Misregulation of Alternative Splicing in a Mouse Model of Rett Syndrome

Mutations in the human MECP2 gene cause Rett syndrome (RTT), a severe neurodevelopmental disorder that predominantly affects girls. Despite decades of work, the molecular function of MeCP2 is not fully understood. Here we report a systematic identification of MeCP2-interacting proteins in the mouse brain. In addition to transcription regulators, we found that MeCP2 physically interacts with several modulators of RNA splicing, including LEDGF and DHX9. These interactions are disrupted by RTT causing mutations, suggesting that they may play a role in RTT pathogenesis. Consistent with the idea, deep RNA sequencing revealed misregulation of hundreds of splicing events in the cortex of Mecp2 knockout mice. To reveal the functional consequence of altered RNA splicing due to the loss of MeCP2, we focused on the regulation of the splicing of the flip/flop exon of Gria2 and other AMPAR genes. We found a significant splicing shift in the flip/flop exon toward the flop inclusion, leading to a faster decay in the AMPAR gated current and altered synaptic transmission. In summary, our study identified direct physical interaction between MeCP2 and splicing factors, a novel MeCP2 target gene, and established functional connection between a specific RNA splicing change and synaptic phenotypes in RTT mice. These results not only help our understanding of the molecular function of MeCP2, but also reveal potential drug targets for future therapies

2014-2015 Master Class - James Dunham (Viola)

https://spiral.lynn.edu/conservatory_masterclasses/1026/thumbnail.jp

Elevated atmospheric CO2 concentration triggers redistribution of nitrogen to promote tillering in rice

Elevated atmospheric CO2 concentration (eCO2) often reduces nitrogen (N) content in rice plants and stimulates tillering. However, there is a general consensus that reduced N would constrain rice tillering. To resolve this contradiction, we investigated N distribution and transcriptomic changes in different rice plant organs after subjecting them to eCO2 and different N application rates. Our results showed that eCO2 significantly promoted rice tillers (by 0.6, 1.1, 1.7, and 2.1 tillers/plant at 0, 75, 150, and 225 kg N ha−1 N application rates, respectively) and more tillers were produced under higher N application rates, confirming that N availability constrained tillering in the early stages of growth. Although N content declined in the leaves (−11.0 to −20.7 mg g−1) and sheaths (−9.8 to −28.8 mg g−1) of rice plants exposed to eCO2, the N content of newly emerged tillers on plants exposed to eCO2 equaled or exceeded the N content of tillers produced under ambient CO2 conditions. Apparently, the redistribution of N within the plant per se was a critical adaptation strategy to the eCO2 condition. Transcriptomic analysis revealed that eCO2 induced less extensive alteration of gene expression than did N application. Most importantly, the expression levels of multiple N-related transporters and receptors such as nitrate transporter NRT2.3a/b and NRT1.1a/b were differentially regulated in leaf and shoot apical meristem, suggesting that multiple genes were involved in sensing the N signal and transporting N metabolites to adapt to eCO2. The redistribution of N in different organs could be a universal adaptation strategy of terrestrial plants to eCO2

Expression of Linear and Novel Circular Forms of an INK4/ARF-Associated Non-Coding RNA Correlates with Atherosclerosis Risk

Human genome-wide association studies have linked single nucleotide polymorphisms (SNPs) on chromosome 9p21.3 near the INK4/ARF (CDKN2a/b) locus with susceptibility to atherosclerotic vascular disease (ASVD). Although this locus encodes three well-characterized tumor suppressors, p16INK4a, p15INK4b, and ARF, the SNPs most strongly associated with ASVD are ∼120 kb from the nearest coding gene within a long non-coding RNA (ncRNA) known as ANRIL (CDKN2BAS). While individuals homozygous for the atherosclerotic risk allele show decreased expression of ANRIL and the coding INK4/ARF transcripts, the mechanism by which such distant genetic variants influence INK4/ARF expression is unknown. Here, using rapid amplification of cDNA ends (RACE) and analysis of next-generation RNA sequencing datasets, we determined the structure and abundance of multiple ANRIL species. Each of these species was present at very low copy numbers in primary and cultured cells; however, only the expression of ANRIL isoforms containing exons proximal to the INK4/ARF locus correlated with the ASVD risk alleles. Surprisingly, RACE also identified transcripts containing non-colinear ANRIL exonic sequences, whose expression also correlated with genotype and INK4/ARF expression. These non-polyadenylated RNAs resisted RNAse R digestion and could be PCR amplified using outward-facing primers, suggesting they represent circular RNA structures that could arise from by-products of mRNA splicing. Next-generation DNA sequencing and splice prediction algorithms identified polymorphisms within the ASVD risk interval that may regulate ANRIL splicing and circular ANRIL (cANRIL) production. These results identify novel circular RNA products emanating from the ANRIL locus and suggest causal variants at 9p21.3 regulate INK4/ARF expression and ASVD risk by modulating ANRIL expression and/or structure

Elevated atmospheric CO2 concentration triggers redistribution of nitrogen to promote tillering in rice

Abstract Elevated atmospheric CO2 concentration (eCO2) often reduces nitrogen (N) content in rice plants and stimulates tillering. However, there is a general consensus that reduced N would constrain rice tillering. To resolve this contradiction, we investigated N distribution and transcriptomic changes in different rice plant organs after subjecting them to eCO2 and different N application rates. Our results showed that eCO2 significantly promoted rice tillers (by 0.6, 1.1, 1.7, and 2.1 tillers/plant at 0, 75, 150, and 225 kg N ha−1 N application rates, respectively) and more tillers were produced under higher N application rates, confirming that N availability constrained tillering in the early stages of growth. Although N content declined in the leaves (−11.0 to −20.7 mg g−1) and sheaths (−9.8 to −28.8 mg g−1) of rice plants exposed to eCO2, the N content of newly emerged tillers on plants exposed to eCO2 equaled or exceeded the N content of tillers produced under ambient CO2 conditions. Apparently, the redistribution of N within the plant per se was a critical adaptation strategy to the eCO2 condition. Transcriptomic analysis revealed that eCO2 induced less extensive alteration of gene expression than did N application. Most importantly, the expression levels of multiple N‐related transporters and receptors such as nitrate transporter NRT2.3a/b and NRT1.1a/b were differentially regulated in leaf and shoot apical meristem, suggesting that multiple genes were involved in sensing the N signal and transporting N metabolites to adapt to eCO2. The redistribution of N in different organs could be a universal adaptation strategy of terrestrial plants to eCO2

Global Transcriptome and Co-Expression Network Analysis Reveal Contrasting Response of Japonica and Indica Rice Cultivar to γ Radiation

Japonica and indica are two important subspecies in cultivated Asian rice. Irradiation is a classical approach to induce mutations and create novel germplasm. However, little is known about the differential response between japonica and indica rice after γ radiation. Here, we utilized the RNA sequencing and Weighted Gene Co-expression Network Analysis (WGCNA) to compare the transcriptome differences between japonica Nipponbare (NPB) and indica Yangdao6 (YD6) in response to irradiation. Japonica subspecies are more sensitive to irradiation than the indica subspecies. Indica showed a higher seedling survival rate than japonica. Irradiation caused more extensive DNA damage in shoots than in roots, and the severity was higher in NPB than in YD6. GO and KEGG pathway analyses indicate that the core genes related to DNA repair and replication and cell proliferation are similarly regulated between the varieties, however the universal stress responsive genes show contrasting differential response patterns in japonica and indica. WGCNA identifies 37 co-expressing gene modules and ten candidate hub genes for each module. This provides novel evidence indicating that certain peripheral pathways may dominate the molecular networks in irradiation survival and suggests more potential target genes in breeding for universal stress tolerance in rice

Distinguishing artificial spin ice states using magnetoresistance effect for neuromorphic computing

Abstract Artificial spin ice (ASI) consisting patterned array of nano-magnets with frustrated dipolar interactions offers an excellent platform to study frustrated physics using direct imaging methods. Moreover, ASI often hosts a large number of nearly degenerated and non-volatile spin states that can be used for multi-bit data storage and neuromorphic computing. The realization of the device potential of ASI, however, critically relies on the capability of transport characterization of ASI, which has not been demonstrated so far. Using a tri-axial ASI system as the model system, we demonstrate that transport measurements can be used to distinguish the different spin states of the ASI system. Specifically, by fabricating a tri-layer structure consisting a permalloy base layer, a Cu spacer layer and the tri-axial ASI layer, we clearly resolve different spin states in the tri-axial ASI system using lateral transport measurements. We have further demonstrated that the tri-axial ASI system has all necessary required properties for reservoir computing, including rich spin configurations to store input signals, nonlinear response to input signals, and fading memory effect. The successful transport characterization of ASI opens up the prospect for novel device applications of ASI in multi-bit data storage and neuromorphic computing

Altered AMPA receptor decay kinetics and synaptic transmission in <i>Mecp2</i> KO cortical neurons can be reversed by ESF.

<p>(a) Representative sample trace of glutamate-evoked current in WT and <i>Mecp2</i> KO cortical neurons. Scale bar = 15ms. (b) Quantification of decay time constant τ of glutamate-evoked current in WT and <i>Mecp2</i> KO cortical neurons. Mean ± S.E.M is plotted. n = 37 for WT, n = 33 for KO. ** <i>P</i> < 0.01; two tailed <i>t</i>-test. (c) Representative mEPSC sample trace in WT and <i>Mecp2</i> KO cortical neurons with or without CTZ treatment. Scale bar = 10ms. (d) Quantification of decay time constant τ of mEPSC. Mean ± S.E.M is plotted. n = 42 for WT, n = 46 for KO, n = 38 for KO + CTZ. ** <i>P</i> < 0.01; one-way ANOVA with Tukey’s multiple comparisons test. (e-f) qRT-PCR analysis the flip/flop ratio (e) and total <i>Gria2</i> minigene (f) expression in HEK 293 cells co-transfected with indicated ESF and the <i>Gria2</i> minigene. Mean ± S.E.M is plotted. n = 4 for each group. * <i>P</i> < 0.05, *** <i>P</i> < 0.001; two tailed <i>t</i>-test. (g) Quantification of decay time constant τ of glutamate-evoked current in WT, <i>Mecp2</i> KO (KO+Ctrl) and <i>Mecp2</i> KO cortical neurons infected with ESF-flop-Gly lentivirus (KO+ESF). Recording was done on acute slice from mouse infected with lentivirus two weeks after stereotaxic injection. Mean ± S.E.M is plotted. n = 50 for WT, n = 26 for KO+Ctrl, n = 27 for KO+ESF. ** <i>P</i> < 0.01, *** <i>P</i> < 0.001; one-way ANOVA with Tukey’s multiple comparisons test. (h) Representative sample trace of AMPAR-mediated current in response to repetitive stimulations in WT and <i>Mecp2</i> KO neurons infected with control or ESF-flop-Gly lentivirus (KO+Ctrl or KO+ESF). Scale bar, 100ms. (i) Relative current amplitude of the xth stimulation to first stimulation in WT and <i>Mecp2</i> KO neurons infected with either control or ESF-flop-Gly lentivirus (KO+Ctrl or KO+ESF). Mean ± S.E.M is plotted. n = 36 for WT, n = 18 for KO+Ctrl, n = 18 for KO+ESF. Asterisk denotes <i>P</i>-value for comparison between WT and KO+Ctrl, *** <i>P</i> < 0.001. Pound sign denotes <i>P</i>-value for comparison between KO+ESF and KO+Ctrl, # <i>P</i> < 0.05, ## <i>P</i> < 0.01; repeated measures two-way ANOVA with Tukey’s multiple comparisons test. (j) Quantification of decay time constant τ of glutamate-evoked current in neurons infected with shCtrl or shLedgf lentivirus. Mean ± S.E.M is plotted. n = 13 for shCtrl, n = 12 for shLedgf, * <i>P</i> < 0.05; two-tailed <i>t</i>-test. (k) Representative sample trace of AMPAR-mediated current in response to repetitive stimulations in neurons infected with shCtrl or shLedgf lentivirus. Scale bar, 100ms. (l) Relative current amplitude of xth stimulation to first stimulation in neurons infected with shCtrl or shLedgf. Mean ± S.E.M is plotted. n = 12 for each group, *** <i>P</i> < 0.001; repeated measures two-way ANOVA with Tukey's multiple comparisons test.</p