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At least 649 records · Page 36Linked to original sources
PubMed1999

Amino-terminal phosphorylation of c-Jun regulates stress-induced apoptosis and cellular proliferation.

c-Jun is a major component of the heterodimeric transcription factor AP-1 and is essential for embryonic development, as fetuses lacking Jun die at mid-gestation with impaired hepatogenesis and primary Jun-/- fibroblasts have a severe proliferation defect and undergo premature senescence in vitro. c-Jun and AP-1 activities are regulated by c-Jun N-terminal phosphorylation (JNP) at serines 63 and 73 through Jun N-terminal kinases(JNKs). JNP is thought to be required for the anti-apoptotic function of c-Jun during hepatogenesis, as mice lacking the JNK kinase SEK1 exhibit liver defects similar to those seen in Jun-/- fetuses. To investigate the physiological relevance of JNP, we replaced endogenous Jun by a mutant Jun allele with serines 63 and 73 mutated to alanines (Jun(tm1wag); hereafter referred to as JunAA). Here we show that primary JunAA fibroblasts have proliferation- and stress-induced apoptotic defects, accompanied by reduced AP-1 activity. JunAA mice are viable and fertile, smaller than controls and resistant to epileptic seizures and neuronal apoptosis induced by the excitatory amino acid kainate. Primary mutant neurons are also protected from apoptosis and exhibit unaltered JNK activity. Our results provide evidence that JNP is dispensable for mouse development, and identify c-Jun as the essential substrate of JNK signalling during kainate-induced neuronal apoptosis.

Animals↗
PubMed2001

The puzzle of zmpB and extensive chain formation, autolysis defect and non-translocation of choline-binding proteins in Streptococcus pneumoniae.

Choline-binding proteins (CBPs) from Streptococcus pneumoniae are involved in several important processes. Inactivation of zmpB, a gene that encodes a surface-located putative zinc metalloprotease, in a S. pneumoniae serotype 4 strain was recently reported to reveal a composite phenotype, including extensive chain formation, lysis defect and transformation deficiency. This phenotype was associated with the lack of surface expression of several CBPs, including the major autolysin LytA. LytA, normally 36 kDa in size, was reported to form an SDS-resistant 80 kDa complex with CinA. ZmpB was therefore proposed to control translocation of CBPs to the surface, possibly through the proteolytic release of CBPs (and RecA) from CinA. Based on the use of 12 independent mariner insertions in the zmpB gene of the well-characterized R6 laboratory strain, we could not confirm several of these observations. Our zmpB mutants: (i) did not form chains; (ii) lysed normally in the presence of deoxycholate, which indicates the presence of a functional autolysin; (iii) transformed at normal frequency; and (iv) contained bona fide CinA and LytA species. Polymorphism of ZmpB between R6 and the serotype 4 isolate could not account for the discrepancy, as inactivation of zmpB (through replacement by transposon-inactivated zmpB R6 alleles) in the latter strain did not affect separation of daughter cells and autolysis. The conflicting observations could be explained by our finding that the reportedly serotype 4 zmpB 'mutant' differed from its S. pneumoniae parent in lacking capsule and in exhibiting characteristic traits of the Streptococcus viridans group, including resistance to optochin.

Amino Acid Sequence↗
PubMed1998

The 16-kDa alpha-crystallin (Acr) protein of Mycobacterium tuberculosis is required for growth in macrophages.

Although the 16-kDa alpha-crystallin homologue of Mycobacterium tuberculosis (MTB) is the dominant protein produced by stationary phase cultures in vitro, it is undetectable in logarithmically growing cultures. By growing bacilli at defined oxygen concentrations, acr transcription was shown to be strongly induced by mildly hypoxic conditions. Acr expression also was found to be induced during the course of in vitro infection of macrophages. The acr gene was replaced with a hygromycin resistance cassette by allelic exchange in MTB H37Rv. The resulting Deltaacr::hpt strain was shown to be equivalent to wild-type H37Rv in in vitro growth rate and infectivity but was significantly impaired for growth in both mouse bone marrow derived macrophages and THP-1 cells. In addition to its proposed role in maintenance of long-term viability during latent, asymptomatic infections, these results establish a role for the Acr protein in replication during initial MTB infection.

Animals↗
PubMed1998

Localization and targeting of the Leishmania donovani hypoxanthine-guanine phosphoribosyltransferase to the glycosome.

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is a key enzyme in the purine salvage pathway of many protozoan parasites. The predicted amino acid sequences of certain HGPRT proteins from parasites of the Trypanosomatidae family reveal a COOH-terminal tripeptide signal that is consistent with the degenerate topogenic signal targeting proteins to the glycosome, a fuel-metabolizing microbody unique to these parasites. To determine definitively the intracellular milieu of HGPRT in these pathogens, polyclonal antiserum to the purified recombinant HGPRT from Leishmania donovani was generated in rabbits, and confocal and immunoelectron microscopy were employed to establish that the L. donovani HGPRT is localized exclusively to the glycosome. No HGPRT protein was detected in delta hgprt null mutants in which both alleles of the HGPRT locus had been replaced by a drug-resistance cassette. Transfectants of the delta hgprt knockout strain in which a wildtype HGPRT was amplified on an expression plasmid contained augmented amounts of HGPRT, all of which was localized to the glycosome. delta hgprt transfectants containing amplified copies of a mutated HGPRT construct in which the Ser-Lys-Val COOH-terminal targeting signal had been deleted expressed HGPRT throughout the parasite, including subcellular organelles such as the nucleus and flagellum. These data demonstrate that the L. donovani HGPRT is compartmentalized exclusively within the glycosome and that the COOH-terminal tripeptide of the protein is necessary to achieve targeting to this organelle.

Animals↗
PubMed2000

Ornithine decarboxylase of Stagonospora (Septoria) nodorum is required for virulence toward wheat.

A knockout strain of Stagonospora (Septoria) nodorum lacking the single ornithine decarboxylase (ODC) allele has been created by targeted gene replacement. A central region of the S. nodorum ODC gene was isolated by polymerase chain reaction using degenerate oligonucleotides and used to probe a lambda genomic library. The gene was sequenced and the encoded ODC protein sequence was shown to be similar to those from other fungi. The functionality of the S. nodorum ODC was confirmed by complementation of an Aspergillus nidulans mutant (puA) strain devoid of ODC activity, restoring growth in the absence of exogenous polyamines. Sporulation of the transformants was reduced suggesting abberant regulation of the S. nodorum gene in A. nidulans. Transformation-mediated gene replacement was used to create strains which were auxotrophic for putrescine and lack ODC coding sequences. Pathogenicity studies on these mutants showed that they are greatly reduced in virulence compared with non-disrupted transformants. This confirms that the strains carrying an ODC disruption cannot obtain sufficient polyamines from the host plant for normal growth and, thus, that fungal ODC may be a suitable target for chemical intervention.

Amino Acid Sequence↗
PubMed2006

Double-stranded RNA-dependent protein kinase phosphorylation of the alpha-subunit of eukaryotic translation initiation factor 2 mediates apoptosis.

As the molecular processes of complex cell stress signaling pathways are defined, the subsequent challenge is to elucidate how each individual event influences the final biological outcome. Phosphorylation of the translation initiation factor 2 (eIF2alpha)atSer(51) is a molecular signal that inhibits translation in response to activation of any of four diverse eIF2alpha stress kinases. We used gene targeting to replace the wild-type Ser(51) allele with an Ala in the eIF2alpha gene to test the hypothesis that translational control through eIF2alpha phosphorylation is a central death stimulus in eukaryotic cells. Homozygous eIF2alpha mutant mouse embryo fibroblasts were resistant to the apoptotic effects of dsRNA, tumor necrosis factor-alpha, and serum deprivation. TNFalpha treatment induced eIF2alpha phosphorylation and activation of caspase 3 primarily through the dsRNA-activated eIF2alpha kinase PKR. In addition, expression of a phospho-mimetic Ser(51) to Asp mutant eIF2alpha-activated caspase 3, indicating that eIF2alpha phosphorylation is sufficient to induce apoptosis. The proapoptotic effects of PKR-mediated eIF2alpha phosphorylation contrast with the anti-apoptotic response upon activation of the PKR-related endoplasmic reticulum eIF2alpha kinase, PERK. Therefore, divergent fates of death and survival can be mediated through phosphorylation at the same site within eIF2alpha. We propose that eIF2alpha phosphorylation is fundamentally a death signal, yet it may promote either death or survival, depending upon coincident signaling events.

Animals↗
PubMed2005

Proceedings of the SMBE Tri-National Young Investigators' Workshop 2005. Molecular genetics of natural populations.

Significant progress in evolutionary genetics has been made by studying, on the one hand, patterns of DNA sequence polymorphism and, on the other, genetic architecture of complex adaptive traits. However, connections between nucleotide variants under selection and adaptively relevant phenotypes are missing. Such connections can be established using precise gene replacement. We review the recent successful introduction of this technique to the analysis of two evolutionarily interesting loci--Odysseus and desaturase2. Both genes have subtle phenotypes that nevertheless could be identified using gene replacement, demonstrating that effects of naturally occurring alleles can be measured in the laboratory. This is an important first step in connecting statistical signatures of selection with adaptation in nature. More candidate genes involved in adaptation, for example, through cloning of genes responsible for reproductive isolation, now need to be identified. Molecular genetic manipulation, DNA polymorphism analysis, and field studies then have to be integrated to provide fresh insights into the mechanisms of evolutionary change.

Alleles↗
PubMed2001

Ornithine decarboxylase knockout in Tapesia yallundae abolishes infection plaque formation in vitro but does not reduce virulence toward wheat.

A knockout strain of Tapesia yallundae lacking the single ornithine decarboxylase (ODC) allele has been created by targeted gene replacement. A central region of the ODC gene was isolated by polymerase chain reaction with degenerate oligonucleotides and used to probe a lambda genomic library. The gene was sequenced, and the encoded ODC protein sequence was shown to be similar to those from other fungi. The functionality of the T. yallundae ODC was confirmed by complementation of an Aspergillus nidulans mutant (puA) strain devoid of ODC activity, restoring growth in the absence of exogenous polyamines. Transformation-mediated gene replacement was used to create strains that were auxotrophic for putrescine and lack ODC coding sequences. ODC knockout strains were unable to differentiate infection structures after in vitro induction and showed an abnormal hyphal branching phenotype. Pathogenicity studies on these mutants showed that, surprisingly, they are not reduced in virulence compared with nondisrupted transformants. This suggests that the strains carrying an ODC disruption can obtain sufficient polyamines from the host plant for normal growth and differentiation and, therefore, that fungal ODC may not be a suitable target for fungicides.

Ascomycota↗
PubMed1993

A proline-rich protein, verprolin, involved in cytoskeletal organization and cellular growth in the yeast Saccharomyces cerevisiae.

A gene (VRP1) encoding a novel proline-rich protein (verprolin) has been isolated from the yeast Saccharomyces cerevisiae as a result of its hybridization to a chick vinculin cDNA probe. The deduced protein sequence contains 24% proline residues present as proline-rich motifs throughout the verprolin sequence. Several of these motifs resemble recently identified sequences shown to bind Src homology 3 (SH3) domains in vitro. Replacement of the wild-type VRP1 allele with a mutant allele results in strains that grow slower than wild-type strains and are temperature sensitive. The vrp1 mutants are impaired in both cell shape and size and display aberrant chitin and actin localization. We propose that verporlin is involved in the maintenance of the yeast actin cytoskeleton, through interactions with other proteins, possibly containing SH3 domains.

Actins↗
PubMed1992

Biogenesis of the yeast vacuole (lysosome). Mutation in the active site of the vacuolar serine proteinase yscB abolishes proteolytic maturation of its 73-kDa precursor to the 41.5-kDa pro-enzyme and a newly detected 41-kDa peptide.

The codon of the catalytic serine in the active site of the vacuolar serine proteinase yscB (PrB) was changed to alanine, yielding the mutant gene prb1-Ala519. Following replacement of the wild-type PRB1 allele with prb1-Ala519, only a 73-kDa molecule was detected by immunoprecipitation with PrB-specific antiserum. The size of the mutant molecule corresponds to the unprocessed cytoplasmic precursor (pre-super-pro-PrB), as detected in sec61 mutants, when translocation into the endoplasmic reticulum is blocked. However, the mutant molecule is completely translocated into the secretory pathway, as indicated by protection from proteinase K digestion in spheroplast lysates in the absence of detergent. When N-glycosylation was inhibited in prb1-Ala519 mutant cells by tunicamycin, a smaller molecule of about 71 kDa appeared consistent with single N-glycosylation and signal-sequence cleavage of the translocated mutant PrB molecule in the endoplasmic reticulum. Thus, the active-site mutation prevents the wild-type processing of the N-glycosylated 73-kDa precursor of PrB to the 41.5 kDa pro-PrB in the endoplasmic reticulum. In order to characterize the processing of wild-type super-pro-PrB in more detail, we generated antibodies against the non-enzymatic superpeptide domain of the 73-kDa precursor expressed in Escherichia coli. We find that, in addition to pro-PrB, a distinct protein (superpeptide) with a mobility of about 41 kDa in SDS/PAGE is generated in the endoplasmic reticulum. Pulse-chase experiments indicate rapid degradation of the 41-kDa superpeptide in wild-type cells. Correspondingly, the superpeptide was virtually undetectable by immunoblotting wild-type cell extracts. In contrast, no degradation of radioactively labeled 41-kDa superpeptide was observed within 60 min in mutant strains deficient in the vacuolar proteinase yscA (PrA), in which maturation of vacuolar pro-PrB to active PrB is blocked. Accordingly, superpeptide antigenic material was readily detected by immunoblotting cell extracts and enriched in vacuolar preparations of PrA deficient mutant cells. These results indicate that the superpeptide and pro-PrB travel to the vacuole, where the superpeptide is rapidly degraded upon pro-PrB activation to PrB. Using purified vacuoles, rapid degradation of the superpeptide was reconstituted in vitro by addition of either mature PrA or mature PrB. However, the PrA-triggered in vitro degradation of the superpeptide required PrB activity, as this process was inhibited in the presence of the PrB inhibitor chymostatin.(ABSTRACT TRUNCATED AT 400 WORDS)

Binding Sites↗
PubMed1992

The polyanion-binding domain of cytoplasmic Lys-tRNA synthetase from Saccharomyces cerevisiae is not essential for cell viability.

Cytoplasmic Lys-tRNA synthetase (LysRS) from Saccharomyces cerevisiae is a dimeric enzyme made up of identical subunits of 68 kDa. By limited proteolysis, this enzyme can be converted to a truncated dimer without loss of activity. Whereas the native enzyme strongly interacts with polyanionic carriers, the modified form displays reduced binding properties. KRS1 is the structural gene for yeast cytoplasmic LysRS. It encodes a polypeptide with an amino-terminal extension composed of about 60-70 amino acid residues, compared to its prokaryotic counterpart. This segment, containing 13 lysine residues, is removed upon proteolytic treatment of the native enzyme. The aim of the present study was to probe in vivo the significance of this amino-terminal extension. We have constructed derivatives of the KRS1 gene, encoding enzymes lacking 58 or 69 amino-terminal residues and, by site-directed mutagenesis, we have changed four or eight lysine residues from the amino-terminal segment of LysRS into glutamic acids. Engineered proteins were expressed in vivo after replacement of the wild-type KRS1 allele. The mutant enzymes displayed reduced specific activities (2-100-fold). A series of carboxy-terminal deletions, encompassing 3, 10 or 15 amino acids, were introduced into the LysRS mutants with modified amino-terminal extensions. The removal of three residues led to a 2-7-fold increase in the specific activity of the mutant enzymes. This partial compensatory effect suggests that interactions between the two extreme regions of yeast LysRS are required for a proper conformation of the native enzyme. All KRS1 derivatives were able to sustain growth of yeast cells, although the mutant cell lines displaying a low LysRS activity grew more slowly. The expression, as single-copy genes, of mutant enzymes with a complete deletion of the amino-terminal extension or with four Lys----Glu mutations, that displayed specific activities close to that of the wild-type LysRS, had no discernable effect on cell growth. We conclude that the polycationic extensions of eukaryotic aminoacyl-tRNA synthetases are dispensable, in vivo, for aminoacylation activities. The results are discussed in relation to the triggering role in in situ compartmentalization of protein synthesis that has been ascribed to the polypeptide-chain extensions that characterize most, if not all, eukaryotic aminoacyl-tRNA synthetases.

Alleles↗
PubMed1993

A transformant of Aspergillus fumigatus deficient in the antigenic cytotoxin ASPFI.

The aspfI gene encoding a ribonucleotoxin, a putative virulence factor of Aspergillus fumigatus, was inactivated by gene disruption. Gene replacement through homologous recombination by the disrupted allele tagged by the hygromycin B resistance marker was performed by transformation of a pathogenic strain. One transformant with the disrupted aspfI gene failed to produce the ASPFI protein and was shown to be pathogenic for mice. We concluded that this ribotoxin is not a main factor in the colonization of the lung tissues by A. fumigatus.

Allergens↗
PubMed1996

Induction of actinorhodin production by rpsL (encoding ribosomal protein S12) mutations that confer streptomycin resistance in Streptomyces lividans and Streptomyces coelicolor A3(2).

A strain of Streptomyces lividans, TK24, was found to produce a pigmented antibiotic, actinorhodin, although S. lividans normally does not produce this antibiotic. Genetic analyses revealed that a streptomycin-resistant mutation str-6 in strain TK24 is responsible for induction of antibiotic synthesis. DNA sequencing showed that str-6 is a point mutation in the rpsL gene encoding ribosomal protein S12, changing Lys-88 to Glu. Gene replacement experiments with the Lys88-->Glu str allele demonstrated unambiguously that the str mutation is alone responsible for the activation of actinorhodin production observed. In contrast, the strA1 mutation, a genetic marker frequently used for crosses, did not restore actinorhodin production and was found to result in an amino acid alteration of Lys-43 to Asn. Induction of actinorhodin production was also detected in strain TK21, which does not harbor the str-6 mutation, when cells were incubated with sufficient streptomycin or tetracycline to reduce the cell's growth rate, and 40 and 3% of streptomycin- or tetracycline-resistant mutants, respectively, derived from strain TK21 produced actinorhodin. Streptomycin-resistant mutations also blocked the inhibitory effects of relA and brgA mutations on antibiotic production, aerial mycelium formation or both. These str mutations changed Lys-88 to Glu or Arg and Arg-86 to His in ribosomal protein S12. The decrease in streptomycin production in relC mutants in Streptomyces griseus could also be abolished completely by introducing streptomycin-resistant mutations, although the impairment in antibiotic production due to bldA (in Streptomyces coelicolor) or afs mutations (in S. griseus) was not eliminated. These results indicate that the onset and extent of secondary metabolism in Streptomyces spp. is significantly controlled by the translational machinery.

Alleles↗
PubMed2002

Cavitary pneumonia in an AIDS patient caused by an unusual Bordetella bronchiseptica variant producing reduced amounts of pertactin and other major antigens.

Although Bordetella bronchiseptica can infect and colonize immunocompromised humans, its role as a primary pathogen in pneumonia and other respiratory processes affecting those patients remains controversial. A case of cavitary pneumonia caused by B. bronchiseptica in an AIDS patient is presented, and the basis of the seemingly enhanced pathogenic potential of this isolate (designated 814) is investigated. B. bronchiseptica was the only microorganism recovered from sputum, bronchoalveolar lavage fluid, and samples taken through the protected brush catheter. Unlike previous work reporting the involvement of B. bronchiseptica in cases of pneumonia, antibiotic treatment selected on the basis of in vitro antibacterial activity resulted in clearance of the infection and resolution of the pulmonary infiltrate. Although isolate 814 produced reduced amounts of several major antigens including at least one Bvg-activated factor (pertactin), the molecular basis of this deficiency was found to be BvgAS independent since the defect persisted after the bvgAS locus of isolate 814 was replaced with a wild-type bvgAS allele. Despite its prominent phenotype, isolate 814 displayed only a modest yet a significant deficiency in its ability to colonize the respiratory tracts of immunocompetent rats at an early time point. Interestingly, the antibody response elicited by isolate 814 in these animals was almost undetectable. We propose that isolate 814 may be more virulent in immunocompromised patients due, at least in part, to its innate ability to produce low amounts of immunogenic factors which may be required at only normal levels for the interaction of this pathogen with its immunocompetent natural hosts.

AIDS-Related Opportunistic Infections↗
PubMed1984

Disruption of regulatory gene GAL80 in Saccharomyces cerevisiae: effects on carbon-controlled regulation of the galactose/melibiose pathway genes.

In Saccharomyces cerevisiae, the transcriptional expression of the galactose-melibiose catabolic pathway genes is under the control of at least three regulatory genes, GAL4, GAL80, and GAL3. We have isolated the GAL80 gene and have studied the effect of a null mutation on the carbon-controlled regulation of the MEL1 and GAL cluster genes. The null mutation was achieved in vivo by replacing the chromosomal wild-type GAL80 allele with an in vitro-created GAL80 deletion-disruption mutation. Enzyme activities and RNA levels for the GAL cluster and MEL1 genes were constitutively expressed in the null mutant strain grown on glycerol-lactate and were higher than in the isogenic wild-type yeast strain when compared after growth on galactose. Carbon catabolite repression of the GAL cluster and MEL1 genes, which occurs at the level of transcription, is retained in the null mutant. Deletion of the GAL80 gene in a gal4 cell does not restore GAL cluster and MEL1 gene expression. The data demonstrate that (i) the GAL80 protein is a purely negative regulator, (ii) the GAL80 protein does not mediate carbon catabolite repression, and (iii) the GAL4 protein is not simply an antagonizer of GAL80-mediated repression.

Chromosome Deletion↗
PubMed2006

Effects of membrane and soluble EPCR on the hemostatic balance and endotoxemia in mice.

Recent studies have shown that endothelial protein C receptor (EPCR) polymorphisms and soluble EPCR levels are associated with thrombotic diseases. It is unknown whether membrane EPCR (mEPCR) heterozygosity and/or physiologically elevated sEPCR levels directly impact the hemostatic balance and the outcome of endotoxemia. In these studies, thrombin infusion experiments revealed that EPCR heterozygosity (Procr+/-) impaired protein C activation by approximately 30%. Infusion of factor Xa with phospholipid demonstrated that the Procr+/- genotype increased the coagulant response relative to wild-type mice. Challenge of the Procr+/- mice with lipopolysaccharide (LPS) did not significantly exaggerate their response compared with wild-type mice. We also generated mice in which one allele of full-length EPCR was replaced by sEPCR (Procrs/+). Compared with Procr+/- mice, Procrs/+ mice had 5-fold higher sEPCR and similar mEPCR levels. Procr+/- and Procrs/+ mice generated similar levels of activated protein C (APC) upon thrombin infusion. They also exhibited a similar coagulant response upon factor Xa/phospholipid infusion. Only supraphysiologic levels of sEPCR could influence protein C activation and exaggerate the coagulant response. In conclusion, mEPCR, but not physiologically elevated sEPCR, regulated protein C activation. Procr heterozygosity results in a mild increase of thrombosis tendency and little influence on the response to endotoxin.

Animals↗
PubMed1999

Craniofacial, vestibular and bone defects in mice lacking the Distal-less-related gene Dlx5.

The Dlx5 gene encodes a Distal-less-related DNA-binding homeobox protein first expressed during early embryonic development in anterior regions of the mouse embryo. In later developmental stages, it appears in the branchial arches, the otic and olfactory placodes and their derivatives, in restricted brain regions, in all extending appendages and in all developing bones. We have created a null allele of the mouse Dlx5 gene by replacing exons I and II with the E. coli lacZ gene. Heterozygous mice appear normal. Beta-galactosidase activity in Dlx5+/- embryos and newborn animals reproduces the known pattern of expression of the gene. Homozygous mutants die shortly after birth with a swollen abdomen. They present a complex phenotype characterised by craniofacial abnormalities affecting derivatives of the first four branchial arches, severe malformations of the vestibular organ, a delayed ossification of the roof of the skull and abnormal osteogenesis. No obvious defect was observed in the patterning of limbs and other appendages. The defects observed in Dlx5-/- mutant animals suggest multiple and independent roles of this gene in the patterning of the branchial arches, in the morphogenesis of the vestibular organ and in osteoblast differentiation.

Animals↗
PubMed1999

IkappaBepsilon-deficient mice: reduction of one T cell precursor subspecies and enhanced Ig isotype switching and cytokine synthesis.

Three major inhibitors of the NF-kappaB/Rel family of transcription factors, IkappaBalpha, IkappaBbeta, and IkappaBepsilon, have been described. To examine the in vivo role of the most recently discovered member of the IkappaB family, IkappaBepsilon, we generated a null allele of the murine IkappaBepsilon gene by replacement of all coding sequences with nlslacZ. Unlike IkappaBalpha nullizygous mice, mice lacking IkappaBepsilon are viable, fertile, and indistinguishable from wild-type animals in appearance and histology. Analysis of beta-galactosidase expression pattern revealed that IkappaBepsilon is mainly expressed in T cells in the thymus, spleen, and lymph nodes. Flow cytometric analysis of immune cell populations from the bone marrow, thymus, spleen, and lymph nodes did not show any specific differences between the wild-type and the mutant mice, with the exception of a reproducible 50% reduction of the CD44-CD25+ T cell subspecies. The IkappaBepsilon-null mice present constitutive up-regulation of IgM and IgG1 Ig isotypes together with a further increased synthesis of these two isotypes after immunization against T cell-dependent or independent Ags. The failure of observable augmentation of constitutive nuclear NF-kappaB/Rel-binding activity is probably due to compensatory mechanisms involving IkappaBalpha and IkappaBbeta, which are up-regulated in several organs. RNase-mapping analysis indicated that IL-1alpha, IL-1beta, IL-1Ra, and IL-6 mRNA levels are constitutively elevated in thioglycolate-elicited IkappaBepsilon-null macrophages in contrast to GM-CSF, G-CSF, and IFN-gamma, which remain undetectable.

Animals↗