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Frank R Jirik

Publications and source records attributed to Frank R Jirik.

17 recordsLinked to original sources
PubMed2006

Differing patterns of genetic instability in mice deficient in the mismatch repair genes Pms2, Mlh1, Msh2, Msh3 and Msh6.

Defects in genes associated with DNA mismatch repair (MMR) have been linked to hereditary colon cancer. Because the MMR pathway includes multiple factors with both overlapping and divergent functions, we sought to compare the impact of deficiencies in each of several MMR genes on genetic instability using a collection of knock-out mouse models. We investigated mutation frequencies and patterns in MMR-deficient mice using two transgenic reporter genes, supFG1 and cII, in the context of mice deficient for Pms2, Mlh1, Msh2, Msh3 or Msh6 or both Msh2 and Msh3 or both Msh3 and Msh6. We found that the mean mutation frequencies of all of the MMR-deficient mice were significantly higher than the mean mutation frequencies of wild-type mice. Mlh1-deficient mice and Msh2-deficient mice had the highest mutation frequencies in a comparison of the single nullizygous mice. Of all the mice studied, mice nullizygous for both Msh2 and Msh3 and those nullizygous for both Msh3 and Msh6 displayed the greatest overall increases in mutation frequencies compared with wild-type mice. Sequence analysis of the mutated reporter genes revealed significant differences between the individual groups of MMR-deficient mice. Taken together, our results further characterize the functions of the MMR factors in mutation avoidance and provide in vivo correlation to biochemical models of the MMR pathway.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Prion diseases and the gastrointestinal tract.

The gastrointestinal (GI) tract plays a central role in the pathogenesis of transmissible spongiform encephalopathies. These are human and animal diseases that include bovine spongiform encephalopathy, scrapie and Creutzfeldt-Jakob disease. They are uniformly fatal neurological diseases, which are characterized by ataxia and vacuolation in the central nervous system. Although they are known to be caused by the conversion of normal cellular prion protein to its infectious conformational isoform (PrPsc) the process by which this isoform is propagated and transported to the brain remains poorly understood. M cells, dendritic cells and possibly enteroendocrine cells are important in the movement of infectious prions across the GI epithelium. From there, PrPsc propagation requires B lymphocytes, dendritic cells and follicular dendritic cells of Peyer's patches. The early accumulation of the disease-causing agent in the plexuses of the enteric nervous system supports the contention that the autonomic nervous system is important in disease transmission. This is further supported by the presence of PrPsc in the ganglia of the parasympathetic and sympathetic nerves that innervate the GI tract. Additionally, the lymphoreticular system has been implicated as the route of transmission from the gut to the brain. Although normal cellular prion protein is found in the enteric nervous system, its role has not been characterized. Further research is required to understand how the cellular components of the gut wall interact to propagate and transmit infectious prions to develop potential therapies that may prevent the progression of transmissible spongiform encephalopathies.

Autonomic Nervous System↗
PubMed2005

Human hormone-sensitive lipase (HSL): expression in white fat corrects the white adipose phenotype of HSL-deficient mice.

In white adipose tissue (WAT), hormone-sensitive lipase (HSL) can mediate lipolysis, a central pathway in obesity and diabetes. Gene-targeted HSL-deficient (HSL-/-) mice with no detectable HSL peptide or activity (measured as cholesteryl esterase) have WAT abnormalities, including low mass, marked heterogeneity of cell diameter, increased diacylglycerol content, and low beta-adrenergic stimulation of adipocyte lipolysis. Three transgenic mouse strains preferentially expressing human HSL in WAT were bred to a HSL-/- background. One, HSL-/- N, expresses normal human HSL (41.3 +/- 9.1% of normal activity); two express a serine-to-alanine mutant (S554A) initially hypothesized to be constitutively active: HSL-/- ML, 50.3 +/- 12.3% of normal, and HSL-/- MH, 69.8 +/- 15.8% of normal. In WAT, HSL-/- N mice resembled HSL+/+ controls in WAT mass, histology, diacylglyceride content, and lipolytic response to beta-adrenergic agents. In contrast, HSL-/- ML and HSL-/- MH mice resembled nontransgenic HSL-/- mice, except that diacylglycerol content and perirenal and inguinal WAT masses approached normal in HSL-/- MH mice. Therefore, 1) WAT expression of normal human HSL markedly improves HSL-/- WAT biochemically, physiologically, and morphologically; 2) similar levels of S554A HSL have a low physiological effect despite being active in vitro; and 3) diacylglycerol accumulation is not essential for the development of the characteristic WAT pathology of HSL-/- mice.

Adipocytes↗
PubMed2005

Degenerative knee joint disease in mice lacking 3'-phosphoadenosine 5'-phosphosulfate synthetase 2 (Papss2) activity: a putative model of human PAPSS2 deficiency-associated arthrosis.

OBJECTIVE: Murine brachymorphism (bm) results from an autosomal recessive mutation of the Papss2 gene that encodes 3'-phosphoadenosine 5'-phosphosulfate synthetase 2, one of the principal enzymes required for the sulfation of extracellular matrix molecules in cartilage and other tissues. A spondyloepimetaphyseal dysplasia has been identified in Pakistani kindred having a mutation of PAPSS2. In addition to skeletal malformations that include short stature evident at birth due to limb shortening, brachydactyly, and kyphoscoliosis, affected individuals demonstrate premature onset degenerative joint disease. We investigated whether loss of Papss2 activity would similarly lead to degenerative joint disease in mice. METHODS: Mice carrying the bm mutation on a C57BL/6 background were obtained from the Jackson Laboratory. Limbs were analyzed by micro-computed tomography (microCT) and histology. RESULTS: At 12 months of age both male and female bm mice exhibited severe degenerative knee joint disease, with cartilage damage being primarily evident in the patello-femoral and medial compartments. Control 12-14-month-old C57BL/6 mice, in contrast, only occasionally demonstrated minimal cartilage damage. muCT imaging of bm limbs revealed shortened diaphyses associated with flared metaphyses in the proximal elements of both fore and hind limbs. Additionally, the bm hind limbs demonstrated extensive structural alterations, characterized by distortion of the patello-femoral groove, and prominent bowing of both tibia and fibula. CONCLUSIONS: The bm mutant, which develops severe articular cartilage lesions of the knee joint by approximately 12 months of age, represents a novel example of murine degenerative joint disease, possibly representing a model of human PAPSS2 deficiency-associated arthrosis.

Animals↗
PubMed2005

Granzyme B induces endothelial cell apoptosis and contributes to the development of transplant vascular disease.

Endothelial cell death induced by cytotoxic T cells is a key initiating event in the development of transplant vascular disease (TVD), the leading cause of late solid organ transplant failure. We studied the role of the granzyme B (GrB) pathwaye, which is one of the main mechanisms by which T cells induce apoptosis of allogeneic targets, in the pathogenesis of TVD. Granzyme B, in combination with perforin (pfn), induced apoptosis of cultured endothelial cells. In hearts transplanted into GrB knockout (GrB-KO) mice, there was a similar level of vasculitis as compared to WT mice, indicating that GrB does not affect immune infiltration into allograft arteries. However, there was a significant reduction in luminal narrowing of allograft arteries from GrB-KO mice as compared to WT recipients. These results indicate that GrB plays a role in endothelial cell death in allograft arteries and in the resultant development of TVD.

Animals↗
PubMed2004

Mutational-reporter transgenes rescued from mice lacking either Mgmt, or both Mgmt and Msh6 suggest that O6-alkylguanine-induced miscoding does not contribute to the spontaneous mutational spectrum.

O6-methylguanine methyltransferase, Mgmt, constitutes the first line of defense against O6-alkylguanine, which can result in G : C to A : T transitions upon DNA replication. Mgmt has been found in organisms as diverse as archaebacteria and mammals. This evolutionary conservation suggests that all organisms may be exposed to either endogenous or environmental alkylating agents. We thus hypothesized that tissues of Mgmt-/- mice would exhibit elevated mutant frequencies. Employing the Big Blue trade mark transgenic system, we evaluated lacI mutants rescued from liver and small intestinal DNA of young Mgmt-/- mice. Interestingly, while there was a small difference between Mgmt-/- mice and controls with respect to lacI mutant frequency, no differences attributable to Mgmt deficiency were apparent in the mutational spectra. Although mutations stemming from O6-guanine alkylations would be predicted to be cumulative, we found no evidence of an Mgmt-dependent alteration in mutation spectrum in DNA samples from 12 month-old mice. To optimize our ability to detect mutations resulting from O6-alkylguanine-induced G : T mismatches, mice with combined deficiencies of Mgmt and the DNA mismatch repair molecule, Msh6, were analysed. In spite of this strategy, we observed no significant differences between Mgmt-/- Msh6-/- and Msh6-/- mouse lacI mutations, except for a trend towards a greater percentage (of total transitions) of G : C to A : T changes in Mgmt-/-Msh6-/- livers. Therefore, despite the striking evolutionary conservation of Mgmt, deficiency of this gene did not significantly impact the spontaneous lacI mutational spectrum in vivo.

Age Factors↗
PubMed2004

Anemia, thrombocytopenia, leukocytosis, extramedullary hematopoiesis, and impaired progenitor function in Pten+/-SHIP-/- mice: a novel model of myelodysplasia.

The myeloproliferative disorder of mice lacking the Src homology 2 (SH2)-containing 5' phosphoinositol phosphatase, SHIP, underscores the need for closely regulating phosphatidylinositol 3-kinase (PI3K) pathway activity, and hence levels of phosphatidylinositol species during hematopoiesis. The role of the 3' phosphoinositol phosphatase Pten in this process is less clear, as its absence leads to embryonic lethality. Despite Pten heterozygosity being associated with a lymphoproliferative disorder, we found no evidence of a hematopoietic defect in Pten(+/-) mice. Since SHIP shares the same substrate (PIP(3)) with Pten, we hypothesized that the former might compensate for Pten haploinsufficiency in the marrow. Thus, we examined the effect of Pten heterozygosity in SHIP(-/-) mice, predicting that further dysregulation of PIP(3) metabolism would exacerbate the pheno-type of the latter. Indeed, compared with SHIP(-/-) mice, Pten(+/-)SHIP(-/-) animals developed a myelodysplastic phenotype characterized by increased hepatosplenomegaly, extramedullary hematopoiesis, anemia, and thrombocytopenia. Consistent with a marrow defect, clonogenic assays demonstrated reductions in committed myeloid and megakaryocytic progenitors in these animals. Providing further evidence of a Pten(+/-)SHIP(-/-) progenitor abnormality, reconstitution of irradiated mice with marrows from these mice led to a marked defect in short-term repopulation of peripheral blood by donor cells. These studies suggest that the regulation of the levels and/or ratios of PI3K-derived phosphoinositol species by these 2 phosphatases is critical to normal hematopoiesis.

Anemia↗
PubMed2004

Inhibition of myogenesis in transgenic mice expressing the human DMPK 3'-UTR.

Myotonic dystrophy (DM1) is a multisystemic disorder caused by a CTG repeat expansion within the 3'-UTR of the DMPK gene. DM1 is characterized by delayed muscle development, muscle weakness and wasting, cardiac conduction abnormalities, cognitive defects and cataracts. Recent studies have demonstrated that the disease mechanism involves a dominant gain-of-function conferred upon mutant transcripts by expanded repeats. However, further attempts to model aspects of DM muscle pathology in cultured myoblasts suggest that 3'-UTR sequences flanking the CTG repeat tract are also required for full expression of the disease phenotype. Here, we report that overexpression of the DMPK 3'-UTR including either wild-type (11) or expanded (91) CTG repeats results in aberrant and delayed muscle development in fetal transgenic mice. In addition, transgenic animals with both expanded and wild-type CTG repeats display muscle atrophy at 3 months of age. Primary myoblast cultures from both 11 and 91 repeat mice display reduced fusion potential, but a greater reduction is observed in the 91 repeat cultures. Taken together, these data indicate that overexpression of the DMPK 3'-UTR interferes with normal muscle development in mice and that this is exacerbated by inclusion of a mutant repeat. This suggests that the delayed muscle development in DM1 involves an interplay between the expanded CTG repeat and adjacent 3'-UTR sequences.

3' Untranslated Regions↗
PubMed2004

Compact intergenic regions of the pufferfish genome facilitate isolation of gene promoters: characterization of Fugu 3'-phosphoadenosine 5'-phosphosulfate synthase 2 (fPapss2) gene promoter function in transgenic Xenopus.

The highly compact nature of the pufferfish (Fugu rubripes) genome renders it a useful tool not only for annotating coding regions within vertebrate genomes, but also for the identification of sequences important to gene regulation. Indeed, owing to this compaction it will be feasible in many instances to initiate analyses using entire intergenic regions when mapping gene promoters; a strategy that is very rarely feasible with the expanded genomes of other species. Stemming from our interest in studying promoters expressed in chondrocytes, we selected for study the intergenic region upstream of Fugu 3'-phosphoadenosine 5'-phosphosulfate synthase 2, fPapss2, a gene required for the normal development of cartilage extracellular matrix. Functional characterization of the entire fPapss2 5' intergenic region was carried out by monitoring expression of the enhanced green fluorescent protein (EGFP) gene reporter in the developing cartilage of transgenic Xenopus laevis. By evaluating a series of 5' intergenic region deletions we defined a minimal fPapss2 sequence of approximately 300 bp that was essential for EGFP expression in tadpole cartilage. This functional analysis of an entire Fugu intergenic region, combined with the efficiency of Xenopus transgenesis, serves as a model for the rapid characterization of evolutionarily-conserved regulatory regions of other pufferfish genes.

Animals↗
PubMed2003

The regulation of retina specific expression of rhodopsin gene in vertebrates.

Rhodopsin genes in most vertebrate species, with the exception of teleost fishes, contain introns. Despite differences in the gene structure, similar regulatory motifs have been identified in fish, amphibian and mammalian rhodopsin promoters, suggesting that rhodopsin gene regulation may be conserved in vertebrates. However, there is no direct evidence to support this notion. To address this, the rhodopsin promoter from the pufferfish, Fugu rubripes, was isolated and tested in transgenic mice and frogs. A 6.5 kb Fugu genomic fragment containing the rhodopsin gene and 4.5 kb 5' flanking region was able to direct expression of the Fugu rhodopsin gene to the retina of transgenic mice. In transgenic tadpoles, photoreceptor rod cell-specific expression of a reporter gene was achieved using only 500 bp Fugu rhodopsin promoter fragment. Mutagenesis of this promoter fragment revealed that a conserved NRE-like motif is crucial for the retina-specific expression. Our investigation suggests that the regulation of retinal specific expression is conserved in the pufferfish, frog and mouse and that the ancestral intron-containing rhodopsin gene has been displaced by an intronless copy in teleosts.

Animals↗
PubMed2003

Increased sensitivity of Fancc-deficient hematopoietic cells to nitric oxide and evidence that this species mediates growth inhibition by cytokines.

Fanconi anemia complementation group C (Fancc)-deficient murine bone marrow progenitors demonstrate increased sensitivity to growth inhibition by interferon gamma (IFNgamma), tumor necrosis factor alpha (TNFalpha), and macrophage inflammatory protein 1alpha (MIP-1alpha). This property has been proposed as a possible pathogenic factor in the marrow failure seen in Fanconi anemia. Supporting our hypothesis that nitric oxide (NO) production might be a common effector in this sensitivity, we found that cytokine-mediated growth inhibition of Fancc(-/-) bone marrow cells was prevented by inhibiting NO synthase activity. Interestingly, Fancc(-/-) hematopoietic cells also exhibited increased growth inhibition on exposure to 2 distinct NO-generating agents, S-nitroso-N-acetyl-D, L-penicillamine (SNAP) and diethylenetriamine nitric oxide adduct (DETA/NO). In keeping with the sensitivity of Fancc(-/-) cells to IFNgamma, inducible nitric oxide synthase (iNOS) levels and nitrite release were both increased following stimulation of Fancc(-/-) macrophages with this cytokine, either alone or in combination with bacterial lipopolysaccharide. Suggesting a plausible mechanism for the increased expression of iNOS, IFNgamma-stimulated Fancc(-/-) macrophages generated higher levels of phospho-Stat1, a positive regulator of inos (nos2) gene expression. These observations, while confined to C57BL/6 Fancc(-/-) hematopoietic cells, raise the possibility that nitric oxide has a role in the pathogenesis of Fanconi anemia.

Animals↗
PubMed2003

Imaging skeletal pathology in mutant mice by microcomputed tomography.

OBJECTIVE: We describe the utility of microcomputed tomography ( micro CT) for imaging skeletal abnormalities in rodent model systems. For the purpose of illustration, the progressive ankylosis (ank) mutant was selected. ank mice develop prominent articular and periarticular calcifications at multiple anatomical sites, including paws, elbows, knees, and vertebrae. METHODS: Forelimbs, hindlimbs, and proximal tail vertebrae of 4-month-old female ank/ank mice were scanned at 15 micro m resolution using a SkyScan 1072 micro CT instrument and images were generated using Analyze 4.0 software. RESULTS: This technique was able to show, in 3-dimensional images, the abnormal calcification of ank/ank mice, which was readily observed within joint surfaces, on periosteal surfaces, sesamoid bones, menisci, and joint capsules, as well as other periarticular ligamentous structures. CONCLUSION: As illustrated by the example of the progressive ankylosis mutant, micro CT represents a powerful emerging tool for identifying and monitoring the progression of developmental or acquired skeletal abnormalities within rodent models.

Animals↗
PubMed2002

Elevated mutant frequencies and predominance of G:C to A:T transition mutations in Msh6(-/-) small intestinal epithelium.

The DNA mismatch repair (MMR) system is primarily responsible for purging newly synthesized DNA of errors incurred during semi-conservative replication. Lesion recognition is initially carried out by one of two heterodimeric protein complexes, MutS(alpha) or MutS(beta). While the former, comprised of MSH2 and MSH6, recognizes mispairs as well as short (1-2 nucleotide) insertions/deletions (IDLs), the latter, made up of MSH2 and MSH3, is primarily responsible for recognizing 2-6 nucleotide IDLs. As most of the functional information on these heterodimers is derived from in vitro studies, it was of interest to study the in vivo consequences of a lack of MutS(alpha). To this end, Big Blue( trade mark ) mice, that carry a lacI(+) transgenic lambda shuttle-phage mutational reporter, were crossed with Msh6(-/-) mice to evaluate the specific contribution of MutS(alpha) to genome integrity. Consistent with the importance of MutS(alpha) in lesion surveillance, small intestine epithelial cell DNA derived from lacI(+) Msh6(-/-) mice exhibited striking increases (average of 41-fold) in spontaneous mutant frequencies. Furthermore, the lacI gene mutation spectrum was dominated by G:C to A:T transitions, highlighting the critical importance of the MutS(alpha) complex in suppressing this frequently observed type of spontaneous mutation.

Animals↗
PubMed2002

Disruption of a single Pten allele augments the chemotactic response of B lymphocytes to stromal cell-derived factor-1.

The tumor suppressor, Pten, has emerged as a critical negative regulator of phosphatidylinositol-3-kinase-dependent intracellular signaling pathways responsible for phenomena such as cellular adhesion, proliferation, and apoptosis. Herein, we present evidence that Pten regulates chemokine-dependent events in B lymphocytes. Primary B cells isolated from Pten(+/-) mice demonstrated increased responsiveness to stromal cell-derived factor-1-induced chemotaxis. This was accompanied by an elevated level of protein kinase B phosphorylation on Ser(473). Our results suggest not only that Pten may be an important regulator of stromal cell-derived factor-1-directed chemotaxis, but also that Pten heterozygosity is associated with increased cellular sensitivity to this chemokine, likely via dysregulation of events lying downstream of phosphatidylinositol-3-kinase. These observations suggest a mechanism by which loss of a single Pten allele may confer a selective advantage on cells during multistep tumor progression.

Adjuvants, Immunologic↗
PubMedDate not supplied

The brachymorph mouse and the developmental-genetic basis for canalization and morphological integration.

Although it is well known that many mutations influence phenotypic variability as well as the mean, the underlying mechanisms for variability effects are very poorly understood. The brachymorph (bm) phenotype results from an autosomal recessive mutation in the phosphoadenosine-phosphosulfate synthetase 2 gene (Papps2). A major cranial manifestation is a dramatic reduction in the growth of the chondrocranium which results from undersulfation of glycosaminoglycans (GAGs) in the cartilage matrix. We found that this reduction in the growth of the chondrocranium is associated with an altered pattern of craniofacial shape variation, a significant increase in phenotypic variance and a dramatic increase in morphological integration for craniofacial shape. Both effects are largest in the basicranium. The altered variation pattern indicates that the mutation produces developmental influences on shape that are not present in the wildtype. As the mutation dramatically reduces sulfation of GAGs, we infer that this influence is variation among individuals in the degree of sulfation, or variable expressivity of the mutation. This variation may be because of genetic variation at other loci that influence sulfation, environmental effects, or intrinsic effects. We infer that chondrocranial development exhibits greater sensitivity to variation in the sulfation of chondroitin sulfate when the degree of sulfation is low. At normal levels, sulfation probably contributes minimally to phenotypic variation. This case illustrates canalization in a particular developmental-genetic context.

Anatomy, Comparative↗