Endogenous retroviruses (ERVs) constitute a significant part of vertebrate genomes. They originated from past retroviral infections and some of them retain transcriptional activity. The key mechanism avoiding uncontrolled ERV transcription is DNA methylation-mediated epigenetic silencing. Despite numerous studies describing the involvement of ERV activity in cellular processes, epigenetic regulation of ERVs is still poorly understood. We previously described a cervid endogenous retrovirus (CrERV) in the mule deer genome. This virus exhibits massive insertional polymorphism, suggesting recent activity. Here we employed NGS-based strategy to determine the methylation pattern of CrERV integrations in four mule deer. Besides the vast majority of methylated integrations, we identified a tiny fraction of demethylated proviral copies. These copies represent evolutionary older integrations located near gene promoters. In general, our work is a first attempt to characterize the epigenetic landscape of insertionally polymorphic ERV on a whole-genome scale and offers insight into its interactions with a host.

The sphingolipid ceramide regulates beta-oxidation of medium and long chain fatty acids in mitochondria. It is not known whether it also regulates oxidation of very long chain fatty acids (VLCFAs) in peroxisomes. Using affinity chromatography, co-immunoprecipitation, and proximity ligation assays we discovered that ceramide interacts with Hsd17b4, an enzyme critical for peroxisomal VLCFA oxidation and docosahexaenoic acid (DHA) generation. Immunocytochemistry showed that Hsd17b4 is distributed to ceramide-enriched mitochondria-associated membranes (CEMAMs). Molecular docking and in vitro mutagenesis experiments showed that ceramide binds to the sterol carrier protein 2-like domain in Hsd17b4 adjacent to peroxisome targeting signal 1 (PTS1), the C-terminal signal for interaction with peroxisomal biogenesis factor 5 (Pex5), a peroxin mediating transport of Hsd17b4 into peroxisomes. Inhibition of ceramide biosynthesis induced translocation of Hsd17b4 from CEMAMs to peroxisomes, interaction of Hsd17b4 with Pex5, and upregulation of DHA. This data indicates a novel role of ceramide as a molecular switch regulating interaction of Hsd17b4 with Pex5 and peroxisomal function.

DNA topoisomerase IB (TOP1) regulates DNA topological structure in many cellular metabolic processes and is a validated target for development of antitumor agents. Our previous study revealed that the benzophenanthridone scaffold is a novel chemotype for the discovery of TOP1 inhibitors. In this work, a series of novel 5-aminoethyl substituted benzophenanthridone derivatives have been synthesized and evaluated for TOP1 inhibition and cytotoxicity. Compound 12 exhibits the most potent TOP1 inhibition (+++) and cytotoxicity in human cancer cell lines with GI50 values at nanomolar concentration range. 12 induces the cellular TOP1cc formation and DNA damage, resulting in HCT116 cell apoptosis. The pharmacokinetics, acute toxicity and antitumor efficiency in vivo of 12 were also studied.

Glioblastoma (GBM) has a poor prognosis despite intensive treatment with surgery and chemoradiotherapy. Previous studies using dose-escalated radiotherapy have demonstrated improved survival; however, increased rates of radionecrosis have limited its use. Development of radiosensitizers could improve patient outcome. In the present study, we report the use of sodium sulfide (Na2S), a hydrogen sulfide (H2S) donor, to selectively kill GBM cells (T98G and U87) while sparing normal human cerebral microvascular endothelial cells (hCMEC/D3). Na2S also decreased mitochondrial respiration, increased oxidative stress and induced γH2AX foci and oxidative base damage in GBM cells. Since Na2S did not significantly alter T98G capacity to perform non-homologous end-joining or base excision repair, it is possible that GBM cell killing could be attributed to increased damage induction due to enhanced reactive oxygen species production. Interestingly, Na2S enhanced mitochondrial respiration, produced a more reducing environment and did not induce high levels of DNA damage in hCMEC/D3. Taken together, this data suggests involvement of mitochondrial respiration in Na2S toxicity in GBM cells. The fact that survival of LN-18 GBM cells lacking mitochondrial DNA (ρ0) was not altered by Na2S whereas the survival of LN-18 ρ+ cells was compromised supports this conclusion. When cells were treated with Na2S and photon or proton radiation, GBM cell killing was enhanced, which opens the possibility of H2S being a radiosensitizer. Therefore, this study provides the first evidence that H2S donors could be used in GBM therapy to potentiate radiation-induced killing.

Feline Leukemia Virus (FeLV) and Feline Immunodeficiency Virus (FIV) are two prevalent transmittable diseases for domestic cats. This paper reports the frequency of these two diseases compared globally across Gross Domestic Product (GDP) at purchasing power parity per capita (PPP). Information around FeLV and FIV rates of infection in specific locations around the world was analyzed from 47 published articles. Results show that based on the data available, the statistical model indicates that the highest percentage of FeLV or FIV infected cats live in areas of lower PPP (p ≤.001) with a decreasing rate of infection of FeLV and FIV with increasing income. Two theories for this could be that the lower PPP locations in this study were also in areas of greater feral cat and cat colony populations, as well as were areas with less emphasis on animal welfare and animal control programs. Additional research should be conducted to strengthen the study size in South America and Africa before further conclusions can be drawn.

Conventional MRI is not sensitive to many pathological processes underpinning multiple sclerosis (MS) ongoing in normal appearing brain tissue (NABT). Quantitative MRI (qMRI) and a multiparameter mapping (MPM) protocol are used to simultaneously quantify magnetization transfer (MT) saturation, transverse relaxation rate R2* (1/T2*) and longitudinal relaxation rate R1 (1/T1), and assess differences in NABT microstructure between MS patients and healthy controls (HC).

The pearl oyster Pinctada fucata martensii (Pfu.), widely cultured in the South China Sea, is a precious source of sea pearls and calcifying materials. A yellow shell variety of Pfu. was obtained after years of artificial breeding. To identify differentially expressed genes between yellow shell and normal black shell pearl oysters, we performed transcriptomic sequencing and proteomic analyses using mantle edge tissues.

Clinical prediction models are not routinely validated. To facilitate validation procedures, the online Evidencio platform ( https://www.evidencio.com ) has developed a tool partly automating this process. This study aims to determine whether semi-automated validation can reliably substitute manual validation.

In patients suffering from Chronic Inflammatory Demyelinating Polyneuropathy (CIDP) disease severity is assessed by Medical Research Counsil (MRC) Scale or Inflammatory Neuropathy Cause and Treatment (INCAT) disability score. However, none of these methods is appropriate to objectively assess muscle weakness or to detect very small subclinical changes. More objective and quantitative measures are needed in order to evaluate treatment efficiency or to detect subclinical affection of upper limps for early diagnosis. The goal of our study was to objectively quantify muscular weakness in CIDP patients with the non-invasive Quantitative Motor (Q-Motor) test of Grip Force Assessment (QGFA) as well as the Involuntary Movement Assessment (QIMA) and to search for differences between typical and atypical CIDP variants. In addition, we hypothesized that Q-Motor findings correlate with disease severity scales such as MRC or INCAT score.

General cognitive ability, sometimes referred to as intelligence, is associated with educational attainment throughout childhood. Most studies that have explored the neural correlates of intelligence in childhood focus on individual brain regions. This analytical approach is designed to identify restricted sets of voxels that overlap across participants. By contrast, we explored the relationship between white matter connectome organization, intelligence, and education. In both a sample of typically-developing children (N = 63) and a sample of struggling learners (N = 139), the white matter connectome efficiency was strongly associated with intelligence and educational attainment. Further, intelligence partially mediated the relationship between connectome efficiency and educational attainment. In contrast, a canonical voxel-wise analysis failed to identify any significant relationships. The results emphasize the importance of distributed brain network properties for cognitive or educational ability in childhood. Our findings are interpreted in the context of a developmental theory, which emphasizes the interaction between different subsystems over developmental time.

Hexokinases (HXKs) have determined to be multifaceted proteins, and they are the only ones able to phosphorylate glucose in plants. However, the binding mode for ATP to plant HXKs remains unclear. Here, we report the crystal structures of rice hexokinase 6 (OsHXK6) in four different forms: (i) apo-form, (ii) binary complex with D-Glc, (iii) quaternary complex with ADP, PO4 and Mg2+, and (iv) pentanary complex with D-Glc, ADP, PO4, and Mg2+. The apo form is in the open state conformation, and the three others are in the closed state, indicating that glucose and ADP-PO4 binding induces a large conformational change by domain rearrangement. The quaternary complex is a novel intermediate during the catalytic reaction we trapped for the first time, which provides a new evidence for the enzymatic mechanism of HXKs. In addition, the latter two complexes reveal the binding mode for ADP-PO4 to plant HXKs, which provide the structural explanation for the dual-function of OsHXK6. In addition, we identified that residues Gly112, Thr261, Gly262, and Gly450 are essential to the binding between ADP-PO4 and OsHXK6 by a series of single mutations and enzymatic assays. Our study provide structural basis for the other functional studies of OsHXK6 in rice.

Autoantibodies against CASPR2 (contactin-associated protein-like 2) have been linked to autoimmune limbic encephalitis that manifests with memory disorders and temporal lobe seizures. According to the growing number of data supporting a role for CASPR2 in neuronal excitability, CASPR2 forms a molecular complex with transient axonal glycoprotein-1 (TAG-1) and shaker-type voltage-gated potassium channels (Kv1.1 and Kv1.2) in compartments critical for neuronal activity and is required for Kv1 proper positioning. Whereas the perturbation of these functions could explain the symptoms observed in patients, the pathogenic role of anti-CASPR2 antibodies has been poorly studied. In the present study, we find that patient autoantibodies alter Caspr2 distribution at the cell membrane promoting cluster formation. We confirm in a HEK cellular model that the anti-CASPR2 antibodies impede CASPR2/TAG-1 interaction and we identify the domains of CASPR2 and TAG-1 taking part in this interaction. Moreover, introduction of CASPR2 into HEK cells induces a marked increase of the level of Kv1.2 surface expression and in cultures of hippocampal neurons Caspr2-positive inhibitory neurons appear to specifically express high levels of Kv1.2. Importantly, in both cellular models, anti-CASPR2 patient autoAb increase Kv1.2 expression. These results provide new insights into the pathogenic role of autoAb in the disease.

Overexpression of Frizzled-7 (FZD7) has been associated with tumor invasion and distant metastases, but little is known about the relationship between FZD7 expression and prognosis in colon cancer.

The computational identification of peptides that can bind the major histocompatibility complex (MHC) with high affinity is an essential step in developing personal immunotherapies and vaccines. We introduce PUFFIN, a deep residual network-based computational approach that quantifies uncertainty in peptide-MHC affinity prediction that arises from observational noise and the lack of relevant training examples. With PUFFIN's uncertainty metrics, we define binding likelihood, the probability a peptide binds to a given MHC allele at a specified affinity threshold. Compared to affinity point estimates, we find that binding likelihood correlates better with the observed affinity and reduces false positives in high-affinity peptide design. When applied to examine an existing peptide vaccine, PUFFIN identifies an alternative vaccine formulation with higher binding likelihood. PUFFIN is freely available for download at http://github.com/gifford-lab/PUFFIN.

Hepatocellular carcinoma (HCC) is a highly fatal malignant cancer worldwide. Elucidating the underlying molecular mechanism of HCC progression is critical for the identification of new therapeutic targets for HCC. This study aimed to determine the role of Non-SMC condensin II complex subunit G2 (NCAPG2) in HCC proliferation and metastasis.

Influenza A virus carries eight negative single-stranded RNAs and uses spliced mRNAs to increase the number of proteins produced from them. Several genome-wide screens for essential host factors for influenza A virus replication revealed a necessity for splicing and splicing-related factors, including Cdc-like kinase 1 (CLK1). This CLK family kinase plays a role in alternative splicing regulation through phosphorylation of serine-arginine rich (SR) proteins. To examine the influence that modulation of splicing regulation has on influenza infection, we analyzed the effect of CLK1 knockdown and inhibition. CLK1 knockdown in A549 cells reduced influenza A/WSN/33 virus replication and increased the level of splicing of segment 7, which encodes the viral M1 and M2 proteins. CLK1-/- mice infected with influenza A/England/195/2009 (H1N1pdm09) virus supported lower levels of virus replication than wild-type mice. Screening of newly developed CLK inhibitors revealed several compounds that have an effect on the level of splicing of influenza A gene segment M in different models and decrease influenza A/WSN/33 virus replication in A549 cells. The promising inhibitor KH-CB19, an indole-based enaminonitrile with unique binding mode for CLK1, and its even more selective analogue NIH39 showed high specificity towards CLK1 and had a similar effect on influenza mRNA splicing regulation. Taken together, our findings indicate that targeting host factors that regulate splicing of influenza mRNAs may represent a novel therapeutic approach.

Src is a known proto-oncogene and its aberrant activity is involved in a variety of cancers, including ovarian cancer, whereas the regulatory mechanism of Src has not been fully clarified. In this study, we identified tripartite motif-containing (TRIM) 50 as a novel negative regulator of Src protein. Our data showed that TRIM50 directly interacted with SH3 domain of Src via its B-box domain; and TRIM50 reduced Src stability by inducing RING domain-dependent K48-linked poly-ubiquitous modification. We further demonstrated that TRIM50 acted as a tumor suppressor in ovarian cancer cells by its negative regulation of Src protein. In vivo animal model verified that TRIM50 inhibited the xenograft tumor growth of ovarian cancer by suppressing Src protein. Clinical investigation showed that expression of TRIM50 in clinical specimens was inversely correlated with the clinical stages, pathology grades and lymph node metastatic status of the patients, which indicated the involvement of aberrant TRIM50 expression in disease progression. Further analysis verified the negative correlation between TRIM50 and Src expression in clinical specimens. Altogether, we identified TRIM50 as a novel suppressor of Src protein, and demonstrated that TRIM50 inhibited ovarian cancer progression by targeting Src and reducing its activity, which provided a novel therapeutic strategy for Src over-activated cancers by positive regulation of TRIM50.

TAR DNA-binding protein 43 (TDP-43) is a hallmark of some neurodegenerative disorders, such as frontotemporal lobar degeneration and amyotrophic lateral sclerosis. TDP-43-related pathology is characterized by its abnormally phosphorylated and ubiquitinated aggregates. It is involved in many aspects of RNA processing, including mRNA splicing, transport, and translation. However, its exact physiological function and role in mechanisms that lead to neuronal degeneration remain elusive. Transgenic rats that were characterized by TDP-43 depletion in neurons exhibited enhancement of the acquisition of fear memory. At the cellular level, TDP-43-depleted neurons exhibited a decrease in the short-term plasticity of intrinsic neuronal excitability. The induction of long-term potentiation in the CA3-CA1 areas of the hippocampus resulted in more stable synaptic enhancement. At the molecular level, the protein levels of an unedited (R) FLOP variant of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) GluR1 and GluR2/3 subunits decreased in the hippocampus. Alterations of FLOP/FLIP subunit composition affected AMPAR kinetics, reflected by cyclothiazide-dependent slowing of the decay time of AMPAR-mediated miniature excitatory postsynaptic currents. These findings suggest that TDP-43 may regulate activity-dependent neuronal plasticity, possibly by regulating the splicing of genes that are responsible for fast synaptic transmission and membrane potential.

Neurodegenerative and neuromuscular disorders can manifest throughout the lifespan of an individual, from infant to elderly individuals. Axonal and synaptic degeneration are early and critical elements of nearly all human neurodegenerative diseases and neural injury, however the molecular mechanisms which regulate this process are yet to be fully elucidated. Furthermore, how the molecular mechanisms governing degeneration are impacted by the age of the individual is poorly understood. Interestingly, in mice which are under 3 weeks of age, the degeneration of axons and synapses following hypoxic or traumatic injury is significantly slower. This process, known as Wallerian degeneration (WD), is a molecularly and morphologically distinct subtype of neurodegeneration by which axons and synapses undergo distinct fragmentation and death following a range of stimuli. In this study, we first use an ex-vivo model of axon injury to confirm the significant delay in WD in neonatal mice. We apply tandem mass-tagging quantitative proteomics to profile both nerve and muscle between P12 and P24 inclusive. Application of unbiased in silico workflows to relevant protein identifications highlights a steady elevation in oxidative phosphorylation cascades corresponding to the accelerated degeneration rate. We demonstrate that inhibition of Complex I prevents the axotomy-induced rise in reactive oxygen species and protects axons following injury. Furthermore, we reveal that pharmacological activation of oxidative phosphorylation significantly accelerates degeneration at the neuromuscular junction in neonatal mice. In summary, we reveal dramatic changes in the neuromuscular proteome during post-natal maturation of the neuromuscular system, and demonstrate that endogenous dynamics in mitochondrial bioenergetics during this time window have a functional impact upon regulating the stability of the neuromuscular system.

We previously reported that the heart-specific choline acetyltransferase (ChAT) gene overexpressing mice (ChAT tg) show specific phenotypes including ischemic tolerance and the CNS stress tolerance. In the current study, we focused on molecular mechanisms responsible for systemic and localized anti-inflammatory phenotypes of ChAT tg. ChAT tg were resistant to systemic inflammation induced by lipopolysaccharides due to an attenuated cytokine response. In addition, ChAT tg, originally equipped with less reactive Kupffer cells, were refractory to brain cold injury, with decreased blood brain barrier (BBB) permeability and reduced inflammation. This is because ChAT tg brain endothelial cells expressed more claudin-5, and their astrocytes were less reactive, causing decreased hypertrophy. Moreover, reconstruction of the BBB integrity in vitro confirmed the consolidation of ChAT tg. ChAT tg were also resistant to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neuronal toxicity due to lower mortality rate and neuronal loss of substantia nigra. Additionally, ChAT tg subjected to MPTP showed attenuated BBB disruption, as evident from reduced sodium fluorescein levels in the brain parenchyma. The activated central cholinergic pathway of ChAT tg lead to anti-convulsive effects like vagus nerve stimulation. However, DSP-4, a noradrenergic neuron-selective neurotoxin against the CNS including the locus ceruleus, abrogated the beneficial phenotype and vagotomy attenuated expression of claudin-5, suggesting the link between the cholinergic pathway and BBB function. Altogether, these findings indicate that ChAT tg possess an anti-inflammatory response potential, associated with upregulated claudin-5, leading to the consolidation of BBB integrity. These characteristics protect ChAT tg against systemic and localized inflammatory pathological disorders, which targets the CNS.