Structure and Engineering of Francisella novicida Cas9.

The RNA-guided endonuclease Cas9 cleaves double-stranded DNA targets complementary to the guide RNA and has been applied to programmable genome editing. Cas9-mediated cleavage requires a protospacer adjacent motif (PAM) juxtaposed with the DNA target sequence, thus constricting the range of targetable sites. Here, we report the 1.7 Å resolution crystal structures of Cas9 from Francisella novicida (FnCas9), one of the largest Cas9 orthologs, in complex with a guide RNA and its PAM-containing DNA targets. A structural comparison of FnCas9 with other Cas9 orthologs revealed striking conserved and divergent features among distantly related CRISPR-Cas9 systems. We found that FnCas9 recognizes the 5'-NGG-3' PAM, and used the structural information to create a variant that can recognize the more relaxed 5'-YG-3' PAM. Furthermore, we demonstrated that the FnCas9-ribonucleoprotein complex can be microinjected into mouse zygotes to edit endogenous sites with the 5'-YG-3' PAM, thus expanding the target space of the CRISPR-Cas9 toolbox.

A Lipid Nanoparticle-Based Method for the Generation of Liver-Specific Knockout Mice.

Knockout mice are useful tools that can provide information about the normal function of genes, including their biochemical, developmental, and physiological roles. One problem associated with the generation of knockout mice is that the loss of some genes of interest produces a lethal phenotype. Therefore, the use of conditioned knockout mice, in which genes are disrupted in specific organs, is essential for the elucidation of disease pathogenesis and the verification of drug targets. In general, conditional knockout mice are produced using the Cre/loxP system; however, the production of the large numbers of Cre/flox knockout and control mice required for analysis requires substantial time and effort. Here, we describe the generation of liver-specific conditional knockout mice via the introduction of lipid nanoparticles encapsulating Cre mRNA into the liver of floxed mice. This technique does not require the production of offspring by mating floxed mice and is therefore more convenient than the conventional method. The results presented here demonstrate that the LNP-based method enables liver-specific gene knockout in a short period of time.

DNA methylation analysis of multiple imprinted DMRs in Sotos syndrome reveals IGF2-DMR0 as a DNA methylation-dependent, P0 promoter-specific enhancer.

Haploinsufficiency of NSD1, which dimethylates histone H3 lysine 36 (H3K36), causes Sotos syndrome (SoS), an overgrowth syndrome. DNMT3A and DNMT3B recognizes H3K36 trimethylation (H3K36me3) through PWWP domain to exert de novo DNA methyltransferase activity and establish imprinted differentially methylated regions (DMRs). Since decrease of H3K36me3 and genome-wide DNA hypomethylation in SoS were observed, hypomethylation of imprinted DMRs in SoS was suggested. We explored DNA methylation status of 28 imprinted DMRs in 31 SoS patients with NSD1 defect and found that hypomethylation of IGF2-DMR0 and IG-DMR in a substantial proportion of SoS patients. Luciferase assay revealed that IGF2-DMR0 enhanced transcription from the IGF2 P0 promoter but not the P3 and P4 promoters. Chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) revealed active enhancer histone modifications at IGF2-DMR0, with high enrichment of H3K4me1 and H3 lysine 27 acetylation (H3K27ac). CRISPR-Cas9 epigenome editing revealed that specifically induced hypomethylation at IGF2-DMR0 increased transcription from the P0 promoter but not the P3 and P4 promoters. NSD1 knockdown suggested that NSD1 targeted IGF2-DMR0; however, IGF2-DMR0 DNA methylation and IGF2 expression were unaltered. This study could elucidate the function of IGF2-DMR0 as a DNA methylation dependent, P0 promoter-specific enhancer. NSD1 may play a role in the establishment or maintenance of IGF2-DMR0 methylation during the postimplantation period.

Synergistic Upregulation of Target Genes by TET1 and VP64 in the dCas9-SunTag Platform.

Overexpression of a gene of interest is a general approach used in both basic research and therapeutic applications. However, the conventional approach involving overexpression of exogenous genes has difficulty achieving complete genome coverage, and is also limited by the cloning capacity of viral vectors. Therefore, an alternative approach would be to drive the expression of an endogenous gene using an artificial transcriptional activator. Fusion proteins of dCas9 and a transcription activation domain, such as dCas9-VP64, are widely used for activation of endogenous genes. However, when using a single sgRNA, the activation range is low. Consequently, tiling of several sgRNAs is required for robust transcriptional activation. Here we describe the screening of factors that exhibit the best synergistic activation of gene expression with TET1 in the dCas9-SunTag format. All seven factors examined showed some synergy with TET1. Among them, VP64 gave the best results. Thus, simultaneous tethering of VP64 and TET1 to a target gene using an optimized dCas9-SunTag format synergistically activates gene expression using a single sgRNA.

[Mail-based intervention for improvement of dietary habits among community-dwelling older adults living in heavy snowfall regions].

Objectives　The aim of this study was to investigate the effect of mail-based intervention using the TAKE10! Program to improve dietary habits in cases where direct intervention is not possible.Methods　Subjects aged 70-91 years (77.6±5.0) were randomly assigned to two groups: 72 in the intervention group and 71 in the control group. The intervention group received monthly mail, which included self-check sheets (TAKE10! Check sheet and TAKE10! Calendar) and a letter with feedback and comments for 5 months. The outcome measures were changes in the intake frequency of 10 food groups, Dietary Variety Score (DVS), and Food Frequency Score (FFS).Results　Compared to baseline, the post-intervention intake frequencies for 9 of 10 food groups, DVS, and FFS significantly increased in the intervention group. No significant differences were observed between baseline and post-intervention in the control group. In the subgroup analysis of the intervention group, post-intervention DVS and FFS of both subjects who cooked their own food and those who did not showed significant increases compared to baseline.Conclusion　The mail-based TAKE10! Program resulted in improved dietary habits and could be shared with families in addition to direct interventions and could also be used in regions with inadequate transportation systems or frequent poor weather conditions.

Arid1b Haploinsufficiency Causes Abnormal Brain Gene Expression and Autism-Related Behaviors in Mice.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder with core symptoms that include poor social communication, restricted interests, and repetitive behaviors. Several ASD mouse models exhibit impaired social interaction, anxiety-like behavior, and elevated perseveration. Large-scale whole exome sequencing studies identified many genes putatively associated with ASD. Like chromodomain helicase DNA binding protein 8 (CHD8), the most frequently mutated gene in individuals with ASD, the candidate gene AT-rich interaction domain 1B (ARID1B) encodes a chromatin remodeling factor. Arid1b heterozygous knockout (hKO) mice exhibited ASD-like traits related to social behavior, anxiety, and perseveration, in addition to associated features reported in some cases of ASD, such as reduced weight, impaired motor coordination, and hydrocephalus. Hydrocephalus was present in 5 of 91 hKO mice, while it was not observed in wild-type littermates (0 of 188). Genome-wide gene expression patterns in Arid1b hKO mice were similar to those in ASD patients and Chd8-haploinsufficient mice, an ASD model, and to developmental changes in gene expression in fast-spiking cells in the mouse brain. Our results suggest that Arid1b haploinsufficiency causes ASD-like phenotypes in mice.

Evaluation of the comprehensive health program "Sumida TAKE10!" for community-dwelling older adults, which aims to prevent or delay the need for long-term nursing care.

Objective The purpose of this study was to evaluate the comprehensive health program "Sumida TAKE10!", which aims to improve dietary habits and promote physical activity among community-dwelling older adults including the pre-frail elderly. This study has been ongoing since 2005 in Sumida Ward, Tokyo with the ultimate aim of preventing or delaying the need for long-term nursing care. We used the term "pre-frail elderly" for older adults who are at risk of requiring long-term care.Methods "Sumida TAKE10!" consists of a general lecture in a public hall followed by 5 educational sessions biweekly at 4-6 community centers. From 2008 to 2013, 402 participants aged ≥65 years were enrolled and included as subjects of the study. The main outcome measures were changes in 10 food intake frequencies, food frequency score (FFS), dietary variety score (DVS), frequency of exercise (obtained via questionnaire) and physical fitness (5-meter maximal walking time, 5-meter walking time, handgrip strength, one-leg standing time with eyes opened (time to upright posture for standing on one leg with eyes open), and the timed up & go test). The secondary outcome measures were changes in the Tokyo Metropolitan Institute of Gerontology (TMIG) Index of Competence score, appetite, frequency of walking and sports, self-rated health, frequency of leaving the house, communication with neighbors, engagement in hobbies, participation in group activities and participation in volunteer activities (obtained via questionnaire).Results Compared to baseline, all outcomes showed significant improvement. "Sumida TAKE10!" can improve dietary habits and increase the physical activity of participants. Positive secondary effects were seen for life function, self-rated health, and social activities. Almost identical positive results were obtained from the pre-frail elderly group, while improvement was also seen in the dietary habits of the subjects who do not cook.Conclusion These results suggest that this program may be useful for population-based approach programs. In addition, comprehensive programs like TAKE10! may increase the health consciousness of community-dwelling older adults.

Associations of eating out and dietary diversity with mild cognitive impairment among in community-dwelling older adults.

Background: Dementia is a critical later life health issue that occurs among members of aging societies. This study examined the relationships between eating out, dietary diversity, and mild cognitive impairment (MCI) among community-dwelling older adults. Methods: We analyzed data from 597 older adults (median age 73.0 years [interquartile range 69.0-78.0] and 62.6% females). We applied the Food Frequency Score (FFS) to evaluate diet variety and the weekly consumption frequencies of ten food items were determined. The Functional Assessment Tool from the National Center for Geriatrics and Gerontology was used to evaluate MCI. Finally, we asked the participants how often they ate out each month; those who replied 'none' were categorized into the "non-eating out" group. Results: The overall prevalence of MCI was 122 (20.4%), with a higher prevalence in the low dietary diversity group than in the high dietary diversity group (28.6% vs. 18.6%). After adjusting for covariates, the participants who self-described as not eating out were independently associated with low dietary diversity (odds ratio [OR]: 1.97, 95% confidence interval [CI]: 1.20-3.20), while low dietary diversity was associated with MCI (OR: 1.72, 95% CI: 1.02-2.87). Structural equation models revealed that not eating out had no direct effect on MCI but was associated with MCI via low dietary diversity (root mean square error of approximation = 0.030, goodness-of-fit index=0.999, and adjusted goodness-of-fit index = 0.984). Conclusions: Although non-eating out may not have a direct effect on MCI, an indirect relationship may exist between eating-out habits and MCI via dietary diversity status.

[11C]PK11195 PET imaging of spinal glial activation after nerve injury in rats.

The role of glial activation has been implicated in the development and persistence of neuropathic pain after nerve injury by recent studies. PK11195 binding to the translocator protein 18kDa (TSPO) has been shown to be enhanced in activated microglia. This study was designed to assess PK11195 imaging in spinal microglia during activation after nerve injury. The development of neuropathic pain was induced by partial sciatic nerve ligation (PSL). PSL rats on days 7 and 14 after nerve injury were subjected to imaging with a small-animal positron emission tomography/computed tomography (PET/CT) scanner using [(11)C]PK11195 to detect spinal microglial activation by means of noninvasive in vivo imaging. Spinal [(3)H]PK11195 autoradiography was performed to confirm the results of [(11)C]PK11195 PET in PSL rats. Quantitative RT-PCR of CD11b and GFAP mRNA, and the immunohistochemistry of Iba1 and GFAP were investigated to detect activated microglia and astrocytes. Mechanical allodynia was observed in the ipsilateral paw of PSL rats from day 3 after nerve injury and stably persisted from days 7 to 14. PET/CT fusion images clearly showed large amounts of accumulation of [(11)C]PK11195 in the lumbar spinal cord on days 7 and 14 after nerve injury. [(11)C]PK11195 enhanced images were restricted to the L3-L6 area of the spinal cord. The standardized uptake value (SUV) of [(11)C]PK11195 was significantly increased in the lumbar spinal cord compared to that of the thoracic region. Increased specific binding of [(11)C]PK11195 to TSPO in the spinal cord of PSL rats was confirmed by competition studies using unlabeled (R, S)-PK11195. Increased [(3)H]PK11195 binding was also observed in the ipsilateral dorsal horn of the L3-L6 spinal cord on days 7 and 14 after nerve injury. CD11b mRNA and Iba1 immunoreactive cells increased significantly on days 7 and 14 after nerve injury by PSL. However, changes in GFAP mRNA and immunoreactivity were slight in the ipsilateral side of PSL rats. In the present study, we showed that glial activation could be quantitatively imaged in the spinal cord of neuropathic pain rats using [(11)C]PK11195 PET, suggesting that high resolution PET using TSPO-specific radioligands might be useful for imaging to assess the role of glial activation, including neuroinflammatory processes, in neuropathic pain patients.

Community-based intervention to improve dietary habits and promote physical activity among older adults: a cluster randomized trial.

BACKGROUND: The fastest growing age group globally is older adults, and preventing the need for long-term nursing care in this group is important for social and financial reasons. A population approach to diet and physical activity through the use of social services can play an important role in prevention. This study examined the effectiveness of a social health program for community-dwelling older adults aimed at introducing and promoting physical activity in the home at each individual's pace, helping participants maintain good dietary habits by keeping self-check sheets, and determining whether long-standing unhealthy or less-than-ideal physical and dietary habits can be changed. METHOD: This cluster randomized trial conducted at 6 community centers in an urban community involved 92 community-dwelling older adults aged 65-90 years. The intervention group (3 community centers; n = 57) participated in the social health program "Sumida TAKE10!" which is an educational program incorporating the "TAKE10!® for Older Adults" program, once every 2 weeks for 3 months. The control group (3 community centers; n=35) was subsequently provided with the same program as a crossover intervention group. The main outcome measures were changes in food intake frequency, food frequency score (FFS), dietary variety score (DVS), and frequency of walking and exercise. The secondary outcome measures were changes in self-rated health, appetite, and the Tokyo Metropolitan Institute of Gerontology (TMIG) Index of Competence score. RESULTS: Compared to baseline, post-intervention food intake frequency for 6 of 10 food groups (meat, fish/shellfish, eggs, potatoes, fruits, and seaweed), FFS, and DVS were significantly increased in the intervention group, and interaction effects of FFS and DVS were seen between the two groups. No significant differences were observed between baseline and post-intervention in the control group. Frequency of walking and exercise remained unchanged in both groups, and no significant difference in improvement rate was seen between the groups. Self-rated health was significantly increased in the intervention group. Appetite and TMIG Index of Competence score were unchanged in both groups. CONCLUSIONS: The social health program resulted in improved dietary habits, as measured by food intake frequency, FFS, and DVS, and may improve self-rated health among community-dwelling older adults. TRIAL REGISTRATION NUMBER: UMIN000007357.

Generation of an ICF syndrome model by efficient genome editing of human induced pluripotent stem cells using the CRISPR system.

Genome manipulation of human induced pluripotent stem (iPS) cells is essential to achieve their full potential as tools for regenerative medicine. To date, however, gene targeting in human pluripotent stem cells (hPSCs) has proven to be extremely difficult. Recently, an efficient genome manipulation technology using the RNA-guided DNase Cas9, the clustered regularly interspaced short palindromic repeats (CRISPR) system, has been developed. Here we report the efficient generation of an iPS cell model for immunodeficiency, centromeric region instability, facial anomalies syndrome (ICF) syndrome using the CRISPR system. We obtained iPS cells with mutations in both alleles of DNA methyltransferase 3B (DNMT3B) in 63% of transfected clones. Our data suggest that the CRISPR system is highly efficient and useful for genome engineering of human iPS cells.

miR-29 represses the activities of DNA methyltransferases and DNA demethylases.

Members of the microRNA-29 (miR-29) family directly target the DNA methyltransferases, DNMT3A and DNMT3B. Disturbances in the expression levels of miR-29 have been linked to tumorigenesis and tumor aggressiveness. Members of the miR-29 family are currently thought to repress DNA methylation and suppress tumorigenesis by protecting against de novo methylation. Here, we report that members of the miR-29 family repress the activities of DNA methyltransferases and DNA demethylases, which have opposing roles in control of DNA methylation status. Members of the miR-29 family directly inhibited DNA methyltransferases and two major factors involved in DNA demethylation, namely tet methylcytosine dioxygenase 1 (TET1) and thymine DNA glycosylase (TDG). Overexpression of miR-29 upregulated the global DNA methylation level in some cancer cells and downregulated DNA methylation in other cancer cells, suggesting that miR-29 suppresses tumorigenesis by protecting against changes in the existing DNA methylation status rather than by preventing de novo methylation of DNA.

Genome-wide analysis of DNA methylation and expression of microRNAs in breast cancer cells.

DNA methylation of promoters is linked to transcriptional silencing of protein-coding genes, and its alteration plays important roles in cancer formation. For example, hypermethylation of tumor suppressor genes has been seen in some cancers. Alteration of methylation in the promoters of microRNAs (miRNAs) has also been linked to transcriptional changes in cancers; however, no systematic studies of methylation and transcription of miRNAs have been reported. In the present study, to clarify the relation between DNA methylation and transcription of miRNAs, next-generation sequencing and microarrays were used to analyze the methylation and expression of miRNAs, protein-coding genes, other non-coding RNAs (ncRNAs), and pseudogenes in the human breast cancer cell lines MCF7 and the adriamycin (ADR) resistant cell line MCF7/ADR. DNA methylation in the proximal promoter of miRNAs is tightly linked to transcriptional silencing, as it is with protein-coding genes. In protein-coding genes, highly expressed genes have CpG-rich proximal promoters whereas weakly expressed genes do not. This is only rarely observed in other gene categories, including miRNAs. The present study highlights the epigenetic similarities and differences between miRNA and protein-coding genes.

Efficient generation of epigenetic disease model mice by epigenome editing using the piggyBac transposon system.

BACKGROUND: Epigenome-edited animal models enable direct demonstration of disease causing epigenetic mutations. Transgenic (TG) mice stably expressing epigenome-editing factors exhibit dramatic and stable changes in target epigenome modifications. Successful germline transmission of a transgene from founder mice to offspring will yield a sufficient number of epigenome-edited mice for phenotypic analysis; however, if the epigenetic mutation has a detrimental phenotypic effect, it can become difficult to obtain the next generation of animals. In this case, the phenotype of founder mice must be analyzed directly. Unfortunately, current TG mouse production efficiency (TG founders per pups born) is relatively low, and improvements would increase the versatility of this technology. RESULTS: In the current study, we describe an approach to generate epigenome-edited TG mice using a combination of both the dCas9-SunTag and piggyBac (PB) transposon systems. Using this system, we successfully generated mice with demethylation of the differential methylated region of the H19 gene (H19-DMR), as a model for Silver-Russell syndrome (SRS). SRS is a disorder leading to growth retardation, resulting from low insulin-like growth factor 2 (IGF2) gene expression, often caused by epimutations at the H19-IGF2 locus. Under optimized conditions, the efficiency of TG mice production using the PB system was approximately threefold higher than that using the conventional method. TG mice generated by this system showed demethylation of the targeted DNA region and associated changes in gene expression. In addition, these mice exhibited some features of SRS, including intrauterine and postnatal growth retardation, due to demethylation of H19-DMR. CONCLUSIONS: The dCas9-SunTag and PB systems serve as a simple and reliable platform for conducting direct experiments using epigenome-edited founder mice.

The Dnmt3b splice variant is specifically expressed in in vitro-manipulated blastocysts and their derivative ES cells.

Manipulation of preimplantation embryos in vitro, such as in vitro fertilization (IVF), in vitro culture (IVC), intracytoplasmic sperm injection (ICSI), somatic cell nuclear transfer (SCNT) and other assisted reproduction technologies (ART), has contributed to the development of infertility treatment and new animal reproduction methods. However, such embryos often exhibit abnormal DNA methylation patterns in imprinted genes and centromeric satellite repeats. These DNA methylation patterns are established and maintained by three DNA methyltransferases: Dnmt1, Dnmt3a and Dnmt3b. Dnmt3b is responsible for the creation of methylation patterns during the early stage of embryogenesis and consists of many alternative splice variants that affect methylation activity; nevertheless, the roles of these variants have not yet been identified. In this study, we found an alternatively spliced variant of Dnmt3b lacking exon 6 (Dnmt3bΔ6) that is specific to mouse IVC embryos. Dnmt3bΔ6 also showed prominent expression in embryonic stem (ES) cells derived from in vitro manipulated embryos. Interestingly, IVC blastocysts were hypomethylated in centromeric satellite repeat regions that could be susceptible to methylation by Dnmt3b. In vitro methylation activity assays showed that Dnmt3bΔ6 had lower activity than normal Dnmt3b. Our findings suggest that Dnmt3bΔ6 could induce a hypomethylation status especially in in vitro manipulated embryos.

[Usefulness of reticulocyte hemoglobin equivalent for the safety of pre-operative autologous blood donation].

Pre-operative autologous blood donation (PABD) provides safe blood for patients at the expense of the risk of iron deficiency anemia that may compromise the patients. The reticulocyte hemoglobin equivalent (RET-He) is an indirect measure of the functional iron available for the erythropoiesis over the previous 2-3 days. The aim of this study was to evaluate the clinical usefulness of RET-He quickly measured by the automated hematology analyzer Sysmex XE-2100 in patients undergoing PABD at our hospital. Receiver-operating characteristic curve analysis revealed that RET-He was reliable in the diagnosis of iron deficiency anemia. Two of 14 patients in the absence of post-PABD iron replacement developed marked anemia with low RET-He levels after PABD, suggesting that this anemia was due to iron deficiency. Of 26 patients receiving post-PABD iron replacement, 8 who had already showed low RET-He levels at PABD developed statistically significant reduction in hemoglobin levels after PABD despite adequate iron replacement, indicating that the 8 patients had iron deficiency prior to PABD. These findings suggest that automated measurement of RET-He may contribute to improve the safety of PABD.

Positive Association between High Protein Food Intake Frequency and Physical Performance and Higher-Level Functional Capacity in Daily Life.

Nutritional factors, including low protein intake and poor dietary variety, affect age-associated impairment in physical performance resulting in physical frailty. This cross-sectional study investigated the association between intake frequency of major high protein foods and both physical performance and higher-level functional capacity using the food frequency score (FFS) and high protein food frequency score (PFFS) among community-dwelling older adults. The data of 1185 older adults categorized into quartiles based on FFS and PFFS were analyzed. After adjusting for covariates, FFS and PFFS were significantly associated with physical performance [FFS, usual gait speed (p for trend = 0.007); PFFS, usual gait speed (p for trend < 0.001), maximum gait speed (p for trend = 0.002), timed up and go (p for trend = 0.025)], and higher-level functional capacity [FFS (p for trend < 0.001); PFFS (p for trend < 0.001)]. After excluding PFFS data, the participants' scores were associated with only higher-level functional capacity. Multi-regression analysis with higher-level functional capacity as the covariate showed that FFS and PFFS were significantly correlated with physical performance. Hence, improving food intake frequency, particularly that of high protein foods, and dietary variety may help maintain higher-level functional capacity and physical performance in community-dwelling older adults.

The Association between Dietary Variety and Physical Frailty in Community-Dwelling Older Adults.

The aim of this cross-sectional study was to examine the association between diet variety and physical frailty in community-dwelling older adults. Data of 577 older adults (mean age: 74.0 ± 6.3 years, women: 62.5%) were analyzed. Diet variety was assessed using the Food Frequency Score (FFS) (maximum, 30 points). The FFS assessed the one-week consumption frequency of ten foods (meat, fish/shellfish, eggs, milk & dairy products, soybean products, green & yellow vegetables, potatoes, fruits, seafood, and fats & oil). Physical frailty was assessed using Fried's component (slowness, weakness, exhaustion, low physical activity, and weight loss). The participants were classified into frail, pre-frail, and non-frail groups. The prevalence of physical frailty was 6.6%. This study found significant associations between physical frailty and low FFS after adjusting for covariates (odds ratio (OR) 0.90, 95% confidence interval (CI) 0.84-0.97, p < 0.01). The optimal cutoff point of the FFS for physical frailty was ≤16 points. FFS lower than the cutoff point were significantly associated with physical frailty after adjusting for covariates (OR 3.46, 95% CI 1.60-7.50, p < 0.01). Diet variety assessed using the FFS cutoff value of ≤16 points was related to the physical frailty status in community-dwelling older adults.

Application of decarboxylation reactions for improvement of dielectric properties of a methacrylic polymer.

In this work, we present a method based on thermally induced decarboxylation to solve the incompatibility of dielectric properties and the developability in aqueous developer solution of dielectric materials for the application in photodefinable insulating materials. Herein, malonic acid 2-hydroxyethyl methacrylate monoester, a methacrylate monomer with ß-keto acid is synthesized via phosphotungstic acid catalyzed esterification of 2-hydroxyethyl methacrylate and malonic acid. Further polymerisation is conducted with the obtained monomer to prepare poly(2-hydroxyethyl methacrylate-co-malonic acid 2-hydroxyethyl acrylate monoester). The thermally induced decarboxylation behaviors of the prepared polymer are confirmed with TGA, pyrolysis-GCMS analysis, and IR analysis. D f/D k tests and dissolution tests are carried out to investigate the influences of decarboxylation on the dielectric properties and dissolubility in aqueous solution, respectively. The results demonstrate that the significant changes in dielectric properties and dissolubility are due to the thermally induced decarboxylation.

MeCP2-dependent repression of an imprinted miR-184 released by depolarization.

Both fragile X syndrome and Rett syndrome are commonly associated with autism spectrum disorders and involve defects in synaptic plasticity. MicroRNA is implicated in synaptic plasticity because fragile X mental retardation protein was recently linked to the microRNA pathway. DNA methylation is also involved in synaptic plasticity since methyl CpG-binding protein 2 (MeCP2) is mutated in patients with Rett syndrome. Here we report that expression of miR-184, a brain-specific microRNA repressed by the binding of MeCP2 to its promoter, is upregulated by the release of MeCP2 after depolarization. The restricted release of MeCP2 from the paternal allele results in paternal allele-specific expression of miR-184. Our finding provides a clue to the link between the microRNA and DNA methylation pathways.