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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 289 records · Page 16Linked to original sources

bioRxiv2016

A Key Cytoskeletal Regulator of Ubiquitination Amplifies TGFβ Signaling during Mouse Developmental Vascular Patterning

Vascular development involves de novo formation of a capillary plexus, which is then pruned and remodeled by angiogenic events. Cytoskeletal remodeling and directional endothelial migration are essential for developmental and pathological angiogenesis. Smad-dependent TGF{beta} signaling controls vascular patterning and is negatively regulated by microtubules. Here we show that a positive regulator of TGF{beta} signaling is essential for developmental vascular patterning and microtubule stability. Rudhira/BCAS3 is known to bind microtubules and to play a nodal role in cytoskeletal remodeling and directional endothelial cell (EC) migration in vitro. We demonstrate that the molecular and cellular function of Rudhira is deployed at critical steps in vascular patterning. We generated the first floxed mice for rudhira and find that global or endothelial knockout of rudhira results in mid-gestation lethality due to aberrant embryonic and extra-embryonic vessel patterning and defective cardiac morphogenesis. Rudhira null yolk sac ECs show random and retarded migration. Yolk sac transcriptome analysis revealed key mediators of angiogenic processes and TGF{beta} receptor signaling were perturbed in rudhira null mutants. Molecular and biochemical analyses showed that rudhira depletion reduced microtubule stability but increased expression of pathway inhibitors leading to high levels of SMAD2/3 ubiquitination and reduced activation. These effects were not rescued by exogenous TGF{beta}. However, TGF{beta} treatment of wild type ECs increased Rudhira expression. Further, exogenous Rudhira, which promotes directional cell migration, caused increased SMAD2/3 nuclear translocation and reduced inhibitor levels. Therefore, we propose that Rudhira and TGF{beta} signaling are mutually dependent. Rudhira has a dual function in promoting TGF{beta} signaling, possibly by sequestering microtubules and simultaneously preventing SMAD2/3 ubiquitination to permit EC migration and vascular patterning. TGF{beta} signaling and aberrant human Rudhira (Breast Cancer Amplified Sequence 3, BCAS3) expression are both associated with tumour metastasis. Our study identifies a cytoskeletal, cell type-specific modulator of TGF{beta} signaling important in development and cancer.

Developmental Biology↗

bioRxiv2016

EXPERIMENTAL GRANULOMATOUS PULMONARY NOCARDIOSIS in BALB/c MICE

Pulmonary nocardiosis is a granulomatous disease with high mortality that affects both immunosuppressed and immunocompetent patients. The mechanisms leading to the establishment and progression of the infection are currently unknown. An animal model to study these mechanisms is sorely needed. We report the first in vivo model of granulomatous pulmonary nocardiosis that closely resembles human pathology. BALB/c mice infected intranasally with two different doses of GFP-expressing Nocardia brasiliensis ATCC700358 (NbGFP), develop weight loss and pulmonary granulomas. Mice infected with 109 CFUs progressed towards death within a week while mice infected with 108 CFUs died after five to six months. Histological examination of the lungs revealed that both the higher and lower doses of NbGFP induced granulomas with NbGFP clearly identifiable at the center of the lesions. Mice exposed to 108 CFUs and subsequently to 109 CFUs were not protected against disease severity but had less granulomas suggesting some degree of protection. Attempts to identify a cellular target for the infection were unsuccessful but we found that bacterial microcolonies in the suspension used to infect mice were responsible for the establishment of the disease. Small microcolonies of NbGFP, incompatible with nocardial doubling times starting from unicellular organisms, were identified in the lung as early as six hours after infection. Mice infected with highly purified unicellular preparations of NbGFP did not develop granulomas despite showing weight loss. Finally, intranasal delivery of nocardial microcolonies was enough for mice to develop granulomas with minimal weight loss. Taken together these results show that Nocardia brasiliensis microcolonies are both necessary and sufficient for the development of granulomatous pulmonary nocardiosis in mice.

Immunology↗

bioRxiv2016

Modelling the effects of bacterial cell state and spatial location on tuberculosis treatment: Insights from a hybrid multiscale cellular automaton model

If improvements are to be made in tuberculosis (TB) treatment, an increased understanding of disease in the lung is needed. Studies have shown that bacteria in a less metabolically active state, associated with the presence of lipid bodies, are less susceptible to antibiotics, and recent results have highlighted the disparity in concentration of different compounds into lesions. Treatment success therefore depends critically on the responses of the individual bacteria that constitute the infection.\n\nWe propose a hybrid, individual-based approach that analyses spatio-temporal dynamics at the cellular level, linking the behaviour of individual bacteria and host cells with the macroscopic behaviour of the microenvironment. The individual elements (bacteria, macrophages and T cells) are modelled using cellular automaton (CA) rules, and the evolution of oxygen, drugs and chemokine dynamics are incorporated in order to study the effects of the microenvironment in the pathological lesion. We allow bacteria to switch states depending on oxygen concentration, which affects how they respond to treatment. This is the first multiscale model of its type to consider both oxygen-driven phenotypic switching of the Mycobacterium tuberculosis and antibiotic treatment. Using this model, we investigate the role of bacterial cell state and of initial bacterial location on treatment outcome. We demonstrate that when bacteria are located further away from blood vessels, less favourable outcomes are more likely, i.e. longer time before infection is contained/cleared, treatment failure or later relapse. We also show that in cases where bacteria remain at the end of simulations, the organisms tend to be slower-growing and are often located within granulomas, surrounded by caseous material.

Microbiology↗

bioRxiv2016

RiVIERA-MT: A Bayesian model to infer risk variants in related traits using summary statistics and functional genomic annotations

Dissecting the physiological circuitry underlying diverse human complex traits associated with heritable common mutations is an ongoing effort. The primary challenge involves identifying the relevant cell types and the causal variants among the vast majority of the associated mutations in the noncoding regions. To address this challenge, we developed an efficient probabilistic framework. First, we propose a sparse group-guided learning algorithm to infer cell-type-specific enrichments. Second, we propose a fine-mapping Bayesian model that incorporates as Bayesian priors the sparse enrichments to infer risk variants. Using the proposed framework to analyze 32 complex human traits revealed meaningful tissue-specific epigenomic enrichments indicative of the relevant disease pathologies. The prioritized variants exhibit prominent tissue-specific epigenomic signatures and significant enrichments for eQTL and conserved elements. Together, we demonstrate the general benefits of the proposed integrative framework in elucidating meaningful tissue-specific epigenomic elements from large-scale correlated annotations and the implicated functional variants for future experimental interrogation.

Bioinformatics↗

bioRxiv2016

NR1H3 p.Arg415Gln is not associated to multiple sclerosis risk

A recent study by Wang et al claims the low-frequency variant NR1H3 p.Arg415Gln is pathological for multiple sclerosis and determines a patients likelihood of primary progressive disease. We sought to replicate this finding in the International MS Genetics Consortium (IMSGC) patient collection, which is 13-fold larger than the collection of Wang et al, but we find no evidence that this variant is associated either with MS or disease subtype. Wang et al also report a common variant association in the region, which we show captures the association the IMSGC reported in 2013. Therefore, we conclude that the reported low-frequency association is a false positive, likely generated by insufficient sample size. The claim of NR1H3 mutations describing a Mendelian form of MS - of which no examples exist - can therefore not be substantiated by data.

Genetics↗

bioRxiv2016

Sub-millisecond optogenetic control of neuronal firing with two-photon holographic photoactivation of Chronos

Optogenetic neuronal network manipulation promises to at last unravel a long-standing mystery in neuroscience: how does microcircuit activity causally relate to behavioral and pathological states? The challenge to evoke spikes with high spatial and temporal complexity necessitates further joint development of light-delivery approaches and custom opsins. Two-photon scanning and parallel illumination strategies applied to ChR2- and C1V1-expressing neurons demonstrated reliable, in-depth generation of action potentials both in-vitro and in-vivo, but thus far lack the temporal precision necessary to induce precisely timed spiking events. Here, we show that efficient current integration enabled by two-photon holographic amplified laser illumination of Chronos, a highly light-sensitive and fast opsin, can evoke spikes with submillisecond precision and repeated firing up to 100 Hz. These results pave the way for optogenetic manipulation with the spatial and temporal sophistication necessary to mimic natural microcircuit activity.

Neuroscience↗

bioRxiv2016

Specific effect of dopamine partial agonist on counterfactual learning: evidence from Gilles de la Tourette syndrome

The dopamine partial agonist aripiprazole is increasingly used to treat pathologies for which other antipsychotics are indicated because it displays fewer side effects, such as sedation and depression-like symptoms, than other dopamine receptor antagonists. Previously, we showed that aripiprazole may protect motivational function by preserving reinforcement-related signals used to sustain reward-maximization behaviour in a simple action-outcome learning task. However, the effect of aripiprazole on more cognitive facets of human reinforcement learning, such as learning from the hypothetical outcomes of alternative courses of action (i.e., counterfactual learning), is unknown.\n\nTo test the influence of aripiprazole on counterfactual learning, we administered a reinforcement-learning task that involves both direct learning from obtained outcomes and indirect learning from forgone outcomes to two groups of Gilles de la Tourette (GTS) patients, one consisting of patients who were completely unmedicated and the other consisting of patients who were receiving aripiprazole monotherapy, and to healthy subjects. We replicated a previous finding showing that aripiprazole does not affect direct learning from obtained outcomes in GTS. We also found that whereas learning performance improved in the presence of counterfactual feedback in both healthy controls and unmedicated GTS patients, this was not the case in aripiprazole-medicated GTS patients.\n\nOur results suggest that whereas aripiprazole preserves direct learning of action-outcome associations, it may impair more complex inferential processes, such as counterfactual learning, from forgone outcomes.

Neuroscience↗

bioRxiv2016

Mechanisms of self-assembly and fibrillization of the prion-like domains

The mechanism of the self-assembly and fibrillization of the prion-like domains lies at the heart of their physiology and pathology. Here with the same methods previously established, we aimed to further decipher the mechanism by characterizing two prion-like sequences with the electrostatic properties very different from that of the full-length TDP-43 prion-like domain with a very basic pI value: namely the C-half of the TDP-43 prion-like domain only abundant in Gly, Ser, Asn and Gln with a pI of ~6.3, and the FUS prion-like domain enriched with Gly, Ser, Gln and Tyr with a pI of ~3.5. Interestingly, the C-half with the TDP-43 unique hydrophobic region removed is no longer able to form insoluble aggregates/fibrils but still capable of self-assembling into the reversible hydrogel with cross-{beta} structures, despite being much slower than the full-length. On the other hand, the FUS prion-like domain rapidly self-assembles into the reversible hydrogel with cross-{beta} fibrillar structures in 1 mM phosphate buffer at pH 6.8 but its self-assembly becomes very slow in 50 mM MES buffer at pH 5.5. Our study reveals that despite having completely different electrostatic properties, the full-length and C-half of the TDP-43 prion-like domain, as well as FUS prion-like domain all have the similar pH-dependence in self-assembly as we previously reported (Lim et al., [2016] PLOS Biol 14:e1002338). This unambiguously indicates that the self-assembly of the prion-like domains is not generally governed by the electrostatic interaction. Rather, their self-assembly and fibrillization are specified by the sequences despite being highly polar and degenerative. Furthermore, our study provides the first evidence that the formation of reversible hydrogel with cross-{beta} structures is separable from fibrillization of the prion-like domain. Finally, our results also successfully reconcile the previous discrepancy about the conformation and mechanism of the self-assembly of the FUS prion-like domain.

Biophysics↗

bioRxiv2016

Case Report: Aggressive Tibial Pseudarthrosis as Primary Symptom in Infant with Neurofibromatosis

Neurofibromatosis (NF1) is a rare genetic neurologic disorder with over 30 distinct clinical manifestations, the top 3 of which include cafe-au-lait spotting, benign tumors and abnormal freckling. Pseudarthrosis (PA), also known as a \"false joint,\" is a rare subset of NF1 symptomology, characterized by bone fractures and nonunion caused by severe bowing of long bones. To date, it is invariably reported as secondary to NF1, commonly at 24 months of age. Here we describe a 4-month old infant who presented with PA as primary symptom, and in absence of an NF1 first-degree relative. Initial manifestation was guarding of the leg and increased irritability upon palpation of the knee, subsequent to light playful jostling. Physician examination revealed gross anterolateral bowing of the left leg. Radiography confirmed tibia-fibula bowing and pathologic transverse fracture at tibia diaphysis, characteristic of PA. Cafe-au-lait spots developed at 6 months subsequent to PA, but with number and size well below the National Institutes of Health criteria for NF1 diagnosis. At 14 months, exome sequencing established definitive NF1 diagnosis. Treatment involved PA takedown surgery. Although healing was seen after 2 months, complications emerged by 6 months. This case suggests that for primary PA without clear etiology, first-contact and consulting physicians should pay careful attention and be vigilant to timing of clinical onset and severity. Early, severe primary PA warrants accelerated NF1 exome sequencing, suggesting expansion of existing federal guidelines may be necessary to improve detection and prognosis of this rare, debilitating but readily managed condition.\n\nFinancial DisclosureThe authors have no financial relationships relevant to this article to disclose\n\nConflict of InterestThe authors have no conflicts of interest to disclose\n\nClinical Trial RegistrationNA

Genetics↗

bioRxiv2016

Genetic diagnosis of Mendelian disorders via RNA sequencing

Across a large variety of Mendelian disorders, 50 to 75% of undiagnosed patients do not receive a genetic diagnosis by whole exome sequencing indicative of underlying regulatory variants. In contrast, whole genome sequencing allows the discovery of all genetic variants, but their significant number, coupled with a poor understanding of the non-coding genome, makes their prioritization challenging. Here, we demonstrate the power of directly sequencing transcriptomes by providing a genetic diagnosis for 11% (5 of 47) of unsolved mitochondrial disease patients and strong candidates for other patients. We found a median of 1 aberrantly expressed gene, 5 aberrant splicing events, and 6 mono-allelically expressed rare variants in patient-derived fibroblasts and established disease-causing roles for each kind. Private exons often arose from sites that are weakly spliced in other individuals, providing an important clue for future variant prioritization. One such intronic exon-creating variant was found in three unrelated families in the complex I assembly factor TIMMDC1, which we consequently established as a novel disease-associated gene. In conclusion, our study expands the diagnostic tools for detecting non-exonic variants of Mendelian disorders and provides examples of intronic loss-of-function variants with pathological relevance.

Genetics↗

bioRxiv2016

Single-cell quantitation of histones and histone post-translational modifications across the cell cycle by high-throughput imaging

Post-translational modifications (PTM) of histone proteins are critical determinants of genome function. Analysis of histone PTM levels has traditionally been performed using biochemical bulk methods to measure individual modifications. These approaches are generally unable to detect differences in histone and histone PTM levels during the cell cycle, do not provide information on variability amongst individual cells, and they are not suitable for multiplexing to comprehensively analyze large numbers of histone modifications in single experiments. We have developed HiHiMap (High-throughput Histone Mapping), an automated high-throughput immunofluorescence technique to determine histone and histone PTM levels across the cell cycle at the single-cell level in a highly parallel format. Our approach uses imaging-based quantification of DNA content and cyclin A levels to stage individual cells in the cell cycle combined with determination of the corresponding histone and histone PMT levels in the same cells. We have applied HiHiMap to a set of 22 histone variants and histone modifications. As proof-of-principle for a biological application, we use HiHiMap to the histone and histone PTMs landscape in primary, immortalized and oncogenically transformed cells. We find differences in the behavior of a specific set of histone PTM during the the cell cycle in transformed cells compared to normal and immortalized cells. The method developed here is widely applicable to the systematic study of histone modifications in physiological and pathological settings at the single-cell level.

Cell Biology↗

bioRxiv2016

DNA damage is a major cause of sequencing errors, directly confounding variant identification.

Pervasive mutations in somatic cells generate a heterogeneous genomic population within an organism and may result in serious medical conditions. While cancer is the most studied disease associated with somatic variations, recent advances in single cell and ultra deep sequencing indicate that a number of phenotypes and pathologies are impacted by cell specific variants. Currently, the accurate identification of low allelic frequency somatic variants relies on a combination of deep sequencing coverage and multiple evidences of the presence of variants. However, in this study we show that false positive variants can account for more than 70% of identified somatic variations, rendering conventional detection methods inadequate for accurate determination of low allelic variants. Interestingly, these false positive variants primarily originate from mutagenic DNA damage which directly confounds determination of genuine somatic mutations. Furthermore, we developed and validated a simple metric to measure mutagenic DNA damage, and demonstrated that mutagenic DNA damage is the leading cause of sequencing errors in widely used resources including the 1000 Genomes Project and The Cancer Genome Atlas.

Genomics↗

bioRxiv2016

Deep Learning and Association Rule Mining for Predicting Drug Response in Cancer

A major challenge in cancer treatment is predicting the clinical response to anti-cancer drugs for each individual patient. For complex diseases such as cancer, characterized by high inter-patient variance, the implementation of precision medicine approaches is dependent upon understanding the pathological processes at the molecular level. While the \"omics\" era provides unique opportunities to dissect the molecular features of diseases, the ability to utilize it in targeted therapeutic efforts is hindered by both the massive size and diverse nature of the \"omics\" data. Recent advances with Deep Learning Neural Networks (DLNNs), suggests that DLNN could be trained on large data sets to efficiently predict therapeutic responses in cancer treatment. We present the application of Association Rule Mining combined with DLNNs for the analysis of high-throughput molecular profiles of 1001 cancer cell lines, in order to extract cancer-specific signatures in the form of easily interpretable rules and use these rules as input to predict pharmacological responses to a large number of anti-cancer drugs. The proposed algorithm outperformed Random Forests (RF) and Bayesian Multitask Multiple Kernel Learning (BMMKL) classification which currently represent the state-of-the-art in drug-response prediction. Moreover, the in silico pipeline presented, introduces a novel strategy for identifying potential therapeutic targets, as well as possible drug combinations with high therapeutic potential. For the first time, we demonstrate that DLNNs trained on a large pharmacogenomics data-set can effectively predict the therapeutic response of specific drugs in different cancer types. These findings serve as a proof of concept for the application of DLNNs to predict therapeutic responsiveness, a milestone in precision medicine.

Bioinformatics↗

bioRxiv2016

Unlocked capacity of proteins to attack membranes characteristic of aggregation: the evil for diseases and aging from Pandora’s box

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC=\"FIGDIR/small/071274_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (61K):\norg.highwire.dtl.DTLVardef@3e897borg.highwire.dtl.DTLVardef@1b429f6org.highwire.dtl.DTLVardef@12f0dbeorg.highwire.dtl.DTLVardef@2f5d8f_HPS_FORMAT_FIGEXP M_FIG C_FIG AbstractAggregation of specific proteins is characteristic of a large spectrum of human diseases including all neurodegenerative diseases, while aggregation of non-specific proteins has been now identified to be a biomarker for cellular aging down to Escherichia coli. Previously, as facilitated with our discovery in 2005 that \"completely insoluble\" proteins could be all solubilized in unsalted water [Song (2009) FEBS Lett. 583: 953], we found that the TDP-43 prion-like domain in fact contains an intrinsic membrane-interacting subdomain [Lim et al. [2016] PLoS Biol. 14, e1002338]. We decrypted that ALS-causing mutations/cofactor-depletion act to render the wellstructured folds of cytosolic VAPB-MSP domain and SOD1 into highly disordered states, thus becoming buffer-insoluble. Most surprisingly, this also unlocks the amphiphilic/hydrophobic regions universally exiting in proteins, which thus acquire a novel capacity in abnormally interacting with membranes [Qin et al. (2013) F1000Res 2-221.v2; Lim (2016) BBA-Biomembranes. 1858: 2223]. Here we aimed extend our discovery to address two fundamental questions: 1) why many E. coli proteins become aggregated in aging; and 2) whether aggregation-prone proteins can also acquire a novel capacity in interacting with membranes; by dissecting the 557-residue S1 ribosomal protein into 7 fragments to disrupt its 6 S1 folds, followed by extensive CD and NMR characterizations. The results reveal that we have successfully eliminated all 6 S1 folds and fragment 4 becomes highly disordered and thus buffer-insoluble. Most strikingly, F4 does acquire a capacity in transforming into a helical conformation in membrane environments. Here, for the first time, our study deciphers that like ALScausing mutants, the disruption of a well-folded E. coli cytosolic protein also unlocks its amphiphilic/hydrophobic regions which are capable of abnormally interacting with membranes. Therefore, proteins, the most important functional players for all forms of life, can transform into membrane-toxic forms triggering diseases and aging, if their hydrophobic/amphiphilic regions are unlocked by genetic, pathological or/and environmental factors, which is characteristic of severe aggregation.

Biophysics↗

bioRxiv2016

Investigating resting-state functional connectivity in the cervical spinal cord at 3T

The study of spontaneous fluctuations in the blood-oxygen-level-dependent (BOLD) signal has recently been extended from the brain to the spinal cord. Two ultra-high field functional magnetic resonance imaging (fMRI) studies in humans have provided evidence for reproducible resting-state connectivity between the dorsal horns as well as between the ventral horns, and a study in non-human primates has shown that these resting-state signals are impacted by spinal cord injury. As these studies were carried out at ultra-high field strengths using region-of-interest (ROI) based analyses, we investigated whether such resting-state signals could also be observed at the clinically more prevalent field strength of 3T. In a reanalysis of a sample of 20 healthy human participants who underwent a resting-state fMRI acquisition of the cervical spinal cord, we were able to observe significant dorsal horn connectivity as well as ventral horn connectivity, but no consistent effects for connectivity between dorsal and ventral horns, thus replicating the human 7T results. These effects were not only observable when averaging along the acquired length of the spinal cord, but also when we examined each of the acquired spinal segments separately, which showed similar patterns of connectivity. Finally, we investigated the robustness of these resting-state signals against variations in the analysis pipeline by varying the type of ROI creation, temporal filtering, nuisance regression and connectivity metric. We observed that - apart from the effects of band-pass filtering - ventral horn connectivity showed excellent robustness, whereas dorsal horn connectivity showed moderate robustness. Together, our results provide evidence that spinal cord resting-state connectivity is a robust and spatially consistent phenomenon that could be a valuable tool for investigating the effects of pathology, disease progression, and treatment response in neurological conditions with a spinal component, such as spinal cord injury.

Neuroscience↗

bioRxiv2016

Quantitative analysis of cryptic splicing associated with TDP-43 depletion

Reliable exon recognition is key to the splicing of pre-mRNAs into mature mRNAs. TDP-43 is an RNA-binding protein whose nuclear loss and cytoplasmic aggregation are a hallmark pathology in amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD). TDP-43 depletion causes the aberrant inclusion of cryptic exons into a range of transcripts, but their extent, relevance to disease pathogenesis and whether they are caused by other RNA-binding proteins implicated in ALS/FTD are unknown. We developed an analysis pipeline to discover and quantify cryptic exon inclusion and applied it to publicly available human and murine RNA-sequencing data. We detected widespread cryptic splicing in TDP-43 depletion datasets but almost none in another ALS/FTD-linked protein FUS. Sequence motif and iCLIP analysis of cryptic exons demonstrated that they are bound by TDP-43. Unlike the cryptic exons seen in hnRNP C depletion, those linked to TDP-43 do not originate from a particular family of transposable element. Cryptic exons are poorly conserved and inclusion overwhelmingly leads to nonsense-mediated decay of the host transcript, with reduced transcript levels observed in differential expression analysis. RNA-protein interaction data on 73 different RNA-binding proteins showed that, in addition to TDP-43, 7 specifically bind TDP-43 linked cryptic exons.This suggests that TDP-43 competes with other splicing factors for binding to cryptic exons and can repress cryptic exon inclusion. Our quantitative analysis pipeline confirms the presence of cryptic exons during TDP-43 depletion and will aid investigation of their relevance to disease.

Genomics↗

bioRxiv2016

Traumatic brain injury causes retention of long introns in metabolic genes via regulation of intronic Histone 3 lysine 36 methylation levels in the sub-acute phase of injury

Traumatic brain injury (TBI) can cause persistent pathological alteration of neurons. This may lead to cognitive dysfunctions, depression, and even increased susceptibility to life threatening diseases, such as epilepsy and Alzheimers Disease. To investigate the underlying genetic and molecular basis of TBI, Wasserman and colleagues developed an inexpensive and reproducible model for simulating TBI in Drosophila melanogaster (Fruit Fly). Using a modified version of this high impact trauma (HIT) device, we subjected w1118 fruit flies to mild closed head trauma. To determine the transcriptomic changes that contribute to survival post TBI, we collected fly heads from the survivors at 2 time points; 4 hours and 24 hours post-trauma. Mild TBI had limited impact on the steady state RNA levels but showed large perturbations in alternative splicing (AS) 24 hours post-trauma. Classification of these AS changes showed selective retention (RI) of long introns (>81bps), with a mean size of ~3000bps. Some of these RI genes also showed a significant reduction in transcript abundance and were specifically enriched in genes involved in mitochondrial metabolism. The RI are enriched in ACACACA motifs known to bind to Smooth (SM), an hnRNPL class of splicing factor. Mutating SM (sm4/Df) resulted in reversal of RI observed 24 hours post-trauma, and in some cases, elimination of basal levels of RI in long introns. This observation suggests that SM is critical regulator of RI and that this process is enhanced by TBI. Interestingly, chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) for histone 3 lysine 36 trimethylation (H3K36me3) revealed increased levels of this histone modification in retained introns post-trauma. Consistent with this observation, mutations in lysine specific demethylase 4A (KDM4A), which de-methylate H3K36me3, increased RI in many of the same long introns affected by TBI. Additionally, higher H3K36me3 levels are observed around intronic SM-binding motifs post-trauma, suggesting interaction between H3K36me3 and SM binding to intronic splicing suppressor sites might be responsible for increasing RI of metabolic genes as a novel mechanism to improve survival after TBI.

Neuroscience↗

bioRxiv2016

ImSig: A resource for the identification and quantification of immune signatures in blood and tissue transcriptomics data

The outcome of many diseases is commonly correlated with the immune response at the site of pathology. The ability to monitor the status of the immune system in situ provides a mechanistic understanding of disease progression, a prognostic assessment and a guide for therapeutic intervention. Global transcriptomic data can be deconvoluted to provide an indication of the cell types present and their activation state, but the gene signatures proposed to date are either disease-specific or have been derived from data generated from isolated cell populations. Here we describe an improved set of immune gene signatures, ImSig, derived based on their co-expression in blood and tissue datasets. ImSig includes validated lists of marker genes for the main immune cell types and a number of core pathways. When used in combination with network analysis, ImSig is an accurate and easy to use approach for monitoring immune phenotypes in transcriptomic data derived from clinical samples.

Immunology↗