/FeJ–dfw/dfw) × C3HeB/FeJ–dfw/dfw], giving the following marker order and sex-averaged distances:D6Mit64–(0.10 + 0.10 cM)–Pang–(1.24 + 0.36 cM)–Itpr1–(0.62 + 0.25 cM)–D6Mit108–(0.52 + 0.23 cM)–D6Mit54–(0.21 + 0.15 cM)–D6Mit23, D6Mit107, D6Mit328–(0.72 + 0.27 cM)–D6Mit11–(0.21 + 0.15 cM)–dfw–(0.93 + 0.31 cM)–Gat4, D6Mit55–(0.10 + 0.10 cM)–D6Mit63–(0.31 + 0.18 cM)–Syn2–(0.62 + 0.25 cM)–D6Mit44(Rho). Female and male genetic maps are similar immediately surrounding thedfwlocus, but show marked differences in other areas. A yeast artificial chromosome-based physical map suggests that the closest markers flanking thedfwlocus,D6Mit11(proximal) andGat4, D6Mit55(distal), are contained within 650–950 kb. The human homologues of the flanking lociItpr1(proximal) andSyn2(distal) map to chromosome 3p25–p26, suggesting that the human homologue of thedfwgene is located within this same region.     

Location of the 9257 and ataxia mutations on mouse Chromosome 18

The location of three mutations on proximal Chromosome (Chr) 18 was determined by analysis of the offspring of several backcrosses. The results demonstrate that ataxia and the insertional mutation TgN9257Mm are separated by less than 1 cM and are located approximately 3 cM from the centromere, while the balding locus is 7 cM more distal. Previous data demonstrated that the twirler locus also maps within 1 cM of ataxia. The corrected locations will contribute to identification of appropriate candidate genes for these mutations. Two polymorphic microsatellite markers for proximal Chr 18 are described, D18Umi1 and D18Umi2. The Lama3 locus encoding the α3 subunit of nicein was mapped distal to ataxia and did not recombine with Tg9257. Received: 19 December 1995 / Accepted: 29 January 1996     

Lupane-triterpene glycosides from leaves of Acanthopanax koreanum

Two new lupane-triterpene glycosides named acankoreosides C and D, were isolated from the leaves of Acanthopanax koreanum. Based on spectroscopic data, the chemical structures were determined as 3-O-β- -glucopyranosyl 3α,11α-dihydroxylup-20(29)-en-28-oic acid 28-O-α- -rhamnopyranosyl-(1→4)-β- -glucopyranosyl-(1→6)-β- -glucopyranosyl ester and 3α,11α-dihydroxylup-23-al-20(29)-en-28-oic acid 28-O-α- -rhamnopyranosyl-(1→4)-β- -glucopyranosyl-(1→6)-β- -glucopyranosyl ester, respectively.     

High-Resolution Linkage Map of Mouse Chromosome 13 in the Vicinity of the Host Resistance LocusLgn1

Natural resistance of inbred mouse strains to infection withLegionella pneumophilais controlled by the expression of a single dominant gene on chromosome 13, designatedLgn1.The genetic difference atLgn1is phenotypically expressed as the presence or absence of intracellular replication ofL. pneumophilain host macrophages. In our effort to identify theLgn1gene by positional cloning, we have generated a high-resolution linkage map of theLgn1chromosomal region. For this, we have carried out extensive segregation analysis in a total of 1270 (A/J × C57BL/6J) × A/J informative backcross mice segregating the resistance allele of C57BL/6J and the susceptibility allele of A/J. Additional segregation analyses were carried out in three preexisting panels of C57BL/6J ×Mus spretusinterspecific backcross mice. A total of 39 DNA markers were mapped within an interval of approximately 30 cM overlapping theLgn1region. Combined pedigree analyses for the 5.4-cM segment overlappingLgn1indicated the locus order and the interlocus distances (in cM):D13Mit128–(1.4)–D13Mit194–(0.1)–D13Mit147–(0.9)–D13Mit36–(0.9)–D13Mit146–(0.2)–Lgn1/D13Mit37–(1.0)–D13Mit70.Additional genetic linkage studies of markers not informative in the A/J × C57BL/6J cross positionedD13Mit30, -72, -195,and-203, D13Gor4, D13Hun35,andMtap5in the immediate vicinity of theLgn1locus. The marker density and resolution of this genetic linkage map should allow the construction of a physical map of the region and the isolation of YAC clones overlapping the gene.     

The ter primordial germ cell deficiency mutation maps near Grl-1 on mouse Chromosome 18

A single recessive gene, ter (teratoma), causes germ cell deficiency and a high incidence of congenital testicular teratomas in the 129/Sv-ter strain of the mouse. Linkage analyses between the ter gene and 36 marker genes of 19 chromosomes were performed with matings between the C57BL/6J-ter congenic strain and four inbred strains. Results showed that the ter gene was linked to D18Mit9, D18Mit14, and D18Mit17 on Chromosome (Chr) 18. Gene order estimated on the basis of recombination distance (in centimorgans) was [centromere-D18Mit14-5.1 (cM)-ter-0 (cM)-D18Mit17-23.8 (cM)-D18Mit9]. D18Mit17 is the microsatellite DNA of the Grl-1 (glucocorticoid receptor-1) locus. We conclude that the ter gene is closely linked to Grl-1 on Chr 18 and is a new mutation involving the developmental modification of primordial germ cells in mice.Deceased     

Host interference with expression of the lambda N gene product

We have searched among E. coli M72 (D, bio11cI857H1) temperature resistant survivors and have found two bacterial mutants, gro100 and gro101 which block λiλ and λi⁴³⁴ phage development but allow growth of their N-independent derivatives λiλ nin and λi⁴³⁴nin. It is not known yet whether these two mutants interfere with the production of the N gene product or with its function. At least part of the gro genotype maps at 12′ of the E. coli genetic map and is co-transductible by Pl with the lac locus.     

Glycosylated nervogenic acid derivatives from Liparis condylobulbon (Reichb.f.) leaves

Three new nervogenic acid glycosides, 1-O-α-l-rhamnopyranosyl 3,5-bis(3-methyl-but-2-enyl)-4-O-[α-l-rhamnopyranosyl-(1→2)-β-d-glucopyranosyl]-benzoate, 3,5-bis(3-methyl-but-2-enyl)-4-O-[α-l-rhamnopyranosyl-(1→2)-β-d-glucopyranosyl]-benzoic acid, and bis{3,5-bis(3-methyl-but-2-enyl)-4-O-[α-l-rhamnopyranosyl-(1→2)-β-d-glucopyranosyl]-benzoyl} 1,2-O-β-d-glucopyranose, which we named condobulbosides A–C, were isolated from a methanol extract of the leaves of Liparis condylobulbon together with an apigenin C-glycoside, schaftoside. Their structures were established on the basis of spectral techniques, namely, UV, IR, HR-MS spectroscopy, both 1D and 2D NMR experiments, and chemical reactions.     

Protein synthesis induced by infection with packaged λdv plasmid

Plasmid λdv or imm²¹dv DNA was joined to a λ arm having a cos site. This recombinant plasmid can be packaged in a λ head, and used to infect Escherichia coli K12 cells. The injected DNA molecules become plasmids in cells. By adding these particles to uv-irradiated uvrA cells, the packageable λdv or imm²¹dv plasmids can be induced to synthesize proteins coded by genes on the plasmid genome. The packageable plasmid system is thus suitable for studying on synthesis and regulation of plasmid-coded biopolymers. Analyses of the dv-coded proteins in gel electrophoreses revealed that among several genes carried on the dv plasmid genome, only those genes that are members of the pRoR-tof-cII-O-P operon can be expressed. Evidence has been presented to show that expression of this operon, which is directly correlated with replication of the genome, is only partially allowed in cells perpetuating the dv plasmid. These observations are discussed in connection with the autorepressor model (D. E. Berg, 1974, Virology 62, 224–233; K. Matsubara, 1976, J. Mol. Biol. 102, 427–439) that genetically accounts for the control mechanism of plasmid replication.     

The Mouse Severe Combined Immune Deficiency (scid) Mutation Is Closely Linked to the B-Cell-Specific Developmental Genes VpreB and λ5

The mouse severe combined immune deficiency (scid) phenotype is due to a recessive, autosomal mutation which results in failed development of lymphocytes. An important step during normal lymphocyte development is the germline rearrangement of DNA segments to assemble functional immunoglobulin or T cell receptor genes. scid lymphocytes fail to rearrange these genes properly, resulting in the absence of mature B and T lymphocytes. This mutation was originally mapped to chromosome 16 by linkage to the immunoglobulin λ light chain genes (Igl-1) and the coat color mutation mahoganoid . We have typed 288 progeny from backcrosses between MOLF/Ei or CAST/Ei and C.B-17-scid for the scid phenotype and nine other loci mapped to the centromeric region of MMU16. We have established a refined map of this region which places the scid gene between Prm-2 and Igl-1 . In addition, no recombinations were found between scid and three other loci, VpreB, λ5, and D16Mit31, providing markers useful for isolating the scid gene by positional cloning.     

Gentisic acid conjugates of Medicago truncatula roots

Three phenolic glycosides 5-O-{[5′′-O-E-(4′′′-O-threo-guaiacylglycerol)-feruloyl]-β-apiofuranosyl-(1→2)-β-xylopyranosyl} gentisic acid, 5-O-[(5′′-O-vanilloyl)-β-apiofuranosyl-(1→2)-β-xylopyranosyl] gentisic acid and 1-O-[E-(4′′′-O-threo-guaiacylglycerol)-feruloyl]-3-O-β-galacturonopyranosyl glycerol were isolated and identified from the roots of Medicago truncatula together with four known 5-O-β-xylopyranosyl gentisic acid, vicenin-2, hovetrichoside C and pterosupin identified for the first time in this species. Structural elucidation was carried out on the basis of UV, mass, ¹H and ¹³C NMR spectral data.     

Genetic mapping of the athymic nude (RNU) locus in the rat to a region on Chromosome 10

The nude trait in the rat is transmitted in an autosomal recessive manner and is associated with thymic aplasia, T-cell deficiency, and hairlessness. Congenic rats homozygous for the RNU (Rowett nude) locus are important models in the study of inflammatory disease, tumor growth, and transplant rejection. The RNU locus has not been previously mapped, and the nature of the gene product is unknown. To determine the map location of this gene, a single F344.rnu/rnu (athymic nude congenic Fischer rat) male congenic rat was bred with 3 LEW/N (NIH stock Lewis rat) female rats to produce F₁ progeny. Twelve F₁ brother-sister breeding pairs were established. Forty-nine phenotypically nude F₂ offspring (198 total) were obtained. Linkage analysis done on F₂ DNA revealed highly significant cosegregation between the nude phenotype and eight polymorphic markers located on Chromosome (Chr) 10. The tightest linkages were with: MYH3 (embryonic, skeletal myosin heavy chain) and SHBG (sex hormone-binding globulin), giving 2 point lod scores of 20.2, and 20.0, respectively. The map order and map distances, determined by multipoint linkage calculations, were: RR24-(16.1 cM)-MYH3-(3.5 cM)-SHBG-(4.7 cM)-RNU-(11.9 cM)-F16F2-(24.1 cM)-CLATP (citrate lyase ATPase)-(2.4 cM)-ACE (angiotensin converting enzyme)/PPY (pancreatic polypeptide)-(14.1 cM)-RR1023. The position of the RNU locus in the rat corresponds closely with that of the recently reported nu locus in the mouse. This finding suggests that the nude phenotype in the rat and the mouse arise from defects in homologous genes.     

Natural resistance to infection with Legionella pneumophila: chromosomal localization of the Lgn1 susceptibility gene

Legionella pneumophila is a strict intracellular pathogen that replicates in the professional phagocytes of the human and guinea pig host. Although murine macrophages from most inbred strains are non-permissive to intracellular replication of L. pneumophila, inflammatory macrophages from the mouse strain A/J are completely permissive to intracellular replication of this bacterium. This genetic difference is controlled by the expression of a single autosomal gene designated Lgn1, with non-permissiveness behaving as completely dominant over permissiveness. We have used a total of 25 AXB/BXA recombinant inbred mouse strains and 182 (A/JxC57BL/6J)xA/J segregating backcross progeny (A/J, permissive; C57BL/6J, non-permissive) to map the Lgn1 gene. Animals were individually type for tolerance to intracellular replication by in vitro infection of their inflammatory macrophages with L. pneumophila. All animals segregated into two non-overlapping groups. Examination of the strain distribution pattern of the AXB/BXA strains for Lgn1 initially identified linkage to Chromosome (Chr) 13 markers. Genotyping of the 25 AXB/BXA strains and the 182 backcross progeny for 11 Chr 13 markers established that Lgn1 mapped to Chr 13, with the gene order and intergene distance D13Mit231-(5.5±1.5)-D13Mit193-(2.2±0.9)-D13Mit194-(1.1±0.6)-D13Mit128-(2.6±1.0)-Lgn1-(2.2±0.9)-D13Mit70-(3.9±1.3)-D13Mit73-(7.2±1.7)-D13Mit53-(0.7±0.5)-D13Mit32-(0.7±0.5)-D13Mit77-(0.7±0.5)-D13Mit78. This portion of Chr 13 is homologous to the distal portion of human Chr 5, 5q11–5q13, suggesting a possible location of a human LGN1 homolog. Understanding the molecular basis of the high permissiveness of A/J macrophage to L. pneumophila may shed light on the survival strategy of this bacterium in highly permissive human phagocytes. This may be achieved by positional cloning of Lgn1, and the identification of the Lgn1 subchromosomal region reported here is a first step towards that goal.     

Damage and mutagenesis of E. coli and bacteriophage λ induced by oxathiolane and aziridinyl steroids

λ-Escherichia coli complexes exhibited remarkable sensitivity to the treatment with test steroidal derivatives in the presence of Cu(II). The decline in plaque-forming units after steroid treatment was more pronounced in complexes with some of the irradiation repair-defective mutants of E. coli K-12, i.e., recA, lexA and polA, as compared to uvrA and wild-type strains. The red gene of λ phage and recA gene of E. coli seem to have a complementary effect on the steroid-induced lesions. An enhanced level of mutagenesis was observed when steroid-treated E. coli cells were transformed with steroid-treated pBR322 plasmid DNA. A remarkable degree of c mutation was also observed when steroid I-treated phage particles were allowed to adsorb on steroid-treated wild-type bacteria. Moreover, the oxathione steroid treatment of λcI857-E. coli lysogen resulted in prophage induction in nutrient broth even at 32°C. Thus on the basis of these results, the role of SOS repair system in steroid-induced mutagenesis and repair of DNA lesions in E. coli and bacteriophage λ has been suggested.     

Microbial transformation of isosteviol oxime and the inhibitory effects on NF-κB and AP-1 activation in LPS-stimulated macrophages

Microbial transformation of isosteviol oxime (ent-16-E-hydroxyiminobeyeran-19-oic acid) (2) with Aspergillus niger BCRC 32720 and Absidia pseudocylindrospora ATCC 24169 yielded several compounds. In addition to bioconverting the d-ring to lactone and lactam moieties, 4α-carboxy-13α-hydroxy-13,16-seco-ent-19-norbeyeran-16-oic acid 13,16-lactone (7) and 4α-carboxy-13α-amino-13,16-seco-ent-19-norbeyeran-16-oic acid 13,16-lactam (10), one known compound, ent-1β,7α-dihydroxy-16-oxo-beyeran-19-oic acid (6), and five new compounds, ent-7α-hydroxy-16-E-hydroxyiminobeyeran-19-oic acid (3), ent-1β,7α-dihydroxy-16-E-hydroxyiminobeyeran-19-oic acid (4), ent-1β-hydroxy-16-E-hydroxyiminobeyeran-19-oic acid (5), ent-8β-cyanomethyl-13-methyl-12-podocarpen-19-oic acid (8), and ent-8β-cyanomethyl-13-methyl-13-podocarpen-19-oic acid (9), were isolated from the microbial transformation of 2. Elucidation of the structures of these isolated compounds was primarily based on 1D and 2D NMR, and HRESIMS data, and 3–5 were further confirmed by X-ray crystallographic analyses. Additionally, the inhibitory effects of all of these compounds were evaluated on NF-κB and AP-1 activation in LPS-stimulated RAW 264.7 macrophages. Among the compounds tested, 5 and 10 significantly inhibited NF-κB activation, with 5 showing equal potency to dexamethasone; 3 and 6–9 significantly inhibited AP-1 activation, particularly 8, which showed more inhibitory activity than dexamethasone.     

Synthetic studies on amino-α-glycosides of aminocyclitols. Synthesis of kanamycin analogues

A diastereoisomer of Kanamycin C has been synthesized by a modified Koenigs—Knorr reaction of 3,4,6-tri-O-acetyl-2-(2,4-dinitroanilino)-2-deoxy-α-D-glucopyranosyl bromide with 4-O-(3-acetamido-2,4,6-tri-O-benzyl-3-deoxy-α-D-glucopyranosyl)-N,N′-di[(benzyloxy)carbonyl]-2-deoxystreptamine. Several Kanamycin analogues were synthesized by a similar condensation reaction. Each of the condensed products was isolated as its crystalline tetra-N-acetyl derivative and was proved by n.m.r. spectroscopy in D₂O to have the α-configuration.     

Chromosomal localization of the mouse titin gene and its relationto “muscular dystrophy with myositis” and nebulin genes on chromosome 2

In the mouse, the genes for the structural components of the myofibril titin and nebulin, Ttn and Neb, map to proximal Chr 2, as does the gene for a muscle disease, “muscular dystrophy with myositis,” mdm. To facilitate the evaluation of Ttn and Neb as possible candidates for mdm, we have determined their relative map positions, using a Mus spretus/Mus musculus interspecific backcross. The gene order (distances in cM) cen-Vim-16.9 ± 4.7-Neb-7.6 ± 3.0-Ttn, Acra-18.0 ± 4.9-Pax-6-17.7 ± 4.9-ahas been determined. Considering the standard deviations, Neb, Ttn, and Acra could colocalize with mdm. Using Ttn and Neb probes, DNAs from mdm/mdm and mdm/+ mice were tested for restriction fragment variants in comparison to the M. musculus wildtype. No variants have been found with 11 restriction nucleases. Our data corroborate a conserved synteny comprising genes NEB, TTN, CHRNA1 on human Chr 2q.