Miracle-dz/UniFAR-FTUs huggingface dataset:
Study of Legionella Effector Domains Revealed Novel and Prevalent Phosphatidylinositol 3-Phosphate Binding Domains

Legionella pneumophila and other Legionella species replicate intracellularly using the Icm/Dot type IV secretion system. In L. pneumophila this system translocates >300 effectors into host cells and in the Legionella genus thousands of effectors were identified, the function of most of which is unknown. ABSTRACT Legionella pneumophila and other Legionella species replicate intracellularly using the Icm/Dot type IV secretion system. In L. pneumophila this system translocates >300 effectors into host cells and in the Legionella genus thousands of effectors were identified, the function of most of which is unknown. Fourteen L. pneumophila effectors were previously shown to specifically bind phosphoinositides (PIs) using dedicated domains. We found that PI-binding domains of effectors are usually not homologous to one another; they are relatively small and located at the effectors' C termini. We used the previously identified Legionella effector domains (LEDs) with unknown function and the above characteristics of effector PI-binding domains to discover novel PI-binding LEDs. We identified three predicted PI-binding LEDs that are present in 14 L. pneumophila effectors and in >200 effectors in the Legionella genus. Using an in vitro protein-lipid overlay assay, we found that 11 of these L. pneumophila effectors specifically bind phosphatidylinositol 3-phosphate (PI3P), almost doubling the number of L. pneumophila effectors known to bind PIs. Further, we identified in each of these newly discovered PI3P-binding LEDs conserved, mainly positively charged, amino acids that are essential for PI3P binding. Our results indicate that Legionella effectors harbor unique domains, shared by many effectors, which directly mediate PI3P binding.
Dispersion of Boron Nitride Nanotubes by Pluronic Triblock Copolymer in Aqueous Solution

Boron nitride nanotubes (BNNTs) have been of interest for their excellent thermal, electrical, and mechanical properties, and they have a broad spectrum of potential applications, such as in piezoelectric materials, reinforcement of materials, and electrothermal insulation materials. For practical use of BNNTs, it is desirable to disperse them in aqueous solution, which improves convenience of handling. However, it is still difficult to make a homogenous and stable BNNT dispersion in aqueous solution, due to their strong van der Waals interactions and hydrophobic surface. To solve these problems, we used Pluronic P85 and F127, which have both hydrophilic groups and hydrophobic groups. Here, we report the wrapped structure of a Pluronic polymer-BNNT dispersion by using small-angle neutron scattering, UV–Vis spectroscopy, thermogravimetric analysis, and atomic force microscopy.
The Evolution of Aging With Pride—National Health, Aging, and Sexuality/Gender Study: Illuminating the Iridescent Life Course of LGBTQ Adults Aged 80 Years and Older in the United States

Aging with Pride: National Health, Aging, and Sexuality/Gender Study is the first federally funded study addressing aging among LGBTQ older adults throughout the United States. This article examines the evolution of this landmark study and explores the well-being of LGBTQ adults aged 80 years and older (n = 200), the most underrepresented group in the field. Based on the Iridescent Life Course, we examined the diverse, intersectional nature of LGBTQ older adults’ lives, finding high levels of education and poverty. Microaggressions were negatively associated with quality-of-life and positively associated with poor physical and mental health; the inverse relationship was found with mastery. When the oldest encountered risks, it resulted in greater vulnerability. This longitudinal study is assessing trajectories in aging over time using qualitative, quantitative, and biological data and testing evidence-based culturally responsive interventions for LGBTQ older adults. Research with LGBTQ oldest adults is much needed before their stories are lost to time.
CAP2 deficiency delays myofibril actin cytoskeleton differentiation and disturbs skeletal muscle architecture and function

Significance Displaced myofibrils termed ring fibers have been reported for human congenital myopathies and for myopathies of unknown etiology, but their natural cause largely remained elusive. Cyclase-associated protein 2 (CAP2) is an actin regulator with unknown physiological function. We identified CAP2 as a regulator for the exchange of α-actin isoforms during myofibril differentiation. In CAP2 mutant mice, myofibrils remained in an undifferentiated stage at the onset of excessive voluntary muscle contractions, predisposing them to form ring fibers that compromise skeletal muscle function. Our data confirm that subtle cytoskeletal changes can cause ring fibers. Moreover, they led us to propose that CAP2 inactivation contributes to defects of human 6p22.3 deletion syndrome such as developmental delay and hypotonia. Actin filaments (F-actin) are key components of sarcomeres, the basic contractile units of skeletal muscle myofibrils. A crucial step during myofibril differentiation is the sequential exchange of α-actin isoforms from smooth muscle (α-SMA) and cardiac (α-CAA) to skeletal muscle α-actin (α-SKA) that, in mice, occurs during early postnatal life. This “α-actin switch” requires the coordinated activity of actin regulators because it is vital that sarcomere structure and function are maintained during differentiation. The molecular machinery that controls the α-actin switch, however, remains enigmatic. Cyclase-associated proteins (CAP) are a family of actin regulators with largely unknown physiological functions. We here report a function for CAP2 in regulating the α-actin exchange during myofibril differentiation. This α-actin switch was delayed in systemic CAP2 mutant mice, and myofibrils remained in an undifferentiated stage at the onset of the often excessive voluntary movements in postnatal mice. The delay in the α-actin switch coincided with the onset of motor function deficits and histopathological changes including a high frequency of type IIB ring fibers. Our data suggest that subtle disturbances of postnatal F-actin remodeling are sufficient for predisposing muscle fibers to form ring fibers. Cofilin2, a putative CAP2 interaction partner, has been recently implicated in myofibril actin cytoskeleton differentiation, and the myopathies in cofilin2 and CAP2 mutant mice showed striking similarities. We therefore propose a model in which CAP2 and cofilin2 cooperate in actin regulation during myofibril differentiation.
Cost of transport is a repeatable trait but is not determined by mitochondrial efficiency in zebrafish (Danio rerio)

ABSTRACT The energy used to move a given distance (cost of transport; CoT) varies significantly between individuals of the same species. A lower CoT allows animals to allocate more of their energy budget to growth and reproduction. A higher CoT may cause animals to adjust their movement across different environmental gradients to reduce energy allocated to movement. The aim of this project was to determine whether CoT is a repeatable trait within individuals, and to determine its physiological causes and ecological consequences. We found that CoT is a repeatable trait in zebrafish (Danio rerio). We rejected the hypothesis that mitochondrial efficiency (P/O ratios) predicted CoT. We also rejected the hypothesis that CoT is modulated by temperature acclimation, exercise training or their interaction, although CoT increased with increasing acute test temperature. There was a weak but significant negative correlation between CoT and dispersal, measured as the number of exploration decisions made by fish, and the distance travelled against the current in an artificial stream. However, CoT was not correlated with the voluntary speed of fish moving against the current. The implication of these results is that CoT reflects a fixed physiological phenotype of an individual, which is not plastic in response to persistent environmental changes. Consequently, individuals may have fundamentally different energy budgets as they move across environments, and may adjust movement patterns as a result of allocation trade-offs. It was surprising that mitochondrial efficiency did not explain differences in CoT, and our working hypothesis is that the energetics of muscle contraction and relaxation may determine CoT. The increase in CoT with increasing acute environmental temperature means that warming environments will increase the proportion of the energy budget allocated to locomotion unless individuals adjust their movement patterns. Summary: The metabolic cost of movement is fixed within individuals, but differences between individuals mean that movement affects energy budgets differentially, and fish with a high cost may alter dispersal patterns.
Extracellular and intracellular sphingosine‐1‐phosphate distinctly regulates exocytosis in chromaffin cells

Sphingosine‐1‐phosphate (S1P) is an essential bioactive sphingosine lipid involved in many neurological disorders. Sphingosine kinase 1 (SphK1), a key enzyme for S1P production, is concentrated in presynaptic terminals. However, the role of S1P/SphK1 signaling in exocytosis remains elusive. By detecting catecholamine release from single vesicles in chromaffin cells, we show that a dominant negative SphK1 (SphK1DN) reduces the number of amperometric spikes and increases the duration of foot, which reflects release through a fusion pore, implying critical roles for S1P in regulating the rate of exocytosis and fusion pore expansion. Similar phenotypes were observed in chromaffin cells obtained from SphK1 knockout mice compared to those from wild‐type mice. In addition, extracellular S1P treatment increased the number of amperometric spikes, and this increase, in turn, was inhibited by a selective S1P3 receptor blocker, suggesting extracellular S1P may regulate the rate of exocytosis via activation of S1P3. Furthermore, intracellular S1P application induced a decrease in foot duration of amperometric spikes in control cells, indicating intracellular S1P may regulate fusion pore expansion during exocytosis. Taken together, our study represents the first demonstration that S1P regulates exocytosis through distinct mechanisms: extracellular S1P may modulate the rate of exocytosis via activation of S1P receptors while intracellular S1P may directly control fusion pore expansion during exocytosis.
Dural Calcitonin Gene-Related Peptide Produces Female-Specific Responses in Rodent Migraine Models

Migraine is the second leading cause for disability worldwide and the most common neurological disorder. It is also three times more common in women; reasons for this sex difference are not known. Using preclinical behavioral models of migraine, we show that application of calcitonin gene-related peptide (CGRP) to the rat dura mater produces cutaneous periorbital hypersensitivity. Surprisingly, this response was observed only in females; dural CGRP at doses from 1 pg to 3.8 μg produce no responses in males. In females, dural CGRP causes priming to a pH 7.0 solution after animals recover from the initial CGRP-induced allodynia. Dural application of interleukin-6 causes acute responses in males and females but only causes priming to subthreshold dural CGRP (0.1 pg) in females. Intracisternal application of BDNF also causes similar acute hypersensitivity responses in males and females but only priming to subthreshold dural CGRP (0.1 pg) in females. Females were additionally primed to a subthreshold dose of the NO-donor sodium nitroprusside (0.1 mg/kg) following dural CGRP. Finally, the sexually dimorphic responses to dural CGRP were not specific to rats as similar female-specific hypersensitivity responses were seen in mice, where increased grimace responses were also observed. These data are the first to demonstrate that CGRP-induced headache-like behavioral responses at doses up to 3.8 μg are female-specific both acutely and following central and peripheral priming. These data further implicate dural CGRP signaling in the pathophysiology of migraine and propose a model where dural CGRP-based mechanisms contribute to the sexual disparity of this female-biased disorder. SIGNIFICANCE STATEMENT Calcitonin gene-related peptide (CGRP) has long been implicated in the pathophysiology of migraine, and CGRP-based therapeutics are efficacious for the treatment of migraine in humans. However, the location of action for CGRP in migraine remains unclear. We show here that application of CGRP to the cranial meninges causes behavioral responses consistent with headache in preclinical rodent models. Surprisingly, however, these responses are only observed in females. Acute responses to meningeal CGRP are female-specific and sensitization to CGRP after two distinct stimuli are also female-specific. These data implicate the dura mater as a primary location of action for CGRP in migraine and suggest that female-specific mechanisms downstream of CGRP receptor activation contribute to the higher prevalence of migraine in women.
Uncontrollable chronic stress affects eating behavior in rats

Abstract The issue of whether the decrease in food intake induced by inescapable shock is due to the uncontrollability of the stressor or the shock per se has not yet been settled. Besides, whether food intake is differentially affected by an uncontrollable chronic stressor has been explored only by a few studies. Thus, we evaluated the effects of chronic escapable or inescapable electric shocks on eating behavior. Rats were exposed to shock sessions for 20 days in two occasions separated by baseline sessions with no shock in an ABAB design. Results showed a reduction in food and water intake and body weight gain during stress periods, especially with inescapable shocks. The findings support a close link between learned helplessness, chronic stress, and eating behavior.
AllergoOncology: Expression platform development and functional profiling of an anti‐HER2 IgE antibody

Efficient strategies for generation of recombinant IgE class antibodies at high-enough yields for pre-clinical screening and mechanistic evaluations remain challenging. We present a transient expression application for the rapid production of recombinant IgE, exemplified with the cloning and functional characterization of a humanised IgE antibody bearing the variable region sequences of the anti-HER2/ neu antibody trastuzumab. Material was generated with high transfection efficiency, in small culture volumes within 7-9-days from design to purified material, and at sufficient yields for functional studies. The antibody conserved the trastuzumab Fab-mediated recognition of the target antigen, and IgE Fc attributes to recognise Fc εRs and to potentiate immune cell-mediated effector functions. Moreover, mast cell and basophil activation tests confirmed lack of activation with IgE in the absence of cross-linking stimuli, supporting potential safe administration in humans. Our study facilitates fast generation of IgE antibodies for speedy screening and functional characterisation and can be readily applied to the design and evaluation of IgE antibodies in Allergy and AllergoOncology. Antibody cloning: We employed polymerase incomplete primer extension (PIPE) PCR cloning and enzyme-free assembly of DNA fragments. The amino acid sequences of the trastuzumab heavy and light variable regions were obtained from the DrugBank database (www.drugbank.ca), translated in nucleotide sequences and manually codon optimized for a human expression host. Optimized sequences were synthesized using GeneArt Gene Synthesis (Thermo Fischer Scientific UK). The DNA sequences of the variable regions were previously reported 1 . The variable region fragments of trastuzumab were cloned into pVitro1-hygro-mcs dual expression vector containing pre-cloned cassettes of the human epsilon chain constant region and kappa light chain constant region cassettes using the polymerase incomplete primer extension (PIPE) PCR cloning as previously described 2 . The PIPE PCR primers used for the fragment amplification are listed in Supplementary Table S1. Briefly, PIPE PCR was performed using pVitro1 plasmid as a template in order to generate two linear PCR fragments with 5` PIPE overhangs, and commercially-generated trastuzumab variable region fragments, in order to generate a variable light (VL) and heavy (VH) region fragments with 5` PIPE overhangs. Correct sizes of the DNA fragments were confirmed with agarose electrophoresis. Anti-HER2 IgE expression was carried out transiently in Expi293F cells using small volume suspension culture (30 mL) for ca. 7-9 days to achieve peak antibody concentrations of ca. 70-80 µg/mL in the culture supernatant. Purification and The antibody was purified using KappaSelect affinity
Biochemical synthesis of gold nanoparticles from leaf protein of Nicotiana tabacum L. cv. xanthi and their physiological, developmental, and ROS scavenging responses on tobacco plant under stress conditions.

The stress conditions imposed by the impact of metal and non-metal oxide nanoparticles over plant systems enhances the synthesis of reactive oxygen species (ROS), resulting in oxidative damage at cellular level. The objective of this study was to synthesise the gold nanoparticles (GNps) from the leaves protein of Nicotiana tabacum L. cv. xanthi, its characterisation, and response on plant physiology and ROS scavenging activity on plants after exposure to different stresses. The authors have treated N. tabacum L. cv. xanthi plants with 100, 200, 300, 400, and 500 ppm biochemically synthesised GNps and examined physiological as well as biochemical changes. Results showed that biochemically synthesised GNps exposure significantly increased the seed germination (P < 0.001), root (P < 0.001), shoot growth (P < 0.001), and antioxidant ability (P < 0.05) of plants depending on bioengineered GNPs concentrations. Low concentrations (200-300 ppm) of GNps boosted growth by ∼50% and significantly increase in photosynthetic parameters such as total chlorophyll content (P < 0.05), membrane ion leakage (P < 0.05) as well as malondialdehyde (P < 0.05) content with respect to untreated plants under stress conditions. The high concentration (400-500 ppm) of GNps affected these parameters in a negative manner. The total antioxidant activity was also elevated in the exposed plants in a dose-dependent manner.
Conformational insights into the inhibitory mechanism of phyto-compounds against Src kinase family members implicated in psoriasis

Abstract Psoriasis is a chronic immune mediated disorder of the skin. There is growing evidence that the Src family tyrosine kinases (SFK) are highly upregulated in psoriasis. The SFK are the key components of the signaling pathways triggering cell growth and differentiation in addition to the immune cascades. In the current work, the interactions between SFK and selective phyto-compounds were studied using molecular docking approach. Based on the results of docking and binding energy calculations quercetin was identified as potential lead compound. To get a deeper insight into the binding of quercetin with the SFK, a combined molecular dynamics and binding free energy calculations were performed. The binding of quercetin disrupted the intra-molecular contacts making the SFK compact except Src kinase. The MM/PBSA free energy decomposition analysis highlighted the significance of hydrophobic and polar residues which are involved in the binding of quercetin. An experimental validation was carried out against the activated forms of Fyn, Lyn and Src kinases, the top three proteins which showed high preference for quercetin. The flow cytometry analysis showed that the expression levels of Fyn, Lyn and Src kinases were dramatically increased in HaCaT cells. However, the treatment of quercetin at the concentration of 51.65 µM for 24 h markedly decreased their expression in HaCaT cells. Besides, similar results were also observed when the HaCaT cells were treated with the kinase inhibitor Ponitinib (1.43 µM) for 24 h. Communicated by Ramaswamy H. Sarma
Galectin‐3 aggravates ox‐LDL‐induced endothelial dysfunction through LOX‐1 mediated signaling pathway

Galectin‐3, a biomarker linking oxidative stress and inflammation, participates in different mechanisms related to atherothrombosis, such as inflammation, proliferation, or macrophage chemotaxis. Accumulating evidence indicates that galectin‐3 may also promote atherogenesis through inducing endothelial dysfunction. Lectin‐like oxidized low‐density lipoprotein (oxLDL) receptor‐1 (LOX‐1), a receptor for oxLDL uptake, contributes to oxLDL‐induced endothelial dysfunction. Whether galectin‐3 induces endothelial dysfunction through modulation of LOX‐1‐mediated signaling remains unclear. In the present study, we explored the mechanisms underlying galectin‐3 enhanced cytotoxicity of oxLDL in human umbilical vein endothelial cells (HUVECs) and the role of LOX‐1. Incubation of HUVECs with galectin‐3 increased the expression of LOX‐1 in RNA and protein levels. In addition, the expression of LOX‐1 induced by oxLDL was promoted by galectin‐3. However, pretreatment of LOX‐1 antibody reduced LOX‐1 mRNA expression level in cells with oxLDL plus galectin‐3 incubation. Compared to cells treated with oxLDL alone, reactive oxygen species (ROS) generation via nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation and subsequent activation of p38 mitogen‐activated protein kinases followed by nuclear factor kappa B (NF‐κB) activation and related inflammatory responses including adhesion molecule expression, adhesiveness of monocytic cells, and IL‐8 release were also aggravated in cells treated with galectin‐3 combined with oxLDL. Compared to cells treated with galectin‐3 plus oxLDL group. We found that LOX‐1 antibody mitigated NADPH oxidase activity, p‐38 up‐regulation, NF‐κB activation, and proinflammatory responses in cells treated with galectin‐3 combined with oxLDL. We conclude that galectin‐3 enhances endothelial LOX‐1 expression and propose a new mechanism by which galectin‐3 may promote endothelial dysfunction by inducing inflammation via LOX‐1/ROS/p38/NF‐κB‐mediated signaling pathway.
Towards controlled terminology for reporting germline cancer susceptibility variants: an ENIGMA report

The vocabulary currently used to describe genetic variants and their consequences reflects many years of studying and discovering monogenic disease with high penetrance. With the recent rapid expansion of genetic testing brought about by wide availability of high-throughput massively parallel sequencing platforms, accurate variant interpretation has become a major issue. The vocabulary used to describe single genetic variants in silico, in vitro, in vivo and as a contributor to human disease uses terms in common, but the meaning is not necessarily shared across all these contexts. In the setting of cancer genetic tests, the added dimension of using data from genetic sequencing of tumour DNA to direct treatment is an additional source of confusion to those who are not experienced in cancer genetics. The language used to describe variants identified in cancer susceptibility genetic testing typically still reflects an outdated paradigm of Mendelian inheritance with dichotomous outcomes. Cancer is a common disease with complex genetic architecture; an improved lexicon is required to better communicate among scientists, clinicians and patients, the risks and implications of genetic variants detected. This review arises from a recognition of, and discussion about, inconsistencies in vocabulary usage by members of the ENIGMA international multidisciplinary consortium focused on variant classification in breast-ovarian cancer susceptibility genes. It sets out the vocabulary commonly used in genetic variant interpretation and reporting, and suggests a framework for a common vocabulary that may facilitate understanding and clarity in clinical reporting of germline genetic tests for cancer susceptibility.