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Biomedical subjects

I Kola

Publications and source records attributed to I Kola.

At least 19 recordsLinked to original sources
PubMed2004

Detailed mapping of the ERG-ETS2 interval of human chromosome 21 and comparison with the region of conserved synteny on mouse chromosome 16.

We have carried out a detailed annotation of 550 kb of genomic DNA on human chromosome 21 containing the ERG and ETS2 genes. Comparative genomic analysis between this region and the interval of conserved synteny on mouse chromosome 16 indicated that the order and orientation of the ERG and ETS2 genes were conserved and revealed several regions containing potential conserved noncoding sequences. Four pseudogenes including those for small protein G, laminin receptor, human transposase protein and meningioma-expressed antigen were identified. A potentially novel gene (C21orf24) with alternative mRNA transcripts, consensus splice donor and acceptor sites, but no coding potential nor murine orthologue, was identified and found to be expressed in a range of human cell lines. We have identified four novel splice variants that arise from a previously undescribed 5' exon of the human ERG gene. Comparison of the cDNA sequences enabled us to determine the complete exon-intron structure of the ERG gene. We have also identified the presence of noncoding RNAs in the first and second introns of the ETS2 gene. Our studies have important implications for Down syndrome as this region contains multiple mRNA transcripts, both coding and potentially noncoding, that may play as yet undescribed roles in the pathogenesis of this disorder.

Alternative Splicing↗
PubMed2003

The chromosome 21 transcription factor ETS2 transactivates the beta-APP promoter: implications for Down syndrome.

The gene that codes for beta-amyloid precursor protein (beta-APP), a protein centrally involved in senile plaque formation in Down syndrome (DS) and Alzheimer's disease (AD), is located on chromosome 21. In DS beta-APP expression is three- to fourfold higher than what is expected from the 1.5-fold increased gene load, suggesting that other genes on chromosome 21 directly or indirectly can further up-regulate beta-APP. Here we show that the chromosome 21 transcription factor ETS2 transactivates the beta-APP gene via specific Ets binding sites in the beta-APP promoter and, in this respect, cooperates with the transcription factor complex AP1. We further show that brains and primary neuronal cultures from Ets2 transgenic mice, as well as 3T3 fibroblasts that overexpress ETS2, display molecular abnormalities also seen in DS, such as elevated expression of beta-APP protein, an increase in presenilin-1 and increased beta-amyloid production. We conclude that ETS2 is a transcriptional regulator of beta-APP and that overexpression of ETS2 in DS may play a role in the pathogenesis of the brain abnormalities in DS and possibly AD.

3T3 Cells↗
PubMed2003

ETS2 overexpression in transgenic models and in Down syndrome predisposes to apoptosis via the p53 pathway.

ETS2 is a transcription factor encoded by a gene on human chromosome 21 and alterations in its expression have been implicated in the pathophysiological features of Down syndrome (DS). This study demonstrates that overexpression of ETS2 results in apoptosis. This is shown in a number of circumstances, including ETS2-overexpressing transgenic mice and cell lines and in cells from subjects with DS. Indeed we report for the first time that the ETS2 overexpression transgenic mouse develops a smaller thymus and lymphocyte abnormalities similar to that observed in DS. In all circumstances of ETS2 overexpression, the increased apoptosis correlated with increased p53 and alterations in downstream factors in the p53 pathway. In the human HeLa cancer cell line, transfection with functional p53 enables ETS2 overexpression to induce apoptosis. Furthermore, crossing the ETS2 transgenic mice with p53(-/-) mice genetically rescued the thymic apoptosis phenotype. Therefore, we conclude that overexpression of human chromosome 21-encoded ETS2 induces apoptosis that is dependent on p53. These results have important consequences for understanding DS and oncogenesis and may provide new insights into therapeutic interventions.

Animals↗
PubMed2003

Transformation induced by Ewing's sarcoma associated EWS/FLI-1 is suppressed by KRAB/FLI-1.

Ewing's sarcoma is a childhood bone tumour with poor prognosis, most commonly associated with a t(11;22)(q24;q12) reciprocal translocation that fuses the EWS and FLI-1 genes, resulting in the production of an aberrant chimeric transcription factor EWS/FLI-1. To elucidate the mechanisms by which EWS/FLI-1 mediates transformation in mouse models, we have generated a murine Ews/Fli-1 fusion protein. We demonstrate that this protein transforms fibroblast cells in vitro similar to human EWS/FLI-1 as demonstrated by serum and anchorage-independent growth, the formation of tumours in nude mice and elevation of the oncogenic marker c-myc. Furthermore, transformation of these cells was inhibited by a specific repressor, KRAB/FLI-1. The KRAB/FLI-1 repressor also suppressed the tumorigenic phenotype of a human Ewing's sarcoma cell line. These findings suggest that the transformed phenotype of Ewing's sarcoma cells can be reversed by using the sequence-specific FLI-1-DNA-binding domain to target a gene repressor domain. The inhibition of EWS/FLI-1 is the first demonstration of the KRAB domain suppressing the action of an ETS factor. This approach provides potential avenues for the elucidation of the biological mechanisms of EWS/FLI-1 oncogenesis and the development of novel therapeutic strategies.

3T3 Cells↗
PubMed2003

An altered antioxidant balance occurs in Down syndrome fetal organs: implications for the "gene dosage effect" hypothesis.

Down syndrome (DS) is the congenital birth defect responsible for the greatest number of individuals with mental retardation. It arises due to trisomy of human chromosome 21 (HSA21) or part thereof. To date there have been limited studies of HSA21 gene expression in trisomy 21 conceptuses. In this study we investigate the expression of the HSA21 antioxidant gene, Cu/Zn-superoxide dismutase-1 (SOD1) in various organs of control and DS aborted conceptuses. We show that SOD1 mRNA levels are elevated in DS brain, lung, heart and thymus. DS livers show decreased SOD1 mRNA expression compared with controls. Since non-HSA21 antioxidant genes are reported to be concomitantly upregulated in certain DS tissues, we examined the expression of glutathione peroxidase-1 (GPX1) in control and DS fetal organs. Interestingly, GPX1 expression was unchanged in the majority of DS organs and decreased in DS livers. We examined the SOD1 to GPX1 mRNA ratio in individual organs, as both enzymes form part of the body's defense against oxidative stress, and because a disproportionate increase of SOD1 to GPX1 results in noxious hydroxyl radical damage. All organs investigated show an approximately 2-fold increase in the SOD1 to GPX1 mRNA ratio. We propose that it is the altered antioxidant ratio that contributes to certain aspects of the DS phenotype.

Antioxidants↗
PubMed2003

Overexpression of the chromosome 21 transcription factor Ets2 induces neuronal apoptosis.

Down syndrome (trisomy 21) neurons display an increased rate of apoptosis in vitro. The genes on chromosome 21 that mediate this increased cell death remain to be elucidated. Here we show that the chromosome 21 transcription factor Ets2, a gene that is overexpressed in Down syndrome, is expressed in neurons, and that moderate overexpression of Ets2 leads to increased apoptosis of primary neuronal cultures from Ets2 tg mice that involves activation of caspase-3. Our data therefore suggest that overexpression of ETS2 may contribute to the increased rate of apoptosis of neurons in Down syndrome.

Animals↗
PubMed2001

Ets-2 is induced by oxidative stress and sensitizes cells to H(2)O(2)-induced apoptosis: implications for Down's syndrome.

An elevated production of hydrogen peroxide mediates the increased rate of apoptosis of cells derived from individuals with Down's syndrome. The mechanism via which this occurs is unknown. Here we show that Ets-2, a transcription factor located on human chromosome 21 and already overexpressed in multiple tissues in Down syndrome (DS, trisomy 21), is induced by low concentrations of hydrogen peroxide. Moreover, cells with an imbalance in the antioxidant enzymes SOD-1/GPX-1, such as occurs in DS through the overexpression of the chromosome 21 gene SOD-1, also results in increased Ets-2 expression. The increase in Ets-2 expression is dependent on mRNA transcription. Importantly, we further demonstrate that 3T3 fibroblasts that overexpress Ets-2 are sensitized to hydrogen peroxide-induced apoptosis. These data implicate Ets-2 in the regulation of oxidant-induced apoptosis and provide a possible rationale for both the (5- to 7-) fold increase in Ets-2 protein level in DS tissues, above the expected gene dosage of 1.5-fold, and the elevated rate of apoptosis in DS cells.

Animals↗
PubMed2001

BACE knockout mice are healthy despite lacking the primary beta-secretase activity in brain: implications for Alzheimer's disease therapeutics.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by accumulation of amyloid plaques and neurofibrillary tangles in the brain. The major components of plaque, beta-amyloid peptides (Abetas), are produced from amyloid precursor protein (APP) by the activity of beta- and gamma-secretases. beta-secretase activity cleaves APP to define the N-terminus of the Abeta1-x peptides and, therefore, has been a long- sought therapeutic target for treatment of AD. The gene encoding a beta-secretase for beta-site APP cleaving enzyme (BACE) was identified recently. However, it was not known whether BACE was the primary beta-secretase in mammalian brain nor whether inhibition of beta-secretase might have effects in mammals that would preclude its utility as a therapeutic target. In the work described herein, we generated two lines of BACE knockout mice and characterized them for pathology, beta-secretase activity and Abeta production. These mice appeared to develop normally and showed no consistent phenotypic differences from their wild-type littermates, including overall normal tissue morphology and brain histochemistry, normal blood and urine chemistries, normal blood-cell composition, and no overt behavioral and neuromuscular effects. Brain and primary cortical cultures from BACE knockout mice showed no detectable beta-secretase activity, and primary cortical cultures from BACE knockout mice produced much less Abeta from APP. The findings that BACE is the primary beta-secretase activity in brain and that loss of beta-secretase activity produces no profound phenotypic defects with a concomitant reduction in beta-amyloid peptide clearly indicate that BACE is an excellent therapeutic target for treatment of AD.

Alzheimer Disease↗
PubMed2001

The soluble murine type I interferon receptor Ifnar-2 is present in serum, is independently regulated, and has both agonistic and antagonistic properties.

The ability to modify responses to type I interferons (IFNs) could alter processes such as hematopoiesis and immunity, which involve endogenous IFNs and responses to exogenous IFNs. The data presented here support a significant role for a recently identified soluble isoform of the murine type I IFN receptor, muIfnar-2a, as an efficient regulator of IFN responses. The messenger RNA (mRNA) transcript encoding muIfnar-2a is generally more abundant than that encoding the transmembrane isoform, muIfnar-2c. Furthermore, the ratio of muIfnar-2a:2c transcripts varied from more than 10:1 in the liver and other organs to less than 1:1 in bone-marrow macrophages, indicating independent regulation of the 2 transcripts encoding receptor isoforms and suggesting that the soluble muIfnar-2a levels are biologically relevant in some organs. Western blot analysis showed that soluble muIfnar-2 was present at high levels in murine serum and other biologic fluids and bound type I IFN. Recombinant muIfnar-2a competitively inhibited the activity of both IFNalpha and beta in reporter assays using the L929 cell line and in antiproliferative and antiviral assays using primary cells. Surprisingly, using primary thymocytes from Ifnar-2(-/-) mice, recombinant muIfnar-2a formed a complex with IFN alpha or beta and muIfnar-1 at the cell surface and transmitted an antiproliferative signal. These data indicate potential dual actions of soluble muIfnar-2 and imply that a signal can be transduced through the Ifnar-1 chain of the receptor complex in the absence of the cytoplasmic domain of Ifnar-2. Therefore, our results suggest that soluble Ifnar-2 is an important regulator of endogenous and systemically administered type I IFN.

Age Factors↗
PubMed2001

Desrt, an AT-rich interaction domain family transcription factor gene, is an early marker for nephrogenic mesoderm and is expressed dynamically during mouse limb development.

Desrt is a mouse gene of the AT-rich interaction domain family of transcription factors. Here we describe the temporal and spatial pattern of expression of Desrt during mouse organogenesis. Desrt expression is first detected in the intermediate plate mesoderm, providing an early embryonic marker for this tissue, and subsequently in the nephrogenic cords of the urogenital ridges. A highly dynamic expression pattern is observed in the developing limb, implicating Desrt in limb patterning. Desrt is also detected in the myotome of the somites, the oro-naso-pharyngeal ectoderm and underlying mesenchyme, otic vesicles, the gut and its derivatives, and transiently in the liver.

Animals↗
PubMed2001

Increased infarct size and exacerbated apoptosis in the glutathione peroxidase-1 (Gpx-1) knockout mouse brain in response to ischemia/reperfusion injury.

Glutathione peroxidase is an antioxidant enzyme that is involved in the control of cellular oxidative state. Recently, unregulated oxidative state has been implicated as detrimental to neural cell viability and involved in both acute and chronic neurodegeneration. In this study we have addressed the importance of a functional glutathione peroxidase in a mouse ischemia/reperfusion model. Two hours of focal cerebral ischemia followed by 24 h of reperfusion was induced via the intraluminal suture method. Infarct volume was increased three-fold in the glutathione peroxidase-1 (Gpx-1) -/- mouse compared with the wild-type mouse; this was mirrored by an increase in the level of apoptosis found at 24 h in the Gpx-1 -/- mouse compared with the wild-type mouse. Neuronal deficit scores correlated to the histologic data. We also found that activated caspase-3 expression is present at an earlier time point in the Gpx-1 -/- mice when compared with the wild-type mice, which suggests an enhanced susceptibility to apoptosis in the Gpx-1 -/- mouse. This is the first known report of such a dramatic increase, both temporally and in level of apoptosis in a mouse stroke model. Our results suggest that Gpx-1 plays an important regulatory role in the protection of neural cells in response to the extreme oxidative stress that is released during ischemia/reperfusion injury.

Animals↗
PubMed2001

Gene targeting of Desrt, a novel ARID class DNA-binding protein, causes growth retardation and abnormal development of reproductive organs.

We have cloned and characterized a novel murine DNA-binding protein Desrt, with a motif characteristic of the ARID (A-T rich interaction domain) family of transcription factors. The Desrt gene encodes an 83-kD protein that is shown to bind DNA and is widely expressed in adult tissues. To examine the in vivo function of Desrt, we have generated mice with a targeted mutation in the ARID domain of Desrt. Homozygous mutants have reduced viability, pronounced growth retardation, and a high incidence of abnormalities of the female and male reproductive organs including cryptorchidism. This may thus serve as a model to dissect the mechanisms involved in the development of the reproductive tract including testicular descent. Gene-targeted mice also display a reduction in the thickness of the zona reticularis of the adrenal gland and transient aberrations of the T and B cell compartments of primary lymphoid organs. These data show that this novel DNA-binding protein, Desrt, has a nonredundant function during growth and in the development of the reproductive system.

AT Rich Sequence↗
PubMed2001

Identification and characterization of alternatively spliced murine Rgs11 isoforms: genomic structure and gene analysis.

The RGS proteins comprise a large family of proteins which were recently identified as negative Regulators of G-protein Signaling. They have been shown to act as GTPase Activating Proteins (GAPs) towards the G(alpha) subunits of heterotrimeric G-proteins. In addition to this GAP activity, which has been shown to occur through the RGS domain, RGS proteins are likely to possess other functions due to the existence of other domains in these molecules (De Vries and Farquhar, 1999; Hepler, 1999). Here, we report the molecular characterization of the murine Rgs11 gene. The gene encodes a protein with high homology to human RGS11 (79.9%), containing conserved DEP (Dishevelled/EGL-10/Pleckstrin) and GGL (G protein gamma-like) domains. The gene is comprised of at least 13 exons, spanning 8-9 kb. Spliced transcript variants were identified which are co-expressed with 5A3, a transcript that contains the largest ORF. Expression of mouse Rgs11 was found to be restricted to specific tissues with a unique pattern of expression observed in brain.

Alternative Splicing↗
PubMed2001

A novel member of the STOMATIN/EPB72/mec-2 family, stomatin-like 2 (STOML2), is ubiquitously expressed and localizes to HSA chromosome 9p13.1.

A cDNA encoding a novel second member of the Band7/stomatin-like/SPFH domain family in humans designated stomatin-like 2 (STOML2) has been isolated using the technique of cDNA Representational Difference Analysis. The STOML2 cDNA encoded a 356 amino acid residue polypeptide with a predicted molecular weight of 38.5 kDa. The predicted polypeptide sequence of STOML2 could be delineated into three major domains: an N-terminal alpha-helical region; a domain with significant similarity to a 172 amino acid region of the HSA stomatin polypeptide, composed of an alternating alpha-helical and beta-sheet structure and a C-terminal domain that was mostly alpha-helical. The stomatin-like domain was observed in 51 other proteins with potentially diverse functions. Based on its homology to stomatin, STOML2 was predicted to be cytoplasmically located. However, unlike most of the other proteins containing stomatin-like domains, the predicted STOML2 polypeptide does not contain a transmembrane region although the presence of N-myristoylation sites suggest that it has the potential to be membrane-associated. Northern blot analysis of a panel of poly(A)(+) mRNA from normal human adult tissues showed that a single 1.3-kb mRNA transcript encoding STOML2 was ubiquitously expressed, with relatively higher levels in skeletal muscle and heart compared to other tissues. Comparison of the STOML2 cDNA sequence with human genomic DNA indicated that the gene encoding STOML2 was 3,250 bp long and consisted of ten exons interrupted by nine introns. We have mapped STOML2 to HSA chromosome 9p13.1, a region that is rearranged in some cancers and thought to contain the gene responsible for acromesomelic dysplasia.

Adult↗
PubMed2000

Phenotypic characterization of an alpha 4 neuronal nicotinic acetylcholine receptor subunit knock-out mouse.

Neuronal nicotinic acetylcholine receptors (nAChR) are present in high abundance in the nervous system (Decker et al., 1995). There are a large number of subunits expressed in the brain that combine to form multimeric functional receptors. We have generated an alpha(4) nAChR subunit knock-out line and focus on defining the behavioral role of this receptor subunit. Homozygous mutant mice (Mt) are normal in size, fertility, and home-cage behavior. Spontaneous unconditioned motor behavior revealed an ethogram characterized by significant increases in several topographies of exploratory behavior in Mt relative to wild-type mice (Wt) over the course of habituation to a novel environment. Furthermore, the behavior of Mt in the elevated plus-maze assay was consistent with increased basal levels of anxiety. In response to nicotine, Wt exhibited early reductions in a number of behavioral topographies, under both unhabituated and habituated conditions; conversely, heightened levels of behavioral topographies in Mt were reduced by nicotine in the late phase of the unhabituated condition. Ligand autoradiography confirmed the lack of high-affinity binding to radiolabeled nicotine, cytisine, and epibatidine in the thalamus, cortex, and caudate putamen, although binding to a number of discrete nuclei remained. The study confirms the pivotal role played by the alpha(4) nAChR subunit in the modulation of a number of constituents of the normal mouse ethogram and in anxiety as assessed using the plus-maze. Furthermore, the response of Mt to nicotine administration suggests that persistent nicotine binding sites in the habenulo-interpeduncular system are sufficient to modulate motor activity in actively exploring mice.

Aggression↗