Abl kinase-mediated FUS Tyr526 phosphorylation alters nucleocytoplasmic FUS localization in FTLD-FUS.

Nuclear to cytoplasmic mislocalization and aggregation of multiple RNA-binding proteins (RBPs), including FUS, are the main neuropathological features of the majority of cases of amyotrophic lateral sclerosis (ALS) and frontotemporal lobular degeneration (FTLD). In ALS-FUS, these aggregates arise from disease-associated mutations in FUS, whereas in FTLD-FUS, the cytoplasmic inclusions do not contain mutant FUS, suggesting different molecular mechanisms of FUS pathogenesis in FTLD that remain to be investigated. We have previously shown that phosphorylation of the C-terminal Tyr526 of FUS results in increased cytoplasmic retention of FUS due to impaired binding to the nuclear import receptor TNPO1. Inspired by the above notions, in the current study we developed a novel antibody against the C-terminally phosphorylated Tyr526 FUS (FUSp-Y526) that is specifically capable of recognizing phosphorylated cytoplasmic FUS, which is poorly recognized by other commercially available FUS antibodies. Using this FUSp-Y526 antibody, we demonstrated a FUS phosphorylation-specific effect on the cytoplasmic distribution of soluble and insoluble FUSp-Y526 in various cells and confirmed the involvement of the Src kinase family in Tyr526 FUS phosphorylation. In addition, we found that FUSp-Y526 expression pattern correlates with active pSrc/pAbl kinases in specific brain regions of mice, indicating preferential involvement of cAbl in the cytoplasmic mislocalization of FUSp-Y526 in cortical neurons. Finally, the pattern of immunoreactivity of active cAbl kinase and FUSp-Y526 revealed altered cytoplasmic distribution of FUSp-Y526 in cortical neurons of post-mortem frontal cortex tissue from FTLD patients compared with controls. The overlap of FUSp-Y526 and FUS signals was found preferentially in small diffuse inclusions and was absent in mature aggregates, suggesting possible involvement of FUSp-Y526 in the formation of early toxic FUS aggregates in the cytoplasm that are largely undetected by commercially available FUS antibodies. Given the overlapping patterns of cAbl activity and FUSp-Y526 distribution in cortical neurons, and cAbl induced sequestration of FUSp-Y526 into G3BP1 positive granules in stressed cells, we propose that cAbl kinase is actively involved in mediating cytoplasmic mislocalization and promoting toxic aggregation of wild-type FUS in the brains of FTLD patients, as a novel putative underlying mechanism of FTLD-FUS pathophysiology and progression.

Microbial agents obtained from tomato straw composting effectively promote tomato straw compost maturation and improve compost quality.

Appropriate management of agricultural organic waste (AOW) presents a significant obstacle in the endeavor to attain sustainable agricultural development. The proper management of AOW is a necessity for sustainable agricultural development. This can be done skillfully by incorporating microbial agents in the composting procedure. In this study, we isolated relevant bacteria strains from tomato straw AOW, which demonstrated efficient degradation of lignocellulose without any antagonistic effects in them. These strains were then combined to create a composite microbial agent called Zyco Shield (ZS). The performance of ZS was compared with a commercially effective microorganism (EM) and a control CK. The results indicate that the ZS treatment significantly prolonged the elevated temperature phase of the tomato straw pile, showing considerable degradation of lignocellulosic material. This substantial degradation did not happen in the EM and CK treatments. Moreover, there was a temperature rise of 4-6 â„ƒ in 2 days of thermophilic phase, which was not the case in the EM and CK treatments. Furthermore, the inoculation of ZS substantially enhanced the degradation of organic waste derived from tomato straw. This method increased the nutrient content of the resulting compost and elevated the enzymatic activity of lignocellulose-degrading enzymes, while reducing the urease enzyme activity within the pile. The concentrations of NH4+-N and NO3--N showed increases of (2.13% and 47.51%), (14.81% and 32.17%) respectively, which is again very different from the results of the EM and CK treatments. To some extent, the alterations observed in the microbial community and the abundance of functional microorganisms provide indirect evidence supporting the fact that the addition of ZS microbial agent facilitates the composting process of tomato straw. Moreover, we confirmed the degradation process of tomato straw through X-ray diffraction, Fourier infrared spectroscopy, and by scanning electron microscopy to analyze the role of ZS microbial inoculum composting. Consequently, reinoculation compost strains improves agricultural waste composting efficiency and enhances product quality.

Higher intake of ß-glucan impairs reproduction in a female teleost, Tor putitora (Hamilton, 1822).

Although the immuno-modulatory and stress-relieving properties of ß-glucan is well elucidated in humans and other animal models, including fish, its role as a dietary supplement on reproduction is extremely scarce. Therefore, in this study, adult female fish were fed one of four test diets having 0 (control), 0.5, 1, and 1.5% ß-D-glucan for 130 days and its effect on reproductive performance, ovarian and liver histology, sex hormones, and transcript abundance of selected reproduction-related genes was assessed. Low dietary intake of ß-glucan improved fertilization and hatching rates (p<0.05). The relative fecundity and percentage of spawning females were higher (non-significant) in 0.5% ß-glucan-fed groups. Surprisingly, even after 130 days, spawning did not occur in 1.5% ß-glucan-fed individuals. Irrespective of ß-glucan intake, all the brooders recorded similar plasma 17ß-estradiol and maturation-inducing hormone (p>0.05). Higher intake of ß-glucan (1.5%) upregulated aromatase genes without a parallel increase in 17ß-estradiol. However, plasma vitellogenin increased with increasing ß-glucan up to 1.0% then declined at 1.5% (p<0.05). The fish that received control, 0.5, and 1.5% ß-glucan recorded similar vitellogenin levels in their plasma. Significantly higher plasma cortisol was evidenced in 1.5% ß-glucan fed brooders (p<0.05). Histologically, higher follicular atresia and leaking of yolk material was evidenced in 1.5% ß-glucan-fed group. Liver histology revealed the highest nutrient/lipid accumulation in fish that received 1.0% and 1.5% ß-glucan. This study demonstrated the stimulatory effect of ß-glucan intake at a lower dose (0.5%) on reproduction. However, higher intake (1.5%) could perturb normal reproductive function in a fish model and caused an increased number of atretic follicles leading to spawning/reproductive failure.

Brown sugar as a carbon source can make agricultural organic waste compost enter the secondary thermophilic stage and promote compost decomposition.

To enhance the efficiency of composting agricultural organic waste (AOW), this study aimed to examine the impact of inoculating tomato straw compost with two distinct microbial agents: ZymoZone (ZZ), a composite microbial agent derived from the straw compost and Effective Microorganisms (EM), a commercial microbial agent. Furthermore, in order to reactivate the microorganisms within the compost during the initial high temperature phase, 10% brown sugar was introduced as a carbon source. The objective of this addition was to assess its influence on the composting process. The findings revealed that compared to the control (CK) group, the ZZ and EM treatments extended the first high-temperature phase by 2 and 1 day, respectively. Furthermore, with the addition of 10% brown sugar, the ZZ and EM treatments remained in the second high-temperature phase for 8 and 7 days, respectively, while the CK treatment had already entered the cooling stage by then. Notably, the inoculation of microbial agents and the addition of brown sugar substantially augmented the activity of lignocellulose-related hydrolases, thereby promoting the degradation of lignocellulose in the ZZ and EM treatment groups. This was confirmed by FTIR analysis, which demonstrated that the addition of microbial agents facilitated the degradation of specific substances, leading to reduced absorbance in the corresponding spectra. XRD analysis further indicated a notable reduction in cellulose crystallinity for both the ZZ (8.00%) and EM (7.73%) treatments. Hence, the incorporation of microbial agents and brown sugar in tomato straw compost effectively enhances the composting process and improves the quality of compost products.

Hsp90-mediated regulation of DYRK3 couples stress granule disassembly and growth via mTORC1 signaling.

Stress granules (SGs) are dynamic condensates associated with protein misfolding diseases. They sequester stalled mRNAs and signaling factors, such as the mTORC1 subunit raptor, suggesting that SGs coordinate cell growth during and after stress. However, the molecular mechanisms linking SG dynamics and signaling remain undefined. We report that the chaperone Hsp90 is required for SG dissolution. Hsp90 binds and stabilizes the dual-specificity tyrosine-phosphorylation-regulated kinase 3 (DYRK3) in the cytosol. Upon Hsp90 inhibition, DYRK3 dissociates from Hsp90 and becomes inactive. Inactive DYRK3 is subjected to two different fates: it either partitions into SGs, where it is protected from irreversible aggregation, or it is degraded. In the presence of Hsp90, DYRK3 is active and promotes SG disassembly, restoring mTORC1 signaling and translation. Thus, Hsp90 links stress adaptation and cell growth by regulating the activity of a key kinase involved in condensate disassembly and translation restoration.

Interactome screening of C9orf72 dipeptide repeats reveals VCP sequestration and functional impairment by polyGA.

Repeat expansions in the C9orf72 gene are a common cause of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, two devastating neurodegenerative disorders. One of the proposed mechanisms of GGGGCC repeat expansion is their translation into non-canonical dipeptide repeats, which can then accumulate as aggregates and contribute to these pathologies. There are five different dipeptide repeat proteins (polyGA, polyGR, polyPR, polyPA and polyGP), some of which are known to be neurotoxic. In the present study, we used BioID2 proximity labelling to identify the interactomes of all five dipeptide repeat proteins consisting of 125 repeats each. We identified 113 interacting partners for polyGR, 90 for polyGA, 106 for polyPR, 25 for polyPA and 27 for polyGP. Gene Ontology enrichment analysis of the proteomic data revealed that these target interaction partners are involved in a variety of functions, including protein translation, signal transduction pathways, protein catabolic processes, amide metabolic processes and RNA-binding. Using autopsy brain tissue from patients with C9orf72 expansion complemented with cell culture analysis, we evaluated the interactions between polyGA and valosin containing protein (VCP). Functional analysis of this interaction revealed sequestration of VCP with polyGA aggregates, altering levels of soluble valosin-containing protein. VCP also functions in autophagy processes, and consistent with this, we observed altered autophagy in cells expressing polyGA. We also observed altered co-localization of polyGA aggregates and p62 in cells depleted of VCP. All together, these data suggest that sequestration of VCP with polyGA aggregates contributes to the loss of VCP function, and consequently to alterations in autophagy processes in C9orf72 expansion disorders.

PolyGA targets the ER stress-adaptive response by impairing GRP75 function at the MAM in C9ORF72-ALS/FTD.

ER stress signaling is linked to the pathophysiological and clinical disease manifestations in amyotrophic lateral sclerosis (ALS). Here, we have investigated ER stress-induced adaptive mechanisms in C9ORF72-ALS/FTD, focusing on uncovering early endogenous neuroprotective mechanisms and the crosstalk between pathological and adaptive responses in disease onset and progression. We provide evidence for the early onset of ER stress-mediated adaptive response in C9ORF72 patient-derived motoneurons (MNs), reflected by the elevated increase in GRP75 expression. These transiently increased GRP75 levels enhance ER-mitochondrial association, boosting mitochondrial function and sustaining cellular bioenergetics during the initial stage of disease, thereby counteracting early mitochondrial deficits. In C9orf72 rodent neurons, an abrupt reduction in GRP75 expression coincided with the onset of UPR, mitochondrial dysfunction and the emergence of PolyGA aggregates, which co-localize with GRP75. Similarly, the overexpression of PolyGA in WT cortical neurons or C9ORF72 patient-derived MNs led to the sequestration of GRP75 within PolyGA inclusions, resulting in mitochondrial calcium (Ca2+) uptake impairments. Corroborating these findings, we found that PolyGA aggregate-bearing human post-mortem C9ORF72 hippocampal dentate gyrus neurons not only display reduced expression of GRP75 but also exhibit GRP75 sequestration within inclusions. Sustaining high GRP75 expression in spinal C9orf72 rodent MNs specifically prevented ER stress, normalized mitochondrial function, abrogated PolyGA accumulation in spinal MNs, and ameliorated ALS-associated behavioral phenotype. Taken together, our results are in line with the notion that neurons in C9ORF72-ALS/FTD are particularly susceptible to ER-mitochondrial dysfunction and that GRP75 serves as a critical endogenous neuroprotective factor. This neuroprotective pathway, is eventually targeted by PolyGA, leading to GRP75 sequestration, and its subsequent loss of function at the MAM, compromising mitochondrial function and promoting disease onset.

Quantitative proteomic analysis of GnRH agonist treated GBM cell line LN229 revealed regulatory proteins inhibiting cancer cell proliferation.

BACKGROUND: Gonadotropin-releasing hormone (GnRH) receptor, a rhodopsin-like G-protein coupled receptor (GPCR) family member involved in GnRH signaling, is reported to be expressed in several tumors including glioblastoma multiforme (GBM), one of the most malignant and aggressive forms of primary brain tumors. However, the molecular targets associated with GnRH receptor are not well studied in GBM or in other cancers. The present study aims at investigating the effect of GnRH agonist (Gosarelin acetate) on cell proliferation and associated signaling pathways in GBM cell line, LN229. METHODS: LN229 cells were treated with different concentrations of GnRH agonist (10-10 M to 10-5 M) and the effect on cell proliferation was analyzed by cell count method. Further, total protein was extracted from control and GnRH agonist treated cells (with maximum reduction in cell proliferation) followed by trypsin digestion, labeling with iTRAQ reagents and LC-MS/MS analysis to identify differentially expressed proteins. Bioinformatic analysis was performed for annotation of proteins for the associated molecular function, altered pathways and network analysis using STRING database. RESULTS: The treatment with different concentrations of GnRH agonist showed a reduction in cell proliferation with a maximum reduction of 48.2% observed at 10-6 M. Quantitative proteomic analysis after GnRH agonist treatment (10-6 M) led to the identification of a total of 29 differentially expressed proteins with 1.3-fold change (23 upregulated, such as, kininogen-1 (KNG1), alpha-2-HS-glycoprotein (AHSG), alpha-fetoprotein (AFP), and 6 downregulated, such as integrator complex subunit 11 (CPSF3L), protein FRG1 (FRG1). Some of them are known [KNG1, AHSG, AFP] while others such as inter-alpha-trypsin inhibitor heavy chain H2 (ITIH2), ITIH4, and LIM domain-containing protein 1 (LIMD1) are novel to GnRH signaling pathway. Protein-protein interaction analysis showed a direct interaction of KNG1, a hub molecule, with GnRH, GnRH receptor, EGFR and other interactors including ITIH2, ITIH4 and AHSG. Overexpression of KNG1 after GnRH agonist treatment was validated using Western blot analysis, while a significant inhibition of EGFR was observed after GnRH agonist treatment. CONCLUSIONS: The study suggests a possible link of GnRH signaling with EGFR signaling pathways likely via KNG1. KNG1 inhibitors may be investigated independently or in combination with GnRH agonist for therapeutic applications.

A serum microRNA sequence reveals fragile X protein pathology in amyotrophic lateral sclerosis.

Knowledge about converging disease mechanisms in the heterogeneous syndrome amyotrophic lateral sclerosis (ALS) is rare, but may lead to therapies effective in most ALS cases. Previously, we identified serum microRNAs downregulated in familial ALS, the majority of sporadic ALS patients, but also in presymptomatic mutation carriers. A 5-nucleotide sequence motif (GDCGG; D = G, A or U) was strongly enriched in these ALS-related microRNAs. We hypothesized that deregulation of protein(s) binding predominantly to this consensus motif was responsible for the ALS-linked microRNA fingerprint. Using microRNA pull-down assays combined with mass spectrometry followed by extensive biochemical validation, all members of the fragile X protein family, FMR1, FXR1 and FXR2, were identified to directly and predominantly interact with GDCGG microRNAs through their structurally disordered RGG/RG domains. Preferential association of this protein family with ALS-related microRNAs was confirmed by in vitro binding studies on a transcriptome-wide scale. Immunohistochemistry of lumbar spinal cord revealed aberrant expression level and aggregation of FXR1 and FXR2 in C9orf72- and FUS-linked familial ALS, but also patients with sporadic ALS. Further analysis of ALS autopsies and induced pluripotent stem cell-derived motor neurons with FUS mutations showed co-aggregation of FXR1 with FUS. Hence, our translational approach was able to take advantage of blood microRNAs to reveal CNS pathology, and suggests an involvement of the fragile X-related proteins in familial and sporadic ALS already at a presymptomatic stage. The findings may uncover disease mechanisms relevant to many patients with ALS. They furthermore underscore the systemic, extra-CNS aspect of ALS.

Spit bubbles, speech bubbles, and COVID-19: creating comics in the age of post-infection India.

Positioning this essay at intersection of comics studies, visual literacy studies, and information literacy studies, we investigate an interdisciplinary liaison between crisis in the age of COVID-19 and its awareness campaign through Indian comics. With a focus on awareness programme, Indian artists designed comics to demonstrate their vital position in social engagement through this visual medium. Following impending threats and growing concerns, people of all ages glued themselves to social media, newspapers, and television to keep them updated on the impact of COVID-19. Indian comics e.g. Nagraj Strikes: The Attack of Coronaman (2020), Priya's Mask (2020), Kids, Vaayu, and Corona: Who Wins the Fight? (2020), and 'Be aware of Droplets & Bubbles!!' (2020) aimed to help children comprehend the precautionary steps to be taken to save themselves from getting infected with Coronavirus. While the first three comics showcase spit-bubbles primarily as the source of COVID-19, infusing the content with a tinge of superhero fantasy, 'Be aware of Droplets & Bubbles!!' (2020) unveils the microbiological evolution and mutation of the pathogen in comics format. The objective of the article is to show how Indian comics on COVID-19 can be an advantageous communicative medium to nurture knowledge and edutainment in post-infection India.

Palmitoylation of acetylated tubulin and association with ceramide-rich platforms is critical for ciliogenesis.

Microtubules are polymers composed of αß-tubulin subunits that provide structure to cells and play a crucial role in in the development and function of neuronal processes and cilia, microtubule-driven extensions of the plasma membrane that have sensory (primary cilia) or motor (motile cilia) functions. To stabilize microtubules in neuronal processes and cilia, α tubulin is modified by the posttranslational addition of an acetyl group, or acetylation. We discovered that acetylated tubulin in microtubules interacts with the membrane sphingolipid, ceramide. However, the molecular mechanism and function of this interaction are not understood. Here, we show that in human induced pluripotent stem cell-derived neurons, ceramide stabilizes microtubules, which indicates a similar function in cilia. Using proximity ligation assays, we detected complex formation of ceramide with acetylated tubulin in Chlamydomonas reinhardtii flagella and cilia of human embryonic kidney (HEK293T) cells, primary cultured mouse astrocytes, and ependymal cells. Using incorporation of palmitic azide and click chemistry-mediated addition of fluorophores, we show that a portion of acetylated tubulin is S-palmitoylated. S-palmitoylated acetylated tubulin is colocalized with ceramide-rich platforms in the ciliary membrane, and it is coimmunoprecipitated with Arl13b, a GTPase that mediates transport of proteins into cilia. Inhibition of S-palmitoylation with 2-bromo palmitic acid or inhibition of ceramide biosynthesis with fumonisin B1 reduces formation of the Arl13b-acetylated tubulin complex and its transport into cilia, concurrent with impairment of ciliogenesis. Together, these data show, for the first time, that ceramide-rich platforms mediate membrane anchoring and interaction of S-palmitoylated proteins that are critical for cilium formation, stabilization, and function.

ß-glucan modulates non-specific immune gene expression, thermal tolerance and elicits disease resistance in endangered Tor putitora fry challenged with Aeromonas salmonicida.

An eight-week feeding trial was performed to assess the effect of different dietary levels (0, 0.5, 1.0, and 1.5%) of ß-glucan (sourced from Saccharomyces cerevisiae) on growth, survival, immunological parameters (immune gene expression, lysozyme, and antiprotease), total antioxidant status, thermal tolerance, and disease resistance of Tor putitora fry. Feeding of moderate doses (0.5 and 1.0%) of ß-glucan significantly improved survival but not weight gain percentage as compared to that received unsupplemented control and highest dose (1.5%) of glucan. Supplementation of ß-glucan in diets differentially influenced the mRNA expression of cytokine and other immune genes. For instance, transcripts of cytokines such as tnf-α and il-1ß were significantly upregulated, while ifn-γ and il-10 were unaffected by ß-glucan intake. Also, the relative mRNA expression of tlr-5 and hepcidin1 along with lysozyme and antiprotease activities were remained largely unchanged by dietary glucan administration. In contrast, ß-glucan induced mRNA expression of defensin1 and c3 while decreased the transcript level of mhc-1. On the other hand, dietary inclusion of ß-glucan markedly improved total antioxidant levels and extended the thermal tolerance limits at both the ends, as shown by increased CTmax and lower CTmin than the control group. After feeding ß-glucan for eight weeks, the fish were bath challenged with a bacterial pathogen, Aeromonas salmonicida. The challenge study results revealed that ß-glucan intake improved most of the studied immune parameters, resulting in lower mortality. Overall, dietary inclusion of ß-glucan (0.5-1.0%) was efficient in improving the immune responses, thermal tolerance, and disease resistance of T. putitora fry.

Molecular characterization of non-specific immune genes of endangered golden mahseer (Tor putitora) and their expression during embryonic and larval development.

The present study was undertaken to characterize and analyze the expression of non-specific immune genes to get an insight into the early immune status of endangered golden mahseer. In this study, the full-length mRNA sequence of IFNγ, TNFα, C3, and IL10 was 927, 1409, 5125 and 1177 bp with an ORF of 558, 765, 4938, and 540 bp, encoding a putative protein of 185, 254, 1645, and 179 amino acid residues, respectively. The deduced amino acid sequences of these genes shared highly conserved structures with those from other cyprinids. Ontogenic real-time qPCR results indicated that expression of IFNγ and TNFα was lower until the morula stage and increased from blastula stage and found maximum at the organogenesis stage. Expression of the C3 gene was lower until the gastrula stage, followed by a linear increase from organogenesis to the pre-metamorphosis stage. The expression of IL10 was significantly lower during early developmental stages (till gastrula stage) and reached maximum at organogenesis. The level of IL1ß was found maximum in unfertilized eggs and remained elevated till the morula stage. TLR4 expression remained lower during the initial developmental stages and reached the maximum at the organogenesis stage. The expression level of defensin1 was substantially low until the organogenesis stage. In comparison, hepcidin1 was found considerably high until the blastula stage and remained significantly lower during later stages of development. Overall, the data generated improves knowledge on the immune status of endangered golden mahseer during embryonic and larval development, which may help develop effective immunomodulatory interventions during nursery rearing of golden mahseer to produce fry with better fitness.

Dietary soy lecithin augments antioxidative defense and thermal tolerance but fails to modulate non-specific immune genes in endangered golden mahseer (Tor putitora) fry.

A 70-day experiment was carried out to assess the effect of different levels (0, 1 and 2%) of soy lecithin in the diet on growth, survival, antioxidant defense markers, immune gene expression and thermal tolerance limits of golden mahseer, Tor putitora fry. Percentage weight gain, specific growth rate (SGR %) and survival of mahseer fed lecithin supplemented diets were not significantly different from those of the control group. Also, the mRNA expression levels of different immune related genes such as tnfα, il-1ß, il-10, complement-3, interferon-gamma (ifnγ) and tlr4 were unaffected by dietary lecithin supplementation. Nevertheless, superoxide dismutase (SOD) activity was significantly greater in the lecithin-fed groups than the control fish. The glutathione-S-transferase (GST) activity was exceptionally high in the 2% lecithin supplemented group compared to the rest two groups. This increase in antioxidant status with dietary lecithin supplementation, however, was not reflected in the whole body malonaldehyde (MDA) levels, as it did not vary significantly among the dietary groups. Importantly, dietary inclusion of soy lecithin significantly increased upper thermal tolerance limits as evidenced by higher CTmax and LTmax values. Likewise, golden mahseer fry fed with lecithin supplemented diets (both 1 and 2%) registered significantly lower critical and lethal thermal minimum (CTmin and LTmin) values than the control group, indicating higher cold tolerance capacity. Our results thus demonstrate that the dietary inclusion of soy lecithin could enhance the upper and lower thermal tolerance limits and antioxidant status of golden mahseer fry and failed to enhance immune related gene expression.

The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta.

BACKGROUND: ADP-ribosylation is a ubiquitous post-translational modification that involves both mono- and poly-ADP-ribosylation. ARTD10, also known as PARP10, mediates mono-ADP-ribosylation (MARylation) of substrate proteins. A previous screen identified protein kinase C delta (PKCδ) as a potential ARTD10 substrate, among several other kinases. The voltage-gated K+ channel Kv1.1 constitutes one of the dominant Kv channels in neurons of the central nervous system and the inactivation properties of Kv1.1 are modulated by PKC. In this study, we addressed the role of ARTD10-PKCδ as a regulator of Kv1.1. RESULTS: We found that ARTD10 inhibited PKCδ, which increased Kv1.1 current amplitude and the proportion of the inactivating current component in HeLa cells, indicating that ARTD10 regulates Kv1.1 in living cells. An inhibitor of ARTD10, OUL35, significantly decreased peak amplitude together with the proportion of the inactivating current component of Kv1.1-containing channels in primary hippocampal neurons, demonstrating that the ARTD10-PKCδ signaling cascade regulates native Kv1.1. Moreover, we show that the pharmacological blockade of ARTD10 increases excitability of hippocampal neurons. CONCLUSIONS: Our results, for the first time, suggest that MARylation by ARTD10 controls neuronal excitability.

Transgenerational effects of ß-glucan on thermal tolerance, growth performance, and immune gene expression of endangered cyprinid Tor putitora progeny.

Nutritional programming signifies a process in which broodstock feeding approaches have long-term effects on the subsequent progeny. The present study aimed to elucidate whether supplementing golden mahseer, Tor putitora broodstock diets with ß-glucan affects progeny growth performance, survival, thermal tolerance, and non-specific immunity. Initially, the growth performance of progeny produced from brooders fed with different levels of ß-glucan was non-significant. However, on the 15th and 35th DPH, the maximum weight was observed in fry obtained from the brooders fed with 0.5% followed by 1.0% ß-glucan. Furthermore, on 50th DPH, significantly higher weight was registered in the fry from the 0.5% ß-glucan fed group while 1.0% ß-glucan group had no transgenerational effect on growth. The condition factor of fry obtained from golden mahseer brooders fed with a 0.5% ß-glucan diet was greater than the control and 1.0% ß-glucan fed group. On the other hand, we did not find any significant transgenerational influence of ß-glucan on the survival of the progeny. The thermal tolerance of fry produced from brooders fed with ß-glucan was significantly modulated at both end-points (CTmax and CTmin). Expression of interleukin-1ß was significantly up-regulated in fry obtained from ß-glucan fed brooders. In contrast, the expression level of tumor necrosis factor-α was significantly higher only in fry produced from 1.0% ß-glucan fed brooders. The expression of immunoglobulin light chain and serum amyloid A gene was significantly higher in fry produced from 0.5% ß-glucan fed brooders. Overall results suggest that the dietary provisioning of ß-glucan in golden mahseer brooders can be a strategy to produce healthy and robust fry in captivity for stock enhancement and conservation programs.

Concurrent changes in thermal tolerance thresholds and cellular heat stress response reveals novel molecular signatures and markers of high temperature acclimation in rainbow trout.

The objective of this study was to better understand the molecular mechanisms which regulate acclimatory responses and thermal safety margins of rainbow trout (Oncorhynchus mykiss) at temperatures above physiological optimum. For this, we investigated the time course of changes in critical thermal tolerance thresholds and associated hepatic and renal transcript abundance of molecular markers related to cellular stress response, during high temperature acclimation. The experimental fish were initially acclimated to 17 °C and later exposed to a gradually raised elevated temperature regime (22 °C) for a period of 30 days. CTmax, CTmin and mRNA expression of candidate markers were examined before the thermal challenge (T0) and over the time-course (days) of high temperature exposure (T1, T3, T7, T15 and T30). With respect to organismal response, CTmax was significantly elevated at T3, but the degree of gain in heat tolerance was not persistent. Contrarily, we observed a gradual loss in cold tolerance with highest CTmin estimate at T30. Based on the time-course of mRNA expression, the studied markers could be categorized into those which were persistently elevated (hsp70a, hsp70b, hspa5, hsp90a, hsp90b, stip1 and serpinh1 in kidney and hsp90b in liver); those which concurred with changes in CTmin (hspbp1, hsp90b, stip1, gr1, hif1a, hyou1, tnfa and tlr5 in kidney); and those which concurred with changes in CTmax (hsp90a, serpinh1, tlr5 and lmo2 in liver). Apparently, transcriptional changes in kidney and liver reflected CTmin and CTmax trend, respectively. Expression profile of stip1 and tlr5 suggest that they are potential novel markers which could reflect thermal limits in rainbow trout. Hepatic metabolic markers were either initially elevated (alt, glud, g6pase1) or down-regulated at different time-points (ast2, gls1, fas, cpt1b, mtor), linked to gluconeogenesis and metabolic depression, respectively. Whereas, growth-axis markers showed no significant differences. Overall, this time-course analysis has revealed potential associations in organismal and tissue-specific cellular response to high temperature acclimation in a thermally sensitive coldwater ectotherm.

FUS pathology in ALS is linked to alterations in multiple ALS-associated proteins and rescued by drugs stimulating autophagy.

Amyotrophic lateral sclerosis (ALS) is a lethal disease characterized by motor neuron degeneration and associated with aggregation of nuclear RNA-binding proteins (RBPs), including FUS. How FUS aggregation and neurodegeneration are prevented in healthy motor neurons remain critically unanswered questions. Here, we use a combination of ALS patient autopsy tissue and induced pluripotent stem cell-derived neurons to study the effects of FUS mutations on RBP homeostasis. We show that FUS' tendency to aggregate is normally buffered by interacting RBPs, but this buffering is lost when FUS mislocalizes to the cytoplasm due to ALS mutations. The presence of aggregation-prone FUS in the cytoplasm causes imbalances in RBP homeostasis that exacerbate neurodegeneration. However, enhancing autophagy using small molecules reduces cytoplasmic FUS, restores RBP homeostasis and rescues motor function in vivo. We conclude that disruption of RBP homeostasis plays a critical role in FUS-ALS and can be treated by stimulating autophagy.

Development of docetaxel nanocapsules for improving in vitro cytotoxicity and cellular uptake in MCF-7 cells.

The aim of this study was to fabricate docetaxel loaded nanocapsules (DTX-NCs) with a high payload using Layer-by-Layer (LbL) technique by successive coating with alternate layers of oppositely charged polyelectrolytes. Developed nanocapsules (NCs) were characterized in terms of morphology, particle size distribution, zeta potential (ζ-potential), entrapment efficiency and in vitro release. The morphological characteristics of the NCs were assessed using transmission electron microscopy (TEM) that revealed coating of polyelectrolytes around the surface of particles. The developed NCs successfully attained a submicron particle size while the ζ-potential of optimized NCs alternated between (+) 34.64 ± 1.5 mV to (-) 33.25 ± 2.1 mV with each coating step. The non-hemolytic potential of the NCs indicated the suitability of the developed formulation for intravenous administration. A comparative study indicated that the cytotoxicity of positively charged NCs (F4) was significant higher (p < 0.05) rather than negative charged NCs (F3), plain drug (DTX) and marketed preparation (Taxotere®) when evaluated in vitro on MCF-7 cells. Furthermore, cell uptake studies evidenced a higher uptake of positive NCs (≥1.2 fold) in comparison to negative NCs. In conclusion, formulated NCs are an ideal vehicle for passive targeting of drugs to tumor cells that may result in improved efficacy and reduced toxicity of encapsulated drug moiety.

Combustion characteristics of refuse-derived fuel pellets having varying plastic compositions.

The plastic fraction of refuse-derived fuel (RDF) pellets influences fuel's physico-chemical, and mechanical properties, which in turn might affect the combustion behavior of RDF. In the present study, the combustion behavior of different plastic fraction-simulated RDF pellets is reported. The simulated pellets were prepared based on the Indian commercial RDF composition. Initially, the plastic content varied from the existing fraction in Indian commercial RDF, i.e., 35% (RDF-1), to a lower plastic content of 5% (RDF-2). Physico-chemical characterization showed that a higher plastic fraction in RDF-1 reduces its pellet density by 25% as compared to RDF-2. The changes in RDF physico-chemical properties due to plastic variation are reflected in the RDF conversion process. Single-particle and packed-bed studies concluded that the lower density for higher plastic RDF-1 leads to lower conversion times (36%), and higher flame front speed (11%), which are desirable conditions for faster conversion. However, packed-bed studies also showed limitations regarding the utilization of high plastic RDF as RDF-1 has a narrow range of operating air mass flux due to the early advent of convective cooling of the bed. Finally, considering the constraints associated with the utilization of high plastic fraction (~ 35%) RDF and to maximize the effective utilization of plastic in RDF, a workable plastic fraction of 15% in RDF was proposed and tested for its combustion properties. RDF with 15% plastic showed faster conversion, higher flame front speed, and a wider range of operating air mass flux before the advent of convective cooling of the bed.