A Resonance-Sensitive Ultralow-Frequency Raman Mode in Twisted Bilayer Graphene.

Twisted bilayer graphene (tBLG) possesses intriguing physical properties including unconventional superconductivity, enhanced light-matter interaction due to the formation of van Hove singularities (vHS), and a divergence of density of states in the electronic band structures. The vHS energy band gap provides optical resonant transition channels that can be tuned by the twist angle and interlayer coupling. Raman spectroscopy provides rich information on the vHS structure of tBLG. Here, we report the discovery of an ultralow-frequency Raman mode at ∼49 cm-1 in tBLG. This mode is assigned to the combination of ZA (an out-of-plane acoustic phonon) and TA (a transverse acoustic phonon) phonons, and the Raman scattering is proposed to occur at the so-called mini-valley. This mode is found to be particularly sensitive to the change in vHS in tBLG. Our findings may deepen the understanding of Raman scattering in tBLG and help to reveal vHS-related electron-phonon interactions in tBLG.

Translational arrest and mRNA decay are independent activities of alphaherpesvirus virion host shutoff proteins.

The herpes simplex virus 1 (HSV1) virion host shutoff (vhs) protein is an endoribonuclease that regulates the translational environment of the infected cell, by inducing the degradation of host mRNA via cellular exonuclease activity. To further understand the relationship between translational shutoff and mRNA decay, we have used ectopic expression to compare HSV1 vhs (vhsH) to its homologues from four other alphaherpesviruses - varicella zoster virus (vhsV), bovine herpesvirus 1 (vhsB), equine herpesvirus 1 (vhsE) and Marek's disease virus (vhsM). Only vhsH, vhsB and vhsE induced degradation of a reporter luciferase mRNA, with poly(A)+ in situ hybridization indicating a global depletion of cytoplasmic poly(A)+ RNA and a concomitant increase in nuclear poly(A)+ RNA and the polyA tail binding protein PABPC1 in cells expressing these variants. By contrast, vhsV and vhsM failed to induce reporter mRNA decay and poly(A)+ depletion, but rather, induced cytoplasmic G3BP1 and poly(A)+ mRNA- containing granules and phosphorylation of the stress response proteins eIF2α and protein kinase R. Intriguingly, regardless of their apparent endoribonuclease activity, all vhs homologues induced an equivalent general blockade to translation as measured by single-cell puromycin incorporation. Taken together, these data suggest that the activities of translational arrest and mRNA decay induced by vhs are separable and we propose that they represent sequential steps of the vhs host interaction pathway.

SSA4 Mediates Cd Tolerance via Activation of the Cis Element of VHS1 in Yeast and Enhances Cd Tolerance in Chinese Cabbage.

Identifying key genes involved in Cadmium (Cd) response pathways in plants and developing low-Cd-accumulating cultivars may be the most effective and eco-friendly strategy to tackle the problem of Cd pollution in crops. In our previous study, Stressseventy subfamily A 4 (SSA4) was identified to be associated with Cd tolerance in yeast. Here, we investigated the mechanism of SSA4 in regulating Cd tolerance in yeast. ScSSA4 binds to POre Membrane 34 (POM34), a key component of nuclear pore complex (NPC), and translocates from the cytoplasm to the nucleus, where it regulates the expression of its downstream gene, Viable in a Hal3 Sit4 background 1 (VHS1), resulting in reduced Cd accumulation in yeast cells. Additionally, we identified a Chinese cabbage SSA4 gene, BrSSA4c, which could enhance the Cd tolerance in Chinese cabbage. This study offers new insights into the regulatory mechanisms of Cd tolerance in yeast, a model organism, and paves the way for the genetic enhancement of Cd tolerance in Chinese cabbage.

A New Design of a CMOS Vertical Hall Sensor with a Low Offset.

Vertical Hall sensors (VHSs), compatible with complementary metal oxide semiconductor (CMOS) technology, are used to detect magnetic fields in the plane of the sensor. In previous studies, their performance was limited by a large offset. This paper reports on a novel CMOS seven-contact VHS (7CVHS), which is formed by adding two additional contacts to a traditional five-contact VHS (5CVHS) to alleviate the offset. The offset voltage and offset magnetic field of the 7CVHS are reduced by 90.20% and 88.31% of those of the 5CVHS, respectively, with a 16.16% current-related sensitivity loss. Moreover, the size and positions of the contacts are optimized in standard GLOBALFOUNDRIES 0.18 µm BCDliteTM technology by scanning parameters using FEM simulations. The simulation data are analyzed in groups to study the influence of the size and contact positions on the current-related sensitivity, offset voltage, and offset magnetic field.

Radiographic and anatomical morphometric assessments of heart size in presumed healthy pet guinea pigs.

Cardiac disease in guinea pigs has been reported in the literature; however, reference intervals for normal radiographic heart size obtained using objective measurement methods have not been provided for this species. The aim of this prospective, reference interval study was to describe cardiac dimensions in presumed healthy guinea pigs using the vertebral heart scale (VHS) from thoracic radiographs, as described for dogs and cats. Furthermore, an anatomical study was carried out to compare the radiographic and anatomical findings. Thoracic radiographs were acquired in right lateral recumbency for 30, client-owned, conscious, presumed healthy guinea pigs and radiographs were acquired in left lateral recumbency for 10 presumed healthy guinea pigs as comparisons. The influence of sex, age, body weight (BW), and recumbency on the VHS and absolute cardiac measurements was investigated. The median (interquartile range; IQR) VHS was 7.4 (7.1-7.6). No differences emerged between the VHS measured in right versus left lateral recumbency (P = .41) or between sexes (P = .16). The VHS values were not influenced by age (P = .53) or BW (P = .26). The anatomical study was carried out on 10 guinea pig cadavers, and in situ and ex situ cardiac measurements were taken using a caliper. A median (IQR) 7.5 (7.2-8.0) VHS was assessed by this anatomical study. The reference intervals provided should be useful tools in the future for the radiographic interpretation of cardiac size in guinea pigs in clinical practice.

The role of artificial intelligence in clinical imaging and workflows.

Evidence-based medicine, outcomes management, and multidisciplinary systems are laying the foundation for radiology on the cusp of a new day. Environmental and operational forces coupled with technological advancements are redefining the veterinary radiologist of tomorrow. In the past several years, veterinary image volumes have exploded, and the scale of hardware and software required to support it seems boundless. The most dynamic trend within veterinary radiology is implementing digital information systems such as PACS, RIS, PIMS, and Voice Recognition systems. While the digitization of radiography imaging has significantly improved the workflow of the veterinary radiology assistant and radiologist, tedious, redundant tasks are abundant and mind-numbing. They can lead to errors with a significant impact on patient care.  Today, these boring and repetitious tasks continue to bog down patient throughput and workflow. Artificial intelligence, particularly machine learning, shows much promise to rocket the workflow and veterinary clinical imaging into a new day where the AI management of mundane tasks allows for efficiency so the radiologist can better concentrate on the quality of patient care. In this article, we briefly discuss the major subsets of artificial intelligence (AI) workflow for the radiologist and veterinary radiology assistant including image acquisition, segmentation and mensuration, rotation and hanging protocol, detection and prioritization, monitoring and registration of lesions, implementation of these subsets, and the ethics of utilizing AI in veterinary medicine.

Vertebral heart score to evaluate cardiac size in thoracic radiographs of 124 healthy rats (Rattus norvegicus).

Dilated cardiomyopathy is a relatively common disease in pet rats (Rattus norvegicus); however, there is a lack of radiographic references for the normal cardiac size in this species. The aim of this prospective, anatomical and reference interval study was to establish quantitative radiographic reference range measurements for the vertebral heart score (VHS) in rats. Right lateral (RL), ventrodorsal (VD), and dorsoventral (DV) radiographs of clinically healthy rats (n = 124) were evaluated. Measurements were performed by 2 expert readers who were unaware of signalment data. The mean values and references intervals of VHS were 7.7 and 7.0-8.5 for the RL, 7.5 and 6.6-8.6 for the VD, and 7.9 and 6.9-9.0 for the DV, with VHS values greater in males than in females. The measurements reported in this study can be used by the clinician as an objective tool to evaluate cardiac size in rats, in order to improve the diagnosis and treatment of cardiac diseases.

A radiographic study of breed-specific vertebral heart score and vertebral left atrial size in Chihuahuas.

Although echocardiography is the gold standard for the diagnosis of cardio-structural disease, thoracic radiography is a rapid, cost-effective, and widely accessible method for evaluating cardiac size in dogs. The vertebral heart score (VHS) and the vertebral left atrial size (VLAS) are established as objective measures of cardiomegaly on thoracic radiographs. However, several studies have shown significant variations in the VHS among different breeds. The Chihuahua is predisposed to both congenital and acquired cardiac diseases. The aim of this prospective, single-center, cross sectional study was thus to evaluate the VHS and the VLAS in healthy adult Chihuahua dogs. A total of 30 Chihuahuas were included. The VHS values in our sample population of Chihuahuas were 10.0 ± 0.6 (95% range, 8.9-11.0). This was significantly greater than the canine reference value of 9.7 ± 0.5 established by Buchanan and Bücheler (P = .002). The VLAS of Chihuahuas in our study was 1.8 ± 0.2 (95% range, 1.3-2.1). This was significantly lower than the values previously reported by Malcolm et al (2.07 ± 0.25; P = .0004). The VHS and the VLAS were not influenced by sex, body weight, short or long hair, and body condition score in normal Chihuahuas. Our results indicated that breed-specific reference values for radiographic VHS and VLAS are needed. In Chihuahuas, the values found in this study can be used as a normal reference in order to help avoid overinterpretation of cardiomegaly in these dogs.

Host shutoff activity of VHS and SOX-like proteins: role in viral survival and immune evasion.

BACKGROUND: Host shutoff refers to the widespread downregulation of host gene expression and has emerged as a key process that facilitates the reallocation of cellular resources for viral replication and evasion of host antiviral immune responses. MAIN BODY: The Herpesviridae family uses a number of proteins that are responsible for host shutoff by directly targeting messenger RNA (mRNA), including virion host shutoff (VHS) protein and the immediate-early regulatory protein ICP27 of herpes simplex virus types 1 (HSV-1) and the SOX (shutoff and exonuclease) protein and its homologs in Gammaherpesvirinae subfamilies, although these proteins are not homologous. In this review, we highlight evidence that host shutoff is promoted by the VHS, ICP27 and SOX-like proteins and that they also contribute to immune evasion. CONCLUSIONS: Further studies regarding the host shutoff proteins will not only contribute to provide new insights into the viral replication, expression and host immune evasion process, but also provide new molecular targets for the development of antiviral drugs and therapies.

Natural allelic variations of Saccharomyces cerevisiae impact stuck fermentation due to the combined effect of ethanol and temperature; a QTL-mapping study.

BACKGROUND: Fermentation completion is a major prerequisite in many industrial processes involving the bakery yeast Saccharomyces cerevisiae. Stuck fermentations can be due to the combination of many environmental stresses. Among them, high temperature and ethanol content are particularly deleterious especially in bioethanol and red wine production. Although the genetic causes of temperature and/or ethanol tolerance were widely investigated in laboratory conditions, few studies investigated natural genetic variations related to stuck fermentations in high gravity matrixes. RESULTS: In this study, three QTLs linked to stuck fermentation in winemaking conditions were identified by using a selective genotyping strategy carried out on a backcrossed population. The precision of mapping allows the identification of two causative genes VHS1 and OYE2 characterized by stop-codon insertion. The phenotypic effect of these allelic variations was validated by Reciprocal Hemyzygous Assay in high gravity fermentations (> 240 g/L of sugar) carried out at high temperatures (> 28 °C). Phenotypes impacted were mostly related to the late stage of alcoholic fermentation during the stationary growth phase of yeast. CONCLUSIONS: Our findings illustrate the complex genetic determinism of stuck fermentation and open new avenues for better understanding yeast resistance mechanisms involved in high gravity fermentations.

Multiple Posttranscriptional Strategies To Regulate the Herpes Simplex Virus 1 vhs Endoribonuclease.

The herpes simplex virus 1 (HSV-1) virion host shutoff (vhs) protein is an endoribonuclease that binds to the cellular translation initiation machinery and degrades associated mRNAs, resulting in the shutoff of host protein synthesis. Hence, its unrestrained activity is considered lethal, and it has been proposed that vhs is regulated by two other virus proteins, VP22 and VP16. We have found that during infection, translation of vhs requires VP22 but not the VP22-VP16 complex. Moreover, in the absence of VP22, vhs is not overactive against cellular or viral transcripts. In transfected cells, vhs was also poorly translated, correlating with the aberrant localization of its mRNA. Counterintuitively, vhs mRNA was predominantly nuclear in cells where vhs protein was detected. Likewise, transcripts from cotransfected plasmids were also retained in the same nuclei where vhs mRNA was located, while poly(A) binding protein (PABP) was relocalized to the nucleus in a vhs-dependent manner, implying a general block to mRNA export. Coexpression of VP16 and VP22 rescued the cytoplasmic localization of vhs mRNA but failed to rescue vhs translation. We identified a 230-nucleotide sequence in the 5' region of vhs that blocked its translation and, when transferred to a heterologous green fluorescent protein transcript, reduced translation without altering mRNA levels or localization. We propose that expression of vhs is tightly regulated by a combination of inherent untranslatability and autoinduced nuclear retention of its mRNA that results in a negative feedback loop, with nuclear retention but not translation of vhs mRNA being the target of rescue by the vhs-VP16-VP22 complex.IMPORTANCE A myriad of gene expression strategies has been discovered through studies carried out on viruses. This report concerns the regulation of the HSV-1 vhs endoribonuclease, a virus factor that is important for counteracting host antiviral responses by degrading their mRNAs but that must be regulated during infection to ensure that it does not act against and inhibit the virus itself. We show that regulation of vhs involves multifaceted posttranscriptional cellular and viral processes, including aberrant mRNA localization and a novel, autoregulated negative feedback loop to target its own and coexpressed mRNAs for nuclear retention, an activity that is relieved by coexpression of two other virus proteins, VP22 and VP16. These studies reveal the interplay of strategies by which multiple virus-encoded factors coordinate gene expression at the time that they are needed. These findings are broadly relevant to both virus and cellular gene expression.

Defining the Role of Stress Granules in Innate Immune Suppression by the Herpes Simplex Virus 1 Endoribonuclease VHS.

In response to virus-induced shutoff host protein synthesis, dynamic aggregates containing mRNA, RNA-binding proteins and translation factors termed stress granules (SGs) often accumulate within the cytoplasm. SGs typically form following phosphorylation and inactivation of the eukaryotic translation initiation factor 2α (eIF2α), a substrate of the double-stranded RNA (dsRNA)-activated kinase protein kinase R (PKR). The detection of innate immune sensors and effectors like PKR at SGs suggests a role in pathogen nucleic acid sensing. However, the functional importance of SGs in host innate responses is unclear and has primarily been examined in response to infection with select RNA viruses. During infection with the DNA virus herpes simplex virus 1 (HSV-1), the virus-encoded virion host shutoff (VHS) endoribonuclease is required to restrict interferon production, PKR activation, and SG formation, although the relationship between these activities remains incompletely understood. Here, we show that in cells infected with a VHS-deficient HSV-1 (ΔVHS) dsRNA accumulated and localized to SGs. Surprisingly, formation of dsRNA and its concentration at SGs was not required for beta interferon mRNA induction, indicating that suppression of type I interferon induction by VHS does not stem from its control of dsRNA accumulation. Instead, STING signaling downstream of cGMP-AMP synthase (cGAS)-dependent DNA sensing is required for beta interferon induction. In contrast, significantly less PKR activation is observed when SG assembly is disrupted by ISRIB, an inhibitor of phosphorylated eIF2α-mediated translation repression, or depleting SG scaffolding proteins G3BP1 or TIA1. This demonstrates that PKR activation is intimately linked to SG formation and that SGs form important hubs to potentiate PKR activation during infection.IMPORTANCE Formation of cytoplasmic stress granules that are enriched for innate immune sensors and effectors is suppressed during many viral infections. It is unclear, however, to what extent this is a side effect of viral efforts to maintain protein synthesis or intentional disruption of a hub for innate immune sensing. In this study, we utilize a herpes simplex virus 1 mutant lacking the RNA nuclease VHS which upon infection induces SGs, PKR activation, and beta interferon to address this question. We show that dsRNA is localized to SGs and that SGs can function to promote PKR activation in the context of a DNA virus infection, but we find no evidence to support their importance for interferon induction during HSV-1 infection.

Time-course study of the protection induced by an interferon-inducible DNA vaccine against viral haemorrhagic septicaemia in rainbow trout.

The highly effective DNA vaccines against diseases caused by fish rhabdoviruses in farmed fish consist of a DNA plasmid vector encoding the viral glycoprotein under the control of a constitutive cytomegalovirus promoter (CMV). Among others, attempts to improve efficacy and safety of these DNA vaccines have focused on regulatory elements of plasmid vectors, which play a major role in controlling expression levels of vaccine antigens. Depending on the context, use of a fish-derived promoter with minimal activity in mammalian cells could be preferable. Another aspect related to the CMV promoter is that constitutive expression of the vaccine antigen may lead to rapid elimination of antigen expressing cells in the fish and thereby potentially reduce the long-term effects of the vaccine. In this study, we compared DNA vaccines with the interferon-inducible Mx promoter from rainbow trout and the CMV promoter, respectively. Plasmid constructs encoding the enhanced green fluorescent protein (EGFP) were used for the in vitro analysis, whereas DNA vaccines encoding the glycoprotein (G) of the viral haemorrhagic septicaemia virus (VHSV) were applied for the in vivo examination. The in vitro analysis showed that while the DNA vaccine with the CMV promoter constitutively drove the expression of EGFP in both fish and human cell lines, the DNA vaccine with the Mx promoter inducibly enhanced the expression of EGFP in the fish cell line. To address the impact on protection, a time-course model was followed as suggested by Kurath et al. (2006), where vaccinated fish were challenged with VHSV at 2, 8 and 78 weeks post-vaccination (wpv). The DNA vaccine with the CMV promoter protected at all times, while vaccination with the DNA vaccine containing the Mx promoter only protected the fish at 8 wpv. However, following induction with Poly (I:C) one week before the challenge, high protection was also evident at 2 wpv. In conclusion, the results revealed a more fish host dependent activity of the trout Mx promoter compared to the traditionally used cross species-active CMV promoter, but improvements will be needed for its application in DNA vaccines to ensure long term protection.

Herpes Simplex Virus 1 Abrogates the cGAS/STING-Mediated Cytosolic DNA-Sensing Pathway via Its Virion Host Shutoff Protein, UL41.

Cyclic GMP-AMP synthase (cGAS) is a key DNA sensor capable of detecting microbial DNA and activating the adaptor protein stimulator of interferon genes (STING), leading to interferon (IFN) production and host antiviral responses. Cells exhibited reduced type I IFN production in response to cytosolic DNA in the absence of cGAS. Although the cGAS/STING-mediated DNA-sensing signal is crucial for host defense against many viruses, especially for DNA viruses, few viral components have been identified to specifically target this signaling pathway. Herpes simplex virus 1 (HSV-1) is a DNA virus that has evolved multiple strategies to evade host immune responses. In the present study, we found that HSV-1 tegument protein UL41 was involved in counteracting the cGAS/STING-mediated DNA-sensing pathway. Our results showed that wild-type (WT) HSV-1 infection could inhibit immunostimulatory DNA-induced activation of the IFN signaling pathway compared with the UL41-null mutant virus (R2621), and ectopic expression of UL41 decreased cGAS/STING-mediated IFN-ß promoter activation and IFN-ß production. Further study indicated that UL41 reduced the accumulation of cGAS to abrogate host recognition of viral DNA. In addition, stable knockdown of cGAS facilitated the replication of R2621 but not WT HSV-1. For the first time, HSV-1 UL41 was demonstrated to evade the cGAS/STING-mediated DNA-sensing pathway by degrading cGAS via its RNase activity.IMPORTANCE HSV-1 is well known for its ability to evade host antiviral responses and establish a lifelong latent infection while triggering reactivation and lytic infection under stress. Currently, whether HSV-1 evades the cytosolic DNA sensing and signaling is still poorly understood. In the present study, we found that tegument protein UL41 targeted the cGAS/STING-mediated cellular DNA-sensing pathway by selectively degrading cGAS mRNA. Knockdown of endogenous cGAS could facilitate the replication of R2621 but not WT HSV-1. Furthermore, UL41 was shown for the first time to act directly on cGAS. Findings in this study could provide new insights into the host-virus interaction and help develop new approaches against HSV-1.

Generation and characterization of UL41 null pseudorabies virus variant in vitro and in vivo.

BACKGROUND: The alphaherpesvirus virion host shutoff (vhs) gene, UL41, can induce degradation of host mRNAs and shut off host protein synthesis. The roles of vhs in HSV-1 and HSV-2 have been studied extensively in previous studies, however, relatively little is known about the vhs protein of PRV. METHODS: A novel method combining CRISPR/Cas9 and Gibson assembly was developed to generate UL41 null PRV variant. The properties of UL41 null PRV in vitro and in vivo were further characterized. And the vhs activity of UL41 protein of PRV variant was evaluated by luciferase assay, Western-blot and RT-qPCR. RESULTS: Gibson assembly based on homologous recombination can accomplish one-step insertion of viral DNA fragments into donor plasmids efficiently (> 80%). Cas9/gRNA further largely enhanced the efficiency of homologous recombination. Using this method we were able to rapidly generate the UL41 null and revertant viruses of PRV variant. Compared to wild type (JS-2012), the UL41 null virus showed significantly smaller plaques and lower titers in Vero cells and impaired lethality and neuroinvasion in mice. Further the UL41 protein from different PRV strains exhibited unequal vhs activity in vitro, which of JS-2012 showed significantly weaker vhs activity than that of European-American strains. In addition UL41 null virus can also significantly decrease the expression of host genes during the early period of infection, which suggests other viral factors may be also involved in host shutoff. CONCLUSIONS: CRISPR/Cas9 combined with Gibson assembly efficiently generated UL41 null PRV. Compared to wild type, UL41 null PRV showed impaired both replication capability in vitro and neuroinvasion in vivo. Further UL41 protein of PRV variant showed significantly weaker vhs activity than that of PRV SC (European-American-like strain), suggesting the deficiency of vhs activity by the PRV variant UL41 protein.

Determination of vertebral heart score in three species of Spider monkey (Ateles fusciceps, A. hybridus and A. paniscus).

BACKGROUND: The vertebral heart score (VHS) is a method of evaluation of cardiac size well documented in domestic mammals and in other primate species, and the aim of this study was to determine the VHS in three species of Spider monkey. METHODS: In this retrospective study, right lateral radiographs of thirty clinically well animals were reviewed and VHS determined. The species included were Ateles fusciceps (n=17), Ateles hybridus (n=8) and Ateles paniscus (n=5). RESULTS: The VHS was found to vary between species and was 9.73±0.81 for A. fusciceps, 10.53±0.37 for A. hybridus and 10.45±0.27 for A. paniscus. CONCLUSIONS: The observed values appear consistent with values determined for other primate species. There was statistically significant variation noted between species, and so VHS should be considered species-specific in this genus. The values determined may be of benefit in objectively evaluating cardiac size in the species investigated.

The Vps27/Hrs/STAM (VHS) Domain of the Signal-transducing Adaptor Molecule (STAM) Directs Associated Molecule with the SH3 Domain of STAM (AMSH) Specificity to Longer Ubiquitin Chains and Dictates the Position of Cleavage.

The deubiquitinating enzyme associated molecule with the SH3 domain of STAM (AMSH) is crucial for the removal of ubiquitin molecules during receptor-mediated endocytosis and lysosomal receptor sorting. AMSH interacts with signal transducing adapter molecule (STAM) 1 or 2, which enhances the activity of AMSH through an unknown mechanism. This stimulation is dependent on the ubiquitin-interacting motif of STAM. Here we investigate the specific mechanism of AMSH stimulation by STAM proteins and the role of the STAM Vps27/Hrs/STAM domain. We show that, in the presence of STAM, the length of the ubiquitin chains affects the apparent cleavage rate. Through measurement of the chain cleavage kinetics, we found that, although the kcat of Lys(63)-linked ubiquitin chain cleavage was comparable for di- and tri-ubiquitin, the Km value was lower for tri-ubiquitin. This increased affinity for longer chains was dependent on the Vps27/Hrs/STAM domain of STAM and required that the substrate ubiquitin chain contain homogenous Lys(63)-linkages. In addition, STAM directed AMSH cleavage toward the distal isopeptide bond in tri-ubiquitin chains. Finally, we generated a structural model of AMSH-STAM to show how the complex binds Lys(63)-linked ubiquitin chains and cleaves at the distal end. These data show how a deubiquitinating enzyme-interacting protein dictates the efficiency and specificity of substrate cleavage.

TOM1 is a PI5P effector involved in the regulation of endosomal maturation.

Phosphoinositides represent a major class of lipids specifically involved in the organization of signaling cascades, maintenance of the identity of organelles and regulation of multiple intracellular trafficking steps. We previously reported that phosphatidylinositol 5-monophosphate (PI5P), produced by the Shigella flexneri phosphatase IpgD, is implicated in the endosomal sorting of the epidermal growth factor receptor (EGFR). Here, we show that the adaptor protein TOM1 is a new direct binding partner of PI5P. We identify the domain of TOM1 involved in this interaction and characterize the binding motif. Finally, we demonstrate that the recruitment of TOM1 by PI5P on signaling endosomes is responsible for the delay in EGFR degradation and fluid-phase bulk endocytosis. Taken together, our data strongly suggest that PI5P enrichment in signaling endosomes prevents endosomal maturation through the recruitment of TOM1, and point to a new function of PI5P in regulating discrete maturation steps in the endosomal system.

Viral and bacterial septicaemic infections modulate the expression of PACAP splicing variants and VIP/PACAP receptors in brown trout immune organs.

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) and PACAP-Related Peptide (PRP) are structurally similar peptides encoded in the same transcripts. Their transcription has been detected not only in the brain but also in a wide range of peripheral tissues, even including organs of the immune system. PACAP exerts pleiotropic activities through G-protein coupled membrane receptors: the PACAP-specific PAC-1 and the VPAC-1 and VPAC-2 receptors that exhibit similar affinities for the Vasoactive Intestinal Peptide (VIP) and PACAP. Recent findings added PACAP and its receptors to the growing list of mediators that allow cross-talk between the nervous, endocrine and immune systems in fish. In this study the expression of genes encoding for PACAP and PRP, as well as VIP/PACAP receptors was studied in laboratory-reared brown trout (Salmo trutta) after septicaemic infections. Respectively Viral Haemorrhagic Septicaemia Virus (VHSV-Ia) or the Gram-negative bacterium Yersinia ruckeri (ser. O1 - biot. 2) were used in infection challenges. Kidney and spleen, the teleost main lymphopoietic organs, were sampled during the first two weeks post-infection. RT-qPCR analysis assessed specific pathogens burden and gene expression levels. PACAP and PRP transcription in each organ was positively correlated to the respective pathogen burden, assessed targeting the VHSV-glycoprotein or Y. ruckeri 16S rRNA. Results showed as the transcription of PACAP splicing variants and VIP/PACAP receptors is modulated in these organs during an acute viral and bacterial septicaemic infections in brown trout. These gene expression results provide clues as to how the PACAP system is modulated in fish, confirming an involvement during active immune responses elicited by both viral and bacterial aetiological agents. However, further experimental evidence is still required to fully elucidate and characterize the role of PACAP and PRP for an efficient immune response against pathogens.

Vertebral heart size and vertebral left atrial size reference ranges in healthy Miniature Schnauzers.

OBJECTIVE: The inclusion of vertebral heart score (VHS) and, more recently, the inclusion of the vertebral left atrial size (VLAS) in radiographic evaluation have become important screening tools for identifying dogs with occult cardiac disease. Several recent papers have shown there are interbreed variations in the VHS reference range. Our hypothesis is that the Miniature Schnauzer would also have a higher reference range for its VHS. ANIMALS: The electronic medical records of IDEXX Telemedicine Consultants were searched for Miniature Schnauzers undergoing thoracic radiographs between March 1, 2022, and February 28, 2023. METHODS: Dogs were included if they had 3 view thoracic radiographs performed and no evidence of cardiopulmonary disease was detected. Dogs with incomplete radiographic studies or cardiac or extracardiac disease were excluded. The VHS and VLAS measurements were performed by 2 board-certified cardiologists independent of one another. RESULTS: A total of 1,000 radiographs were obtained of which 272 were included for the study. The overall range for the VHS in this cohort was 9.68 to 12.07 with a median of 10.9. For VLAS measurements, a range of 1.71 to 2.4 was documented with a median of 2.0. CLINICAL RELEVANCE: The VHS for Miniature Schnauzers without cardiac disease was confirmed to be higher than the canine reference range.