Chromosome loops, insulators, and histone methylation: new insights into regulation of imprinting in clusters.
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Animals↗
Biomedical subjects
K Mitsuya
Publications and source records attributed to K Mitsuya.
Epigenetic heterogeneity at imprinted loci in normal populations.
Genomic imprinting is the phenomenon by which the two alleles of certain genes are differentially expressed according to their parental origin. Extensive analysis of allelic expression at multiple imprinted loci in a normal population has not performed so far. In the present study, we examined the allelic expression pattern of three imprinted genes in a panel of 262 Japanese normal individuals. We observed differences in the extent of maintenance of allele-specific expression of the three genes. The allelic expression of small nuclear ribonucleoprotein N (SNRPN) was stringently regulated while that of multimembrane-spanning polyspecific transporter-like gene 1 (IMPT1) showed a large degree of variation. Significant biallelic expression of insulin-like growth factor II (IGF2) was observed in about 10% of normal individuals. Our findings add to the accumulating evidence for variable allelic expression at multiple loci in a normal human population. This epigenetic heterogeneity can be a stable trait and potentially influence individual phenotypes.
Alleles↗
Large-scale evaluation of imprinting status in the Prader-Willi syndrome region: an imprinted direct repeat cluster resembling small nucleolar RNA genes.
Loss of paternal gene expression at the imprinted domain on proximal human chromosome 15 causes Prader-Willi syndrome (PWS), a complex multiple-anomaly disorder involving variable mental retardation, hyperphasia leading to obesity and infantile hypotonia with failure to thrive. Although numerous paternally expressed transcripts have been identified that reside in the candidate region, the individual contributions to the development of PWS have not been firmly established. Recent studies of mouse models carrying a cytogenetic deletion suggest that paternal deficiency of the SNRPN-IPW interval is critical for perinatal lethality of potential relevance to PWS. Here we determined the allelic expression profiles of a total of 118 cDNA clones using monochromosomal hybrids retaining either a paternal or maternal human chromosome 15. Our results demonstrated a preponderance of unusual transcripts lacking protein-coding potential that were expressed exclusively from the paternal copy of the critical interval. This interval was also found to encompass a large direct repeat (DR) cluster displaying a potentially active chromatin conformation of paternal origin, as suggested by enhanced sensitivity to nuclease digestion. Database searches revealed an unexpected organization of tandemly repeated consensus elements, all of which possessed well-defined box C and D sequences characteristic of small nucleolar RNAs (snoRNAs). Southern blot analysis further demonstrated a considerable degree of phylogenetic conservation of the DR locus in the genomes of all mammalian species tested, but not in chicken, Xenopus and Drosophila. These findings imply a potential direct contribution of the DR locus, representing a cluster of multiple snoRNA genes, to certain phenotypic features of PWS.
Base Sequence↗
Construction of 700 human/mouse A9 monochromosomal hybrids and analysis of imprinted genes on human chromosome 6.
As an in vitro assay system for the identification of human imprinted genes, a library of human/mouse A9 monochromosomal hybrids containing a single, intact bsr-tagged human chromosome of known parental origin, derived from normal human fibroblasts, has been previously generated by microcell-mediated chromosome transfer (MMCT). To supplement this assay system, we constructed additional 700 A9 monochromosomal hybrids, using a pSTneo or pPGKneo selection marker. To validate the A9 hybrids, we screened them with chromosome-specific polymorphic markers, and identified the hybrids containing either human chromosome 6, 7, 14, 18, or 21 of known parental origin. Matching paternal and maternal chromosome pairs of A9 hybrids were identified for chromosomes 6, 7, 14, and 18. The paternal-specific expression of ZAC (zinc finger protein, which regulates apoptosis and cell cycle arrest) and HYMAI (hydatidiform mole-associated and imprinted transcript), and the maternal-specific methylation of a CpG island within an imprinted domain on human chromosome 6q24, were maintained in A9 hybrids. For an example, we profiled the expression of expressed sequence tags (ESTs) and the methylation of CpG islands in the 300-kb imprinted domain around 6q24, which may be associated with cancers and transient neonatal diabetes mellitus (TNDM). Thus, the 700 A9 hybrids should be useful for various aspects of imprinting studies.
Animals↗
A novel maternally expressed gene, ATP10C, encodes a putative aminophospholipid translocase associated with Angelman syndrome.
Lack of a maternal contribution to the genome at the imprinted domain on proximal chromosome 15 causes Angelman syndrome (AS) associated with neurobehavioral anomalies that include severe mental retardation, ataxia and epilepsy. Although AS patients have infrequent mutations in the gene encoding an E6-AP ubiquitin ligase required for long-term synaptic potentiation (LTP), most cases are attributed to de novo maternal deletions of 15q11-q13. We report here that a novel maternally expressed gene, ATP10C, maps within the most common interval of deletion and that ATP10C expression is virtually absent from AS patients with imprinting mutations, as well as from patients with maternal deletions of 15q11-q13.
Adenosine Triphosphatases↗
Comprehensive analysis of 28 patients with latex allergy and prevalence of latex sensitization among hospital personnel.
Recently anaphylactic shock caused by latex gloves and medical instruments has been discussed as an important problem in surgical operations. Patients with contact urticaria or anaphylaxis due to natural rubber latex were first reported in Japan in 1993, and the number of cases is gradually increasing. In the present study, we analyzed 28 patients examined in our clinic from 1993 to 1998. The diagnosis of latex allergy was made on the basis of clinical history, latex specific IgE antibody, skin test, and use test. The 3 male and 25 female patients included 20 nurses, 4 doctors, 2 housewives, one animal hospital employee, and one worker in a senile rehabilitation center. The majority were health care workers. Contact urticaria from rubber gloves was the most common clinical symptom. Some of the patients developed severe attacks of anaphylaxis. During the period from 1995 to 1997, we also performed a questionnaire study and a serum examination of latex specific IgE antibody among the personnel of our university hospital. The screening test for the antibody was more reliable than our questionnaire study in detecting latex-sensitive persons. The prevalence of latex allergy was found to be 4.6%. This indicates that not only dermatologists but also all hospital workers should be aware of this type of allergy.
Adult↗
Loss of imprinting of long QT intronic transcript 1 in colorectal cancer.
Loss of imprinting (LOI) of the insulin-like growth factor 2 (IGF2) and H19 genes on human chromosome 11 has been found not only in childhood tumors but also in common adult cancers including colorectal cancer. Recently, a transcript called LIT1 (long QT intronic transcript 1) has been identified within the KvLQT1 locus on chromosome 11. LIT1 is expressed preferentially from the paternal allele and is transcribed in most human tissues. LOI of LIT1 was found in a considerable number of Beckwith-Wiedemann syndrome (BWS) patients, suggesting that it is associated with the etiology of BWS. Since LOI of IGF2 was observed in association with overexpression of IGF2 in colorectal cancer in our previous study, we examined the status of genomic imprinting of LIT1 and H19 in comparison with IGF2 in colorectal cancer. We examined 44 surgically dissected colorectal cancer tissues. Ten of them represented informative cases for LIT1. None of these patients exhibited loss of heterozygosity (LOH) of LIT1, and LOI of LIT1 was observed in 4 of the 10 (40%) informative patients, but not in non-cancerous tissues. Neither LOH nor LOI of H19 was observed. LOI of IGF2 was observed in 4 of 18 (22%) informative patients. These results suggest that LOI of LIT1 is frequently observed in colorectal cancer and may be a useful marker for diagnosis of colorectal cancer.
Chromosomes, Human, Pair 11↗
Tentorial enhancement on MR images is a sign of cavernous sinus involvement in patients with sellar tumors.
BACKGROUND AND PURPOSE: This study was undertaken to analyze enhancement patterns of the dura around sellar tumors and to compare the results with tumor invasion or compression of the cavernous sinuses. Postoperative enhancement patterns on MR images were compared with preoperative findings. METHODS: Contrast-enhanced coronal and sagittal MR images were examined prospectively in 96 patients with sellar tumors (65 macroadenomas, 15 microadenomas, 14 Rathke cleft cysts, and two chordomas at the sella). All patients underwent surgical treatment, and pre- and postsurgical features on MR images were compared. RESULTS: Presurgical MR images showed dural enhancement in 36.5% of the patients: asymmetric tentorial enhancement in 24 patients, symmetric tentorial enhancement in seven, and sphenoidal ridge or clivus enhancement in four. Asymmetric tentorial enhancement disappeared after surgical decompression in seven patients. For evaluation of cavernous sinus invasion ipsilateral to the enhancement, sensitivity and specificity of the asymmetric tentorial enhancement sign were 81.3% and 86.3%, respectively. Sensitivity and specificity of the sign were 42.9% and 93.6% for cavernous sinus involvement, including compression and invasion. CONCLUSION: Asymmetric tentorial enhancement is a useful sign in the diagnosis of invasion or severe compression of the cavernous sinus by sellar tumor. The sign may represent venous congestion or collateral flow in the tentorium due to obstructed flow in the medial portion of the cavernous sinus.
Adenoma↗
Targeted disruption of the human LIT1 locus defines a putative imprinting control element playing an essential role in Beckwith-Wiedemann syndrome.
Human chromosome 11p15.5 harbors an intriguing imprinted gene cluster of 1 Mb. This imprinted domain is implicated in a wide variety of malignancies and Beckwith-Wiedemann syndrome (BWS). Recently, several lines of evidence have suggested that the BWS-associated imprinting cluster consists of separate chromosomal domains. We have previously identified LIT1, a paternally expressed antisense RNA within the KvLQT1 locus through a positional screening approach using human monochromosomal hybrids. KvLQT1 encompasses the translocation breakpoint cluster in BWS and patients exhibit frequent loss of maternal methylation at the LIT1 CpG island, implying a regulatory role for the LIT1 locus in coordinate control of the imprinting cluster. Here we generated modified human chromosomes carrying a targeted deletion of the LIT1 CpG island using recombination-proficient chicken DT40 cells. Consistent with the prediction, this mutation abolished LIT1 expression on the paternal chromosome, accompanied by activation of the normally silent paternal alleles of multiple imprinted loci at the centromeric domain including KvLQT1 and p57(KIP2). The deletion had no effect on imprinting of H19 located at the telomeric end of the cluster. Our findings demonstrate that the LIT1 CpG island can act as a negative regulator in cis for coordinate imprinting at the centromeric domain, thereby suggesting a role for the LIT1 locus in a BWS pathway leading to functional inactivation of p57(KIP2). Thus, the targeting and precise modification of human chromosomal alleles using the DT40 cell shuttle system can be used to define regulatory elements that confer long-range control of gene activity within chromosomal domains.
Alleles↗
Osteochondroma of the pubic symphysis associated with sexual disturbance.
Osteochondroma of the pubic symphysis is a rare benign skeletal tumor. We report here a case of an osteochondroma of the pubic symphysis associated with a sexual disturbance, where a computed tomography scan clearly showed a tumor lesion of the pubic symphysis. The case is reported not only because of its rarity but also because it is important that gynecologists should bear this disease in mind, since a patient with this tumor may not visit an orthopedist but a gynecologist.
Adult↗
Multipoint imprinting analysis in sporadic colorectal cancers with and without microsatellite instability.
Disrupted imprinting is implicated in certain tumorigenesis. Since aberrant methylation has been described for a majority of microsatellite instability (MSI)-positive sporadic colorectal cancers, we have investigated alteration to the imprinting in 55 sporadic colorectal cancers with or without MSI. Loss of imprinting (LOI) of IGF2 and PEG1/MEST was observed in 42% and 35% of informative cancers, respectively. H19 expression was not detected in 24% of informative cancers. SNRPN and NDN retained monoallelic expression in all the cancers examined. These findings indicate no simultaneous disruption of the imprinted genes. LOI of IGF2 and PEG1/MEST was also observed in colorectal mucosa from almost all the patients with LOI in tumor tissue. Moreover, MSI-positive colorectal cancers exhibit LOI of IGF2 with a high frequency compared to MSI-negative cancers (P=0.013). These observations, consistent with a previous report, establish an association between LOI of IGF2 and MSI in colorectal cancers and provide insight into susceptibility of tumor development.
Alleles↗
Mouse A9 cells containing single human chromosomes for analysis of genomic imprinting.
To develop an systematic in vitro approach for the study of genomic imprinting, we generated a new library of human/mouse A9 monochromosomal hybrids. We used whole cell fusion and microcell-mediated chromosome transfer to generate A9 hybrids containing a single, intact, bsr-tagged human chromosome derived from primary fibroblasts. A9 hybrids were identified that contained either human chromosome 1, 2, 4, 5, 7, 8, 10, 11, 15, 18, 20, or X. The parental origin of these chromosomes was determined by polymorphic analysis using microsatellite markers, and matched hybrids containing maternal and paternal chromosomes were identified for chromosomes 5, 10, 11 and 15. The imprinted gene KVLQT1 on human chromosome 11p15.5 was expressed exclusively from the maternal chromosome in A9 hybrids, and the parental-origin-specific expression patterns of several other imprinted genes were also maintained. This library of human monochromosomal hybrids is a valuable resource for the mapping and cloning of human genes and is a novel in vitro system for the screening of imprinted genes and for their functional analysis.
Animals↗
Loss of imprinting of a paternally expressed transcript, with antisense orientation to KVLQT1, occurs frequently in Beckwith-Wiedemann syndrome and is independent of insulin-like growth factor II imprinting.
Genomic imprinting plays a fundamental role in cancer and some hereditary diseases, including Beckwith-Wiedemann syndrome (BWS), a disorder of prenatal overgrowth and predisposition to embryonal malignancies such as Wilms tumor. We have previously shown that the KVLQT1 gene on chromosomal band 11p15 is imprinted, with expression of the maternal allele, and that the maternal allele is disrupted in rare BWS patients with balanced germ-line chromosomal rearrangements. We now show that an antisense orientation transcript within KVLQT1, termed LIT1 (long QT intronic transcript 1) is expressed normally from the paternal allele, from which KVLQT1 transcription is silent, and that in the majority of patients with BWS, LIT1 is abnormally expressed from both the paternal and maternal alleles. Eight of sixteen informative BWS patients (50%) showed biallelic expression, i.e., loss of imprinting (LOI) of LIT1. Similarly, 21 of 36 (58%) BWS patients showed loss of maternal allele-specific methylation of a CpG island upstream of LIT1. Surprisingly, LOI of LIT1 was not linked to LOI of insulin-like growth factor II (IGF2), which was found in 2 of 10 (20%) BWS patients, even though LOI of IGF2 occurs frequently in Wilms and other tumors, and in some patients with BWS. Thus, LOI of LIT1 is the most common genetic alteration in BWS. We propose that 11p15 harbors two imprinted gene domains-a more centromeric domain including KVLQT1 and p57(KIP2), alterations in which are more common in BWS, and a more telomeric domain including IGF2, alterations in which are more common in cancer.
Beckwith-Wiedemann Syndrome↗
LIT1, an imprinted antisense RNA in the human KvLQT1 locus identified by screening for differentially expressed transcripts using monochromosomal hybrids.
Mammalian imprinted genes are frequently arranged in clusters on particular chromosomes. The imprinting cluster on human chromosome 11p15 is associated with Beckwith-Wiedemann syndrome (BWS) and a variety of human cancers. To clarify the genomic organization of the imprinted cluster, an extensive screen for differentially expressed transcripts in the 11p15 region was performed using monochromosomal hybrids with a paternal or maternal human chromosome 11. Here we describe an imprinted antisense transcript identified within the KvLQT1 locus, which is associated with multiple balanced chromosomal rearrangements in BWS and an additional breakpoint in embryonal rhabdoid tumors. The transcript, called LIT1 (long QT intronic transcript 1), was expressed preferentially from the paternal allele and produced in most human tissues. Methylation analysis revealed that an intronic CpG island was specifically methylated on the silent maternal allele and that four of 13 BWS patients showed complete loss of maternal methylation at the CpG island, suggesting that antisense regulation is involved in the development of human disease. In addition, we found that eight of eight Wilms' tumors exhibited normal imprinting of LIT1 and five of five tumors displayed normal differential methylation at the intronic CpG island. This contrasts with five of six tumors showing loss of imprinting of IGF2. We conclude that the imprinted gene domain at the KvLQT1 locus is discordantly regulated in cancer from the imprinted domain at the IGF2 locus. Thus, this positional approach using human monochromosomal hybrids could contribute to the efficient identification of imprinted loci in humans.
Animals↗
Changes in platelet ATP secretion and aggregation during pregnancy and in preeclampsia.
BACKGROUND: Platelet secretion plays an important role in the aggregation of platelets. However, the quantitative relationship between platelet aggregation and secretion of ATP during pregnancy and in pre-eclampsia has yet to be clarified. This study is designed to determine whether platelet count, volume, aggregation, and the amount of secreted ATP change in healthy, nonpregnant women, nonpreeclamptic pregnant women, and preeclamptic pregnant women and whether beta-thromboglobulin (BTG) and platelet factor 4 (PF-4) concentrations alter in nonpreeclamptic and pre-eclamptic women. METHODS: Peripheral blood was collected from 114 women. Nonpreeclamptic pregnant women were divided into four groups (gestational weeks 10, 20, 30, and 35). Platelet aggregation and ATP secretion were investigated with the use of a lumi-aggregometer. BTG and PF-4 concentrations in peripheral blood were determined in 12 pregnant and 11 preeclamptic women. RESULTS: The amount of secreted ATP upon induction by 5 microM ADP increased significantly (P < 0.05-0.01) with gestational age. On the other hand, the amount of secreted ATP induced by 5 microg/mL collagen reached the maximal value from gestational weeks 20 to 35 in nonpreeclamptic women. Significantly more platelet aggregation was induced by the ADP and collagen in nonpreeclamptic women in gestational weeks 20 and 30 than in the gestational weeks 10 or 35 (P < 0.05-0.005). The amount of secreted ATP and platelet count were significantly lower (P < 0.05) in preeclampsia than in normal pregnancy. The BTG and PF-4 concentrations were significantly higher (P < 0.05) in preeclampsia than in normal pregnancy. CONCLUSIONS: The sensitivity of platelets for ATP secretion may intensify with progression of pregnancy. In normal pregnancy, around gestational week 35, the platelets may exhibit weaker ability to aggregate but maintain the capacity to secrete ATP. In preeclampsia, secreted ATP decreased because platelets may be stimulated to undergo a partial secretion.
Adenosine Triphosphate↗
Aspirin-sensitive urticaria: provocation with a leukotriene receptor antagonist.
The diagnosis of two patients with aspirin-induced urticaria (AIU) was confirmed by oral provocation with aspirin, other non-steroidal anti-inflammatory drugs, and food additives. Low doses of a novel leukotriene (LT) receptor antagonist, ONO-1078 (ONO, Japan) induced urticaria in these patients, while the same doses of ONO-1078 did not provoke any eruptions in 10 normal healthy volunteers or five patients with aspirin-unrelated chronic urticaria. This is the first report that a selective LTD4/LTE4 receptor antagonist that is effective as an antiasthmatic agent evoked urticaria in patients with AIU. Our observation suggests that the pathogenesis of AIU depends on the stimulation of LT receptors. The accumulation of results of anti-LT therapy may provide clues to resolve the pathogenetic mechanisms of this disorder.
Administration, Oral↗
Epigenetic reprogramming of the human H19 gene in mouse embryonic cells does not erase the primary parental imprint.
BACKGROUND: Genomic imprinting in mammals is thought to result from epigenetic modifications to chromosomes during gametogenesis, which leads to differential allelic expression during development. There is a requirement for an appropriate experimental system to enable the analysis of the mechanisms of genomic imprinting during embryogenesis. RESULTS: To develop a novel in vitro system for studying the molecular basis of genomic imprinting, we constructed mouse cell lines containing either a paternal or maternal human chromosome 11, by microcell-mediated chromosome transfer. Allele-specific expression and DNA methylation studies revealed that the imprinting status of the human H19 gene was maintained in mouse A9 mono-chromosomal hybrids. Each parental human chromosome was introduced independently into mouse near-diploid immortal fibroblasts (m5S) and two embryonal carcinoma (EC) cell lines (OTF9-63 and P19). The paternal allele of human H19 remained in a repressed state in m5S cells, but was de-repressed in both EC cells. The paternal H19 allele was demethylated extensively in OTF9-63 cells, whereas the only alteration in P19 hybrids was de novo methylation on both alleles in the 3' region. Following in vitro differentiation, the expressed paternal H19 allele was selectively repressed in differentiated derivatives of EC hybrids. CONCLUSION: These results indicated that human imprint marks could function effectively in mouse cells, and that the imprinting process was epigenetically reprogrammed in embryonal carcinoma cells, without erasure of the primary imprint that marked the parental origin. Therefore, these mono-chromosomal hybrids could provide a valuable in vitro system to study the mechanisms involved in the regulation of imprinted gene expression.
Animals↗
[Studies of genomic imprinting using mouse cells containing a human chromosome].
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Animals↗