The influence of N6-methyladenosine (m6A) RNA methylation on synaptic function and local protein synthesis

Leading theories on learning and memory postulate the importance of strengthening specific synaptic pathways, communication of post- and pre-synaptic terminals, and dendritic local protein synthesis. RNA-binding proteins which recognise epigenomic N6-methyladenosine (m6A) messenger RNA modifications can regulate a number of molecular functions including polyribosome loading, RNA decay, splicing, and translation. To investigate whether and how synaptic plasticity may be regulated by m6A RNA processing, three approaches were undertaken. First, I analysed m6A-sequencing data of human grey matter, white matter, and fetal tissue to identify transcripts that were m6A-methylated in human brain, as well as the biological processes and diseases enriched in this data. Several transcripts whose protein product is important in synaptic structure and function, particularly neurotransmitter receptors, were found to be methylated. The most highly enriched gene ontology terms included “synapses” and “nervous system development,” suggesting a function for m6A methylation in plasticity. Second, I performed immunofluorescent assays on quiescent and activated neuronal cell lines to examine changes in colocalisation of m6A modifications with m6A-binding proteins at synapses. Colocalisation of m6A and YTHDF1, YTHDF3, and ALKBH5 at synapses all increased significantly after 15 minutes of glutamate receptor activation. Changes were also observed 24 hours after activation. These results suggest the binding of these proteins to m6A-mRNAs responds to plasticity processes and is time-dependent. Third, abundance of m6A-modified RNA and expression of YTHDF1/YTHDF3 in the cerebellum, frontal and cingulate gyrus cortex, and hippocampus of normal and neurological disease-affected human brain was examined. Using machine learning quantitative analysis, I found m6A abundance and YTH protein expression to be differently dysregulated in each region and neuronal population of brain affected by Parkinson’s disease, Lewy Body Dementia, and cognitive defects. Overall, these findings provide evidence highlighting the importance of m6A-mediated regulation of local protein synthesis at synapses, providing a clearer understanding of the processes involved in normal synaptic and neuronal function and how m6A regulation may contribute to the pathophysiology of brain disease

From form to function: m6A methylation links mRNA structure to metabolism

Reversible N6-methyladenosine (m6A) RNA modification is a posttranscriptional epigenetic modification of the RNA that regulates many key aspects of RNA metabolism and function. In this review, we highlight major recent advances in the field, with special emphasis on the potential link between m6A modifications and RNA structure. We will also discuss the role of RNA methylation of neuronal transcripts, and the emerging evidence of a potential role in RNA transport and local translation in dendrites and axons of transcripts involved in synaptic functions and axon growth

Modifying the m6A brain methylome by ALKBH5-mediated demethylation: a new contender for synaptic tagging

Synaptic plasticity processes, which underlie learning and memory formation, require RNA to be translated local to synapses. The synaptic tagging hypothesis has previously been proposed to explain how mRNAs are available at specific activated synapses. However how RNA is regulated, and which transcripts are silenced or processed as part of the tagging process is still unknown. Modification of RNA by N6-methyladenosine (m6A/m) influences the cellular fate of mRNA. Here, by advanced microscopy, we showed that m6A demethylation by the eraser protein ALKBH5 occurs at active synaptic ribosomes and at synapses during short term plasticity. We demonstrated that at activated glutamatergic post-synaptic sites, both the YTHDF1 and YTHDF3 reader and the ALKBH5 eraser proteins increase in co-localisation to m6A-modified RNAs; but only the readers showed high co-localisation to modified RNAs during late-stage plasticity. The YTHDF1 and YTHFDF3 readers also exhibited differential roles during synaptic maturation suggesting that temporal and subcellular abundance may determine specific function. m6A-sequencing of human parahippocampus brain tissue revealed distinct white and grey matter m6A methylome profiles indicating that cellular context is a fundamental factor dictating regulated pathways. However, in both neuronal and glial cell-rich tissue, m6A effector proteins are themselves modified and m6A epitranscriptional and posttranslational modification processes coregulate protein cascades. We hypothesise that the availability m6A effector protein machinery in conjunction with RNA modification, may be important in the formation of condensed synaptic nanodomain assemblies through liquid-liquid phase separation. Our findings support that m6A demethylation by ALKBH5 is an intrinsic component of the synaptic tagging hypothesis and a molecular switch which leads to alterations in the RNA methylome, synaptic dysfunction and potentially reversible disease states

The influence of N6-methyladenosine (m6A) RNA methylation on synaptic function and local protein synthesis

Leading theories on learning and memory postulate the importance of strengthening specific synaptic pathways, communication of post- and pre-synaptic terminals, and dendritic local protein synthesis. RNA-binding proteins which recognise epigenomic N6-methyladenosine (m6A) messenger RNA modifications can regulate a number of molecular functions including polyribosome loading, RNA decay, splicing, and translation. To investigate whether and how synaptic plasticity may be regulated by m6A RNA processing, three approaches were undertaken. First, I analysed m6A-sequencing data of human grey matter, white matter, and fetal tissue to identify transcripts that were m6A-methylated in human brain, as well as the biological processes and diseases enriched in this data. Several transcripts whose protein product is important in synaptic structure and function, particularly neurotransmitter receptors, were found to be methylated. The most highly enriched gene ontology terms included “synapses” and “nervous system development,” suggesting a function for m6A methylation in plasticity. Second, I performed immunofluorescent assays on quiescent and activated neuronal cell lines to examine changes in colocalisation of m6A modifications with m6A-binding proteins at synapses. Colocalisation of m6A and YTHDF1, YTHDF3, and ALKBH5 at synapses all increased significantly after 15 minutes of glutamate receptor activation. Changes were also observed 24 hours after activation. These results suggest the binding of these proteins to m6A-mRNAs responds to plasticity processes and is time-dependent. Third, abundance of m6A-modified RNA and expression of YTHDF1/YTHDF3 in the cerebellum, frontal and cingulate gyrus cortex, and hippocampus of normal and neurological disease-affected human brain was examined. Using machine learning quantitative analysis, I found m6A abundance and YTH protein expression to be differently dysregulated in each region and neuronal population of brain affected by Parkinson’s disease, Lewy Body Dementia, and cognitive defects. Overall, these findings provide evidence highlighting the importance of m6A-mediated regulation of local protein synthesis at synapses, providing a clearer understanding of the processes involved in normal synaptic and neuronal function and how m6A regulation may contribute to the pathophysiology of brain disease

m6A mRNA methylation in human brain is disrupted in Lewy body disorders

N6-methyladenosine modification of RNA (m6A) regulates translational control which may influence neuronal dysfunction underlying neurodegenerative diseases. Using microscopy and a machine learning approach, we performed cellular profiling of m6A-RNA abundance and YTHDF1/YTHDF3 m6A reader expression within four regions of the human brain from non-affected individuals and individuals with Parkinson’s disease, Dementia with Lewy bodies (DLB) or Mild Cognitive Impairment (MCI). In non-diseased tissue, we found m6A-modified RNAs showed cell-type and sub-compartment-specific variation. YTHDF1 and YTHDF3 showed opposing expression patterns in the cerebellum, and frontal and cingulate cortices. Machine learning quantitative image analysis revealed that m6A-modified transcripts were significantly altered in localisation and abundance in disease tissue with significant decreases in m6A-RNAs in Parkinson’s disease, and significant increases in m6A-RNA abundance in Dementia with Lewy bodies. MCI tissue showed variability across regions but similar to DLB, in brain areas with an overall significant increase in m6A-RNAs, modified RNAs within dendritic processes were reduced. Using mass spectrometry proteomic datasets to corroborate our findings, we found significant changes in YTHDF3 and m6A anti-reader protein abundance in Alzheimer’s disease (AD) and asymptomatic AD/MCI tissue and correlation with cognitive resilience. These results provide evidence for disrupted m6A regulation in Lewy body diseases and a plausible mechanism through which RNA processing could contribute to the formation of Lewy bodies and other dementia-associated pathological aggregates. The findings suggest that manipulation of epitranscriptomic processes influencing translational control may lead to new therapeutic approaches for neurodegenerative diseases

A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons

Summary: Brain-derived neurotrophic factor (BDNF) promotes neuronal differentiation and survival and is implicated in the pathogenesis of many neurological disorders. Here, we identified a novel intergenic enhancer located 170 kb from the Bdnf gene, which promotes the expression of Bdnf transcript variants during mouse neuronal differentiation and activity. Following Bdnf activation, enhancer-promoter contacts increase, and the region moves away from the repressive nuclear periphery. Bdnf enhancer activity is necessary for neuronal clustering and dendritogenesis in vitro, and for cortical development in vivo. Our findings provide the first evidence of a regulatory mechanism whereby the activation of a distal enhancer promotes Bdnf expression during brain development

International Nosocomial Infection Control Consortiu (INICC) report, data summary of 43 countries for 2007-2012. Device-associated module

We report the results of an International Nosocomial Infection Control Consortium (INICC) surveillance study from January 2007-December 2012 in 503 intensive care units (ICUs) in Latin America, Asia, Africa, and Europe. During the 6-year study using the Centers for Disease Control and Prevention's (CDC) U.S. National Healthcare Safety Network (NHSN) definitions for device-associated health care–associated infection (DA-HAI), we collected prospective data from 605,310 patients hospitalized in the INICC's ICUs for an aggregate of 3,338,396 days. Although device utilization in the INICC's ICUs was similar to that reported from ICUs in the U.S. in the CDC's NHSN, rates of device-associated nosocomial infection were higher in the ICUs of the INICC hospitals: the pooled rate of central line–associated bloodstream infection in the INICC's ICUs, 4.9 per 1,000 central line days, is nearly 5-fold higher than the 0.9 per 1,000 central line days reported from comparable U.S. ICUs. The overall rate of ventilator-associated pneumonia was also higher (16.8 vs 1.1 per 1,000 ventilator days) as was the rate of catheter-associated urinary tract infection (5.5 vs 1.3 per 1,000 catheter days). Frequencies of resistance of Pseudomonas isolates to amikacin (42.8% vs 10%) and imipenem (42.4% vs 26.1%) and Klebsiella pneumoniae isolates to ceftazidime (71.2% vs 28.8%) and imipenem (19.6% vs 12.8%) were also higher in the INICC's ICUs compared with the ICUs of the CDC's NHSN

International Nosocomial Infection Control Consortium report, data summary of 50 countries for 2010-2015: Device-associated module

•We report INICC device-associated module data of 50 countries from 2010-2015.•We collected prospective data from 861,284 patients in 703 ICUs for 3,506,562 days.•DA-HAI rates and bacterial resistance were higher in the INICC ICUs than in CDC-NHSN's.•Device utilization ratio in the INICC ICUs was similar to CDC-NHSN's. Background: We report the results of International Nosocomial Infection Control Consortium (INICC) surveillance study from January 2010-December 2015 in 703 intensive care units (ICUs) in Latin America, Europe, Eastern Mediterranean, Southeast Asia, and Western Pacific. Methods: During the 6-year study period, using Centers for Disease Control and Prevention National Healthcare Safety Network (CDC-NHSN) definitions for device-associated health care-associated infection (DA-HAI), we collected prospective data from 861,284 patients hospitalized in INICC hospital ICUs for an aggregate of 3,506,562 days. Results: Although device use in INICC ICUs was similar to that reported from CDC-NHSN ICUs, DA-HAI rates were higher in the INICC ICUs: in the INICC medical-surgical ICUs, the pooled rate of central line-associated bloodstream infection, 4.1 per 1,000 central line-days, was nearly 5-fold higher than the 0.8 per 1,000 central line-days reported from comparable US ICUs, the overall rate of ventilator-associated pneumonia was also higher, 13.1 versus 0.9 per 1,000 ventilator-days, as was the rate of catheter-associated urinary tract infection, 5.07 versus 1.7 per 1,000 catheter-days. From blood cultures samples, frequencies of resistance of Pseudomonas isolates to amikacin (29.87% vs 10%) and to imipenem (44.3% vs 26.1%), and of Klebsiella pneumoniae isolates to ceftazidime (73.2% vs 28.8%) and to imipenem (43.27% vs 12.8%) were also higher in the INICC ICUs compared with CDC-NHSN ICUs. Conclusions: Although DA-HAIs in INICC ICU patients continue to be higher than the rates reported in CDC-NSHN ICUs representing the developed world, we have observed a significant trend toward the reduction of DA-HAI rates in INICC ICUs as shown in each international report. It is INICC's main goal to continue facilitating education, training, and basic and cost-effective tools and resources, such as standardized forms and an online platform, to tackle this problem effectively and systematically