bluuebunny/crossref_metadata_2025 huggingface dataset:
Trade-offs constrain adaptive pathways to T6 survival
Authors: MacGillivray Kathryn A. , Ng Siu Lung , Wiesenfeld Sophia , Guest Randi L. , Jubery Tahrima , Silhavy Thomas J. , Ratcliff William C. , Hammer Brian K.
Published: September 2022

Many microbial communities are characterized by intense competition for nutrients and space. One way for an organism to gain control of these resources is by eliminating nearby competitors. The Type VI Secretion System (T6) is a nano-harpoon used by many bacteria to inject toxins into neighboring cells. While much is understood about mechanisms of T6-mediated toxicity, little is known about the ways that competitors can defend themselves against this attack, especially in the absence of their own T6. Here we use directed evolution to examine the evolution of T6 resistance, subjecting eight replicate populations of Escherichia coli to T6 attack by Vibrio cholerae. Over ~500 generations of competition, the E. coli evolved to survive T6 attack an average of 27-fold better than their ancestor. Whole genome sequencing reveals extensive parallel evolution. In fact, we found only two pathways to increased T6 survival: apaH was mutated in six of the eight replicate populations, while the other two populations each had mutations in both yejM and yjeP. Synthetic reconstruction of individual and combined mutations demonstrate that yejM and yjeP are synergistic, with yejM requiring the mutation in yejP to provide a benefit. However, the mutations we identified are pleiotropic, reducing cellular growth rates, and increasing susceptibility to antibiotics and elevated pH. These trade-offs underlie the effectiveness of T6 as a bacterial weapon, and help us understand how the T6 shapes the evolution of bacterial interactions.

Significance

Bacteria are the most abundant organisms on Earth and often live in dense, diverse communities, where they interact with each other. One of the most common interactions is antagonism. While most research has focused on diffusible toxins (e.g., antibiotics), bacteria have also evolved a contact-dependent nano-harpoon, the Type VI Secretion System (T6), to kill neighboring cells and compete for resources. While the co-evolutionary dynamics of antibiotic exposure is well understood, no prior work has examined how targets of T6 evolve resistance. Here, we use experimental evolution to observe how an Escherichia coli target evolves resistance to T6 when it is repeatedly competing with a Vibrio cholerae killer. After 30 rounds of competition, we identified mutations in three genes that improve E. coli survival, but found that these mutations come at a cost to other key fitness components. Our findings provide new insight into how contact-dependent antagonistic interaction drives evolution in a polymicrobial community.
Derivation of a minimal functional XIST by combining human and mouse interaction domains
Authors: Navarro-Cobos Maria Jose , Morales-Guzman Suria Itzel , Baldry Sarah E.L. , Brown Carolyn J.
Published: September 2022

XIST is a 17-19 kb long non-coding RNA critical for X-chromosome inactivation. Tandem repeats within the RNA serve as functional domains involved in the cis-limited recruitment of heterochromatic changes and silencing. To explore the sufficiency of these domains while generating a functional miniXIST for targeted silencing approaches, we tested inducible constructs integrated into 8p in a male cell line. Previous results suggested silencing could be accomplished with a transgene comprised of the repeat A which is highly conserved and critical for silencing; the repeat F which overlaps regulatory elements and the repeat E region which contributes to XIST localization by binding proteins such as CIZ1. As PRC1 is recruited through HNRNPK binding of repeats B - C - D we included a second ‘miniXIST’ comprising AFE with the mouse PID, a 600-nucleotide region of repeat B and C. Silencing of nearby genes was possible with and without PID, however, silencing more distally required the addition of PID. The recruitment of heterochromatic marks, evaluated by IF combined with RNA FISH, revealed that the AFE domains were sufficient only for the recruitment of CIZ1. However, miniXIST transgene recruited all marks, albeit not to full XIST levels. The ability of the PID domain to facilitate silencing and heterochromatic marks recruitment was unexpected, and inhibition of PRC1 suggested that many of these are PRC1-independent. These results suggest that the addition of this small region allowed the partial recruitment of all the changes induced by a full XIST, demonstrating the feasibility of finding a minimal functional XIST.
Single-Molecule Structure and Topology of Kinetoplast DNA Networks
Authors: He Pinyao , Katan Allard J. , Tubiana Luca , Dekker Cees , Michieletto Davide
Published: September 2022

The Kinetoplast DNA (kDNA) is a two-dimensional Olympic-ring-like network of mutually linked 2.5 kb-long DNA minicircles found in certain parasites called Trypanosomes. Understanding the self-assembly and replication of this structure are not only major open questions in biology but can also inform the design of synthetic topological materials. Here we report the first high-resolution, single-molecule study of kDNA network topology using AFM and steered molecular dynamics simulations. We map out the DNA density within the network and the distribution of linking number and valence of the minicircles. We also characterise the DNA hubs that surround the network and show that they cause a buckling transition akin to that of a 2D elastic thermal sheet in the bulk. Intriguingly, we observe a broad distribution of density and valence of the minicircles, indicating heterogeneous network structure and individualism of different kDNA structures. Our findings explain outstanding questions in the field and offer single-molecule insights into the properties of a unique topological material.
Discovery of thyrocyte heterogeneity reveals an essential role of Notch signaling in thyroid function and mammalian homeostasis
Authors: Mosteiro Lluc , Thao Nguyen Thi Thu , Hankeova Simona , Reichelt Mike , Vandriel Shannon M. , Lai Zijuan , Choudhury Feroza K. , Sangaraju Dewakar , Kamath Binita M. , Scherl Alexis , Piskol Robert , Siebel Christian W.
Published: September 2022

The thyroid functions at the apex of a web of endocrine organs that control cell growth, differentiation and metabolic homeostasis. Thyroid dysregulation significantly impacts human health in myriad ways with thyroid diseases standing as the most common endocrine disorder. Despite the essential role of the thyroid in human health, a high-resolution view of the cellular composition as well as molecular mechanisms that govern function of this crucial organ have been lacking. Employing the first single-cell analyses of adult mouse thyroid, we here report the discovery of unexpected thyrocyte heterogeneity, specifically three distinct thyrocyte subtypes marked by different metabolic and Notch signaling patterns. Using a battery of pharmacologic and genetic methods, we find that selective inhibition of Notch ligands and receptors disrupts thyrocyte mitochondrial activity and ROS production, thus decreasing levels of circulating thyroid hormones, inducing hypothyroidism and disrupting whole-body thermoregulation. We find an enriched frequency of hypothyroidism in children with Alagille Syndrome, a genetic disorder marked by Notch loss-of-function mutations, suggesting that our Notch-thyroid mechanisms are relevant in humans and directly account for Alagille hypothyroidism. Overall, our work reveals that Notch, although classically described as a developmental pathway that determines cell fate, controls homeostasis and thermoregulation in the adult through a mitochondria-based mechanism in a subset of thyrocytes. Our fine-grained picture of the thyroid unveils a novel understanding of this key metabolic organ and provides clinically impactful insights into its pathological dysfunctions.
Utility of stromal lymphocytes in diagnosis and predicting upgrade of B3 breast lesions from core biopsies
Authors: Kader Tanjina , Hendry Shona , Provenzano Elena , Jayawardana Madawa W , Pang Jia-Min , Elder Kenneth , Byrne David J , Tjoeka Lauren , Frazer Helen ML , House Eloise , Jayasinghe Sureshni , Keane Holly , Murugasu Anand , Rajan Neeha , Miligy Islam M , Green Andrew R , Rakha Emad A , Fox Stephen B , Mann G. Bruce , Campbell Ian G , Gorringe Kylie L
Published: September 2022

For more than two decades attempts have been made to identify a subset of women diagnosed with lesions with uncertain malignant potential (B3 lesions) who could safely be observed rather than being treated with surgical excision and/or chemoprevention. Various histopathological, clinical and imaging parameters for risk recommendation have been evaluated, with little impact on clinical practice. The primary reason for surgery is to rule out an upgrade lesion to either ductal carcinomain situ(DCIS) or invasive breast cancer (IBC). While on average 30% of these patients are upgraded after diagnostic biopsy, a large number are over treated,making this an important harm of screening.

Here we evaluated stromal lymphocytes from B3 biopsies (n=264) as a predictive biomarker for upgrade. A higher number of stromal lymphocytes were observed in upgraded B3 lesions than non-upgraded (p< 0.01, zero inflated binomial model) for both ductal and papillary lesions (n=174). This observation was validated in an independent cohort (p<0.001, p<0.05, zero binomial model, ductal and papillary lesions, respectively) (n=90). Our data suggested that the presence of ≥5% of lymphocytes in the surrounding specialised stroma of B3 lesions are predictive of B3 lesions being upgraded with a specificity of 93% and 87% in our discovery and validation cohorts, respectively. The area under the curve (AUC) for the discovery cohort using lymphocyte count and age as variables was 0.77 and was validated with an AUC of 0.81 in the validation cohort.

In conclusion, we can identify a subset of the patients at risk of upgrade with high specificity. Assessing the tumour microenvironment including stromal lymphocytes may contribute to reducing unnecessary surgeries in the clinic.
Detection and Role of Feline Apolipoprotein B mRNA-editing Enzyme Catalytic Polypeptide Subunit 3G-Like Protein in Feline Cells and Tissues
Authors: Assogba Barnabe D. , Chaudhary Shannon , Kaur Harmanpreet , Soo Raymond , Dela Cruz Mark A. P. , Assogba Jovy M. G.
Published: September 2022

The innate host defence system is designed to resist pathogenic microorganism infections. Despite the compelling scientific evidence, our understanding of the full potential of the mechanism is still unclear due to the complex interactions between hosts and invaders. We previously reported latency in cat mucosal infected with low-dose cell-associated feline immunodeficiency virus (102 and 103 infected cells). Here we investigated the expression of Apolipoprotein B mRNA-editing enzyme catalytic subunit 3G (APOBEC3G or A3G) in feline cells and tissues and whether its presence antagonizes the viral pre-integration complex resulting in partial or complete FIV latency. Total RNA and protein lysates were collected from cell lines, blood, and tissue samples. Real-time RT-PCR and western blot assays were used to quantify fA3G-like protein in cats exposed to high versus low-dose cell-associated FIV. We consistently detected fA3G-like protein in mock T-cell lines (E-CD4+, MYA-1, Crandell feline kidney cells) and primary bone marrow-derived macrophages with variable expressions in feline peripheral blood mononuclear cells (PBMC). In addition, the fA3G-like protein was found to interact with FIV group-specific antigen (Gag) protein through immunoprecipitation assays. The protein expression was utterly abrogated following FIV infection. However, in lytic FIV infection (in vivo), fA3G-like protein decreased in early post-infection, whereas latently infected cats showed stable expression. These data are the first report of the fA3G-like protein expression in felines and its abrogation in lytic but not in latent FIV-infected individuals. These results might provide new insight into the role of fA3G-like protein in the host defence mechanism against retrovirus infections.
Microphysiological vascular malformation model reveals a role of dysregulated Rac1 and mTORC1/2 in lesion formation
Authors: Aw Wen Yih , Cho Crescentia , Wang Hao , Cooper Anne Hope , Doherty Elizabeth L. , Rocco David , Huang Stephanie A. , Kubik Sarah , Whitworth Chloe P. , Armstrong Ryan , Hickey Anthony J. , Griffith Boyce , Kutys Matthew L. , Blatt Julie , Polacheck William J.
Published: September 2022

Somatic activating mutations ofPIK3CAare associated with the development of vascular malformations (VMs). Here, we describe a microfluidic model ofPIK3CA-driven VMs consisting of human umbilical vein endothelial cells (HUVECs) expressingPIK3CAactivating mutations embedded in 3D hydrogels. We observed enlarged and irregular vessel phenotypes, consistent with clinical signatures and concomitant with PI3K-driven upregulation of Rac1/PAK, MEK/ERK, and mTORC1/2 signaling. We observed differential effects between Alpelisib, a PIK3CA inhibitor, and Rapamycin, an mTORC1 inhibitor, in mitigating matrix degradation and vascular network topology. While both drugs are effective in preventing vessel enlargement, Alpelisib suppressed mTORC2-dependent AKT1 phosphorylation and MEK/ERK signaling. Rapamycin failed to reduce MEK/ERK and mTORC2 activity and resulted in vascular hyperbranching, while inhibiting PAK, MEK1/2, and mTORC1/2 signaling mitigates abnormal growth and vascular dilation. Collectively, these findings establish anin vitroplatform for modeling VMs and confirm a role of dysregulated Rac1/PAK and mTORC1/2 signaling inPIK3CA-driven VMs.
Aquaporin regulates cell rounding through vacuole formation during endothelial-to-hematopoietic transition
Authors: Sato Yuki , Shigematsu Mugiho , Shibata-Kanno Maria , Maejima Sho , Tamura Chie , Sakamoto Hirotaka
Published: September 2022

Endothelial-to-hematopoietic transition (EHT) is crucial for hematopoietic stem cell (HSC) generation. During EHT, the morphology of hemogenic endothelial cells (HECs) changes from flat and adherent to spherical HSCs, which detach from the dorsal aorta. HECs attain a rounded shape in a mitosis-independent manner before cell adhesion termination, suggesting an atypical cell-rounding mechanism. However, the direct mechanisms underlying this change in cell morphology during EHT remain unclear. Here, we show that large vacuoles were transiently formed in HECs and that aquaporin-1 (AQP1) was localized in the vacuole and plasma membranes. Overexpression of AQP1 in non-HECs induced ectopic vacuole expansion, cell rounding, and subsequent cell detachment from the endothelium into the bloodstream, mimicking EHT. Loss of redundant AQP functions by CRISPR/Cas9 gene editing in HECs impeded the morphological EHT. Our findings provide the first evidence indicating that morphological segregation of HSCs from endothelial cells is regulated by water influx into vacuoles. These findings provide important insights for further exploration of the mechanisms underlying cell/tissue morphogenesis through water-adoptive cellular responses.

SUMMARY STATEMENT

Hemogenic endothelial cells transiently form large vacuoles during endothelial-to-hematopoietic transition. Aquaporin water channels regulate cell rounding and detachment of emerging hematopoietic stem cells through vacuole formation.
Development of the alpha rhythm is linked to visual white matter pathways and visual detection performance
Authors: Caffarra Sendy , Kanopka Klint , Kruper John , Richie-Halford Adam , Roy Ethan , Rokem Ariel , Yeatman Jason D.
Published: September 2022

Alpha is the strongest electrophysiological rhythm in awake humans at rest. Despite its predominance in the EEG signal, strong variations can be observed in alpha properties during development, with an increase of alpha frequency over childhood and adulthood. Here we tested the hypothesis that these changes of alpha rhythm are related to the maturation of visual white matter pathways. We capitalized on a large dMRI-EEG dataset (dMRI n=2,747, EEG n=2,561) of children and adolescents (age range: 5-21 years old) and showed that maturation of the optic radiation specifically accounts for developmental changes of alpha frequency. Behavioral analyses also confirmed that variations of alpha frequency are related to maturational changes in visual perception. The present findings demonstrate the close link between developmental variations in white matter tissue properties, electrophysiological responses, and behavior.
Genetic tools that target mechanoreceptors produce reliable labeling of bladder afferents
Authors: Tran Emily L. , Stuedemann Sara A. , Link Olivia D. , Crawford LaTasha K.
Published: September 2022

Mechanosensitive bladder sensory neurons are critical for sensing bladder distention, but their role in bladder pain and bladder pathology is poorly understood, due in part, to the challenges of identifying mechanoreceptors in tissue sections. A lot of what is known about how disease alters sensory innervation of the bladder comes from studies that traditionally focus on nociceptive nerve terminals. In seeking tools to characterize the role of non-nociceptive afferents in the bladder, we first examined neurofilament heavy (NFH), a marker for medium to large-diameter myelinated A fibers, in combination with the common marker for peptidergic nociceptors, calcitonin gene-related peptide (CGRP). While there was partial overlap between NFH and CGRP, 87% of NFH fibers were CGRP-negative, underscoring the abundance of non-nociceptive A fibers nerve terminals in the bladder. Two mouse lines that have been used for genetic labeling of mechanoreceptors of the skin were tested for their ability to label bladder afferents. Once crossed to Cre-dependent reporter lines, tyrosine kinase B (TrkB) TrkBCreER2 mice can be used to label A-delta mechanoreceptors while receptor tyrosine kinase ret protooncogene (Ret) RetCreER2 mice can label a combination of A-beta mechanoreceptors and non-peptidergic nociceptors in adult mice (late Ret). Both mouse lines produced successful labeling of bladder nerve terminals demonstrating partial overlap with NFH and minimal overlap with CGRP. Thus, we have identified new genetic strategies to investigate CGRP-negative subpopulations of bladder afferents that remain largely uncharacterized in studies that target peptidergic nociceptors. These tools can help elucidate the role of mechanosensitive afferents in bladder pathophysiology and urologic chronic pain.
NPAS4 supports drug-cue associations and relapse-like behavior through regulation of the cell type-specific activation balance in the nucleus accumbens
Authors: Hughes Brandon W. , Tsvetkov Evgeny , Siemsen Benjamin M. , Snyder Kirsten. K. , Akiki Rose Marie , Wood Daniel J. , Penrod Rachel D. , Scofield Michael D. , Berto Stefano , Taniguchi Makoto , Cowan Christopher W.
Published: September 2022

Use of addictive substances creates powerful drug-cue associations that often trigger relapse. Drug seeking is gated in the nucleus accumbens (NAc) by competing activation of D1 dopamine receptor-expressing medium spiny neurons (D1-MSNs) that promote, and D2 dopamine receptor-expressing neurons (D2-MSNs) that oppose, drug seeking. We show here that the ensemble of neurons in the NAc that induce the neuronal activity-regulated transcription factor, Neuronal PAS Domain Protein 4 (NPAS4), is required for cocaine-context associations. In addition, NPAS4 functions within NAc D2-MSNs to govern the activation balance of NAc D1-MSNs and D2-MSNs necessary for drug-context memories and cue-induced cocaine, but not sucrose, seeking. NPAS4 regulates drug-cue associations and preponderant D1-MSN activation by influencing a program of gene expression that blocks cocaine-induced potentiation of prefrontal cortical excitatory drive onto D2-MSNs. Together our findings reveal that NPAS4 is a key player governing NAc MSN cell-type activation balance and promoting drug-cue associations and relapse vulnerability.
How inhibitory and excitatory inputs gate output of the inferior olive
Authors: Loyola Sebastián , Hoogland Tycho M. , Negrello Mario , Hoedemaker Hugo , Romano Vincenzo , De Zeeuw Chris I.
Published: September 2022

The inferior olive provides the climbing fibers to Purkinje cells in the cerebellar cortex, where they elicit all-or-none complex spikes and control major forms of plasticity. Given their important role in both short-term and long-term coordination of cerebellum-dependent behaviors, it is paramount to understand the factors that determine the output of olivary neurons. Here, we investigate how the inhibitory and excitatory inputs to the olivary neurons interact with each other, generating spiking patterns of olivary neurons that ride on their intrinsic oscillations. Using dual color optogenetic stimulation and whole cell recordings we demonstrate how intervals between the inhibitory input from the cerebellar nuclei and excitatory input from the mesodiencephalic junction affect phase and gain of the olivary output at both the sub- and suprathreshold level. When the excitatory input is activated approximately 50 ms after the inhibitory input, the phase of the intrinsic oscillations becomes remarkably unstable and the excitatory input can hardly generate any olivary spike. Instead, when the excitatory input is activated approximately 150 ms after the inhibitory input, the excitatory input can optimally drive olivary spiking, riding on top of the first cycle of the subthreshold oscillations that have been powerfully reset by the preceding inhibitory input. Simulations of a large-scale network model of the inferior olive highlight to what extent the synaptic interactions penetrate in the neuropil, generating quasi-oscillatory spiking patterns in large parts of the olivary subnuclei, the size of which also depends on the relative timing of the inhibitory and excitatory inputs.
Pseudokinase HPO-11 inhibits nonsense-mediated decay to ensure genome stability in C. elegans
Authors: Zhao Qian , Gromoff Erika D , Yang Wei , Schwarz Jennifer , Tittel Lena , Schulze Ekkehard , Warscheid Bettina , Baumeister Ralf , Qi Wenjing
Published: September 2022

DNA double-strand breaks (DSBs) are highly toxic DNA lesions that can induce mutations and chromosome rearrangement therefore causing genome instability (GIN). In response to DSBs, cells activate the DNA damage response by hierarchical assembly of signaling and repair mechanisms. This involves recruitment of the repair factors at DSB sites, local chromatin remodeling, cell cycle arrest and, eventually, DNA repair or apoptosis. Studies investigating mechanisms ensuring genome stability have so far mostly focused on DNA-protein interactions and signal transduction in response to DNA damage. Emerging evidence in the last decade suggests that post-transcriptional control of gene expression by RNA-binding proteins also participates in maintaining genome integrity. However, how specific control of RNA fate mechanistically affects genome stability is still poorly understood. Here, we report that the pseudokinase HPO-11 ensures genome integrity in C. elegans. Loss of hpo-11 leads to accumulation of R-loops, increased DSBs and germline apoptosis, as well as an elevated mutation rate in the somatic cells. In addition, inhibition of nonsense mediated decay (NMD) reduces DSBs and germline apoptosis in the absence of hpo-11. We find that HPO-11 physically interacts with SMG-2, the core factor of NMD, and prevents degradation of specific transcripts by NMD, thus contributing to maintenance of genome stability. Furthermore, knock-down of hpo-11 human homologs NRBP1/2 also results in increased DNA DSBs, and NRBP1/2 physically interact with the human SMG-2 orthologue UPF1. In summary, our work identifies an evolutionarily conserved role of HPO-11 to protect genome stability via suppressing abnormal mRNA decay by NMD.
Early detection of late blight in potato by whole-plant redox imaging
Authors: Hipsch Matanel , Michael Yaron , Lampl Nardy , Sapir Omer , Cohen Yigal , David Helman , Rosenwasser Shilo
Published: September 2022

Late blight caused by the oomycete Phytophthora infestans is a most devastating disease of potatoes (Solanum tuberosum). Its early detection is crucial for suppressing disease spread. Necrotic lesions are normally seen in leaves at 4 dpi (days post inoculation) when colonized cells are dead, but early detection of the initial biotrophic growth stage, when the pathogen feeds on living cells, is challenging. Here, the biotrophic growth phase of P. infestans was detected by whole-plant redox imaging of potato plants expressing chloroplast-targeted reduction-oxidation sensitive green fluorescent protein (chl-roGFP2). Clear spots on potato leaves with a lower chl-roGFP2 oxidation state were detected as early as 2 dpi, before any visual symptoms were recorded. These spots were particularly evident during light-to-dark transitions and reflected mislocalization of chl-roGFP2 outside the chloroplasts, demonstrating perturbation of the chloroplast import system by the pathogen. Image analysis based on machine learning enabled systematic identification and quantification of spots and unbiased classification of infected and uninfected leaves in inoculated plants. Comparing redox to chlorophyll fluorescence imaging showed that infected leaf areas which exhibit mislocalized chl-roGFP2 also showed reduced non-photochemical quenching (NPQ) and enhanced quantum PSII yield (ΦPSII) compared to the surrounding leaf areas. The data suggest that mislocalization of chloroplast-targeted proteins is an efficient marker of late blight infection and demonstrate how it can be utilized for nondestructive monitoring of the disease biotrophic stage using whole-plant redox imaging.
PKCβ facilitates leukemogenesis in chronic lymphocytic leukaemia by promoting constitutive BCR-mediated signaling
Authors: Hay Jodie , Tarafdar Anuradha , Holroyd Ailsa , Moka Hothri A. , Dunn Karen M. , Alshayeb Alzahra , Lloyd Bryony H. , Cassels Jennifer , Malik Natasha , Khan Ashfia F. , Sou IengFong , Lees Jamie , Almuhanna Hassan N. B. , Kalakonda Nagesh , Slupsky Joseph R. , Michie Alison M.
Published: September 2022

B cell antigen receptor (BCR) signaling competence is critical for pathogenesis of chronic lymphocytic leukemia (CLL). Defining key proteins that facilitate these networks aid in the identification of targets for therapeutic exploitation. We previously demonstrated that reduced PKCα function in mouse hematopoietic stem/progenitor cells (HPSCs) resulted in PKCβII upregulation and generation of a poor-prognostic CLL-like disease. Here, prkcb knockdown in HSPCs leads to reduced survival of PKCα-KR-expressing CLL-like cells, concurrent with reduced expression of the leukemic markers CD5 and CD23. SP1 promotes elevated expression of prkcb in PKCα-KR expressing cells enabling leukemogenesis. Global gene analysis revealed an upregulation of genes associated with B cell activation in PKCα-KR expressing cells, coincident with upregulation of PKCβII: supported by activation of key signaling hubs proximal to the BCR and elevated proliferation. Ibrutinib (BTK inhibitor) or enzastaurin (PKCβII inhibitor) treatment of PKCα-KR expressing cells and primary CLL cells showed similar patterns of Akt/mTOR pathway inhibition, supporting the role for PKCβII in maintaining proliferative signals in our CLL mouse model. Ibrutinib or enzastaurin treatment also reduced PKCα-KR-CLL cell migration towards CXCL12. Overall, we demonstrate that PKCβ expression facilitates leukemogenesis and identify that BCR-mediated signaling is a key driver of CLL development in the PKCα-KR model.

Statement of Significance

PKCβ facilitates leukemogenesis of CLL, driven through an SP1-regulated transcriptional program and promotes BCR signaling. Thus far, PKCβ is the only kinase within the BCR signaling pathway, a key pathway in driving CLL pathogenesis, implicated in the generation of neoplastic B lineage cells.
Distinct patterns of PV and SST GABAergic neuronal activity in the basal forebrain during olfactory-guided behavior in mice
Authors: Moss Elizabeth H. , Tantry Evelyne K. , Le Elaine , Chin Pey-Shyuan , Ambrosi Priscilla , Brandel-Ankrapp Katie L. , Arenkiel Benjamin R.
Published: September 2022

Sensory perception relies on the flexible detection and interpretation of stimuli across variable contexts, conditions, and behavioral states. The basal forebrain is a hub for behavioral state regulation, supplying dense cholinergic and GABAergic projections to various brain regions involved in sensory processing. Of GABAergic neurons in the basal forebrain, parvalbumin (PV) and somatostatin (SST) subtypes serve opposing roles towards regulating behavioral states. To elucidate the role of basal forebrain circuits in sensory-guided behavior, we investigated GABAergic signaling dynamics during odor-guided decision-making. We used fiber photometry to record cell type-specific basal forebrain activity during an odor discrimination task and correlated temporal patterns of PV and SST neuronal activity with olfactory task performance. We found that while both PV-expressing and SST-expressing GABAergic neurons were excited during trial initiation, PV neurons were selectively suppressed by reward whereas SST neurons were excited. Notably, chemogenetic inhibition of BF SST neurons modestly altered decision bias to favor reward-seeking while optogenetic inhibition of BF PV neurons during odor presentations improved discrimination accuracy. Together, these results suggest that the bidirectional activity of GABAergic basal forebrain neuron subtypes distinctly influence perception and decision-making during olfactory guided behavior.

SIGNIFICANCE STATEMENT

This study reveals distinct roles for basal forebrain GABAergic neurons in odor perception and odor-guided decision-making. Fiber photometry shows that basal forebrain parvalbumin-expressing neurons are selectively suppressed by rewards, while somatostatin-expressing neurons are activated, establishing the unique recruitment of these GABAergic neurons during behavioral reinforcement. Chemogenetic and optogenetic interventions demonstrate divergent roles for these neuronal subtypes in reward-seeking behavior and odor perception. This research provides new insights into how GABAergic neurons in the basal forebrain shape sensory perception and decision-making.
Nearest neighbor search on embeddings rapidly identifies distant protein relations
Authors: Schütze Konstantin , Heinzinger Michael , Steinegger Martin , Rost Burkhard
Published: September 2022

Since 1992, all state-of-the-art (SOTA) methods for fast and sensitive identification of evolutionary, structural, and functional relations between proteins (also referred to as “homology detection”) use sequences and sequence-profiles (PSSMs). Protein Language Models (pLMs) generalize sequences, possibly capturing the same constraints as PSSMs, e.g., through embeddings. Here, we explored how to use such embeddings for nearest neighbor searches to identify relations between protein pairs with diverged sequences (remote homology detection for levels of <20% pairwise sequence identity, PIDE). While this approach excelled for proteins with single domains, we demonstrated the current challenges applying this to multi-domain proteins and presented some ideas how to overcome existing limitations, in principle. We observed that sufficiently challenging data set separations were crucial to provide deeply relevant insights into the behavior of nearest neighbor search when applied to the protein embedding space, and made all our methods readily available for others.
Linking mesozooplankton and SAR11 bacteria in Oxygen Deficient Zones and the open ocean
Authors: Fuchsman Clara A. , Hays Matthew D. , Huanca-Valenzuela Paulina , Gregory Benjamin P. , Plough Louis V. , Duffy Megan E. , Keil Richard G. , Peng Xuefeng
Published: September 2022

The gravitational biological pump is not large enough to account for microbial heterotrophic activity in the mesopelagic ocean. Migrating zooplankton may be a key source of organic matter transport to depth. Here we show signatures of zooplankton in the suspended organic matter at the zooplankton vertical migration depth in the Eastern Tropical North Pacific Oxygen Deficient Zone (ETNP ODZ). We examine the mesozooplankton community in metagenomic depth profiles using the mitochondrial cytochrome c oxidase (COI) gene as a marker in the ETNP and Eastern Tropical South Pacific (ETSP) ODZs and at the oxic Hawaii Ocean Timeseries (HOT). Additionally, eukaryotic transcripts (polyA-selected) were examined for zooplankton in the ETNP. While zooplankton eDNA increased in the ODZ, zooplankton eRNA decreased in the ODZ, similar to previous net-based data, implying that eDNA is better preserved under anoxia. At all stations, Cnidaria, often missed in net-based data, contributed greatly to the zooplankton eDNA/eRNA. SAR11 abundance, determined from the single-copy core gene (rpoB), significantly correlated with zooplankton eDNA, with R2values >0.8 at all stations. Strong correlations between SAR11 and zooplankton have not been previously reported, but are logical as SAR11 bacteria consume and zooplankton excrete simple dissolved organic compounds. SAR11 bacteria possessed genes to utilize urea and taurine in the ODZ, both compounds known to be excreted by zooplankton. In ODZs, SAR11 bacteria preferentially used the taurine degradation pathway leading to C and N assimilation, not the pathway for organic S assimilation, probably due to additional sources of organic S in ODZs.
RTEL-1 and DNA polymerase theta promote subtelomeric DNA synthesis and telomere fusion in C. elegans
Authors: Lister-Shimauchi Evan H. , Brady Morgan , Frenk Stephen , Harris Braxton , Ortiz Ana-Maria Leon , Memili Aylin , Nguyen Minh , Boulton Simon , Ahmed Shawn
Published: September 2022

Interstitial telomere sequences (ITS) are degenerate telomere tracts scattered along chromosome arms whose functions are not well understood. We found that critically shortened telomeres of C. elegans telomerase mutants initiate DNA synthesis within ITS tracts that were close to or far from a telomere. Some ITS tracts were targeted recurrently. RTEL-1 dismantles T-loops and recombination intermediates, and DNA polymerase theta (POLQ-1) promotes end-joining using short segments of microhomology. In telomerase mutants, RTEL-1 and POLQ-1 promoted telomere fusion and DNA synthesis at subtelomeric ITS tracts. RTEL-1 is known to suppress homologous recombination, and we found that RTEL-1 similarly suppressed POLQ-1-mediated double-strand break repair. Mutation signatures characteristic of repair by POLQ-1 occurred during initiation of subtelomeric DNA synthesis and at subsequent template shifting events. We propose that RTEL-1 and POLQ-1 play distinct essential roles in subtelomeric DNA synthesis, a process that may contribute significantly to telomere fusion and tumor genome evolution.
Simple, low-cost production of DNA MS2 virus-like particles as molecular diagnostic controls
Authors: Crone Michael A. , Freemont Paul S.
Published: September 2022

Suitable controls are integral for the validation and continued quality assurance of diagnostic workflows. Plasmids, DNA or in vitro transcribed RNA are often used to validate novel diagnostic workflows, however, they are poorly representative of clinical samples. RNA phage virus-like particles packaged with exogenous RNA have been used in clinical diagnostics as workflow controls, serving as surrogates for infectious viral particles. Comparable controls for DNA viruses are more challenging to produce, with analogous DNA phages being infectious and packaging of DNA within RNA phages requiring complex purification procedures and expensive chemical linkers. We present a simple and inexpensive method to produce MS2 virus-like particles, packaged with DNA, that makes use of affinity chromatography for purification and enzymatic production of exogenous DNA suitable for packaging. The produced virus-like particles were packaged with Hepatitis B Virus DNA and were then quantified using droplet digital PCR and calibrated against the WHO international standard using a commercial assay in an accredited clinical laboratory.
Effects of stimulus rate and periodicity on auditory cortical entrainment to continuous sounds
Authors: Momtaz Sara , Bidelman Gavin M.
Published: September 2022

The neural mechanisms underlying the exogenous coding and neural entrainment to rapid auditory stimuli have seen a recent surge of interest. However, few studies have characterized how parametric changes in stimulus presentation alter entrained responses. Applying inter-trial phase-locking (ITPL) and phase-locking value (PLV) analyses applied to high-density human electroencephalogram (EEG) data, we investigated the degree to which the brain entrains to speech vs. non-speech (i.e., click) sounds within and across tokens. Passive cortico-acoustic tracking was investigated in N=24 normal young adults utilizing EEG time-frequency and source analyses that isolated neural activity stemming from both auditory temporal cortices. We parametrically manipulated the rate and periodicity of repetitive, continuous speech and click stimuli to investigate how speed and jitter in ongoing sounds stream affect oscillatory entrainment. Both stimulus domains showed rightward hemisphere asymmetry in phase-locking strength with stronger and earlier responses to speech vs. clicks. Neuronal synchronization to speech was enhanced at 4.5 Hz (the putative universal rate of speech) and showed a differential pattern to that of clicks, particularly at higher rates. Phase-locking to speech decreased with increasing jitter but entrainment to speech remained superior to clicks. Surprisingly, click were invariant to periodicity manipulations. Our findings provide evidence that the brain’s neural entrainment to complex sounds is enhanced and more sensitized when processing speech relative to non-speech sounds. That this specialization is apparent even under passive listening suggests a priority of the auditory system for synchronizing to behaviorally-relevant signals.
LARP1 regulates metabolism and mTORC1 activity in cancer
Authors: Chettle James , Dedeic Zinaida , Fischer Roman , Vendrell Iolanda , Campo Leticia , Easton Alistair , Browne Molly , Morris Josephine , Schwenzer Hagen , Lascaux Pauline , Gijsbers Rik , Pires Elisabete , Royston Daniel J. , Ferguson David J. P. , Coosemans An , Kessler Benedikt , McCullagh James , Ahmed Ahmed A. , Ramadan Kristijan , Bushell Martin , Harris Adrian L. , Goding Colin R. , Blagden Sarah P.
Published: September 2022

The protein mammalian target of rapamycin (mTOR) is a master regulator of cell homeostasis. Although mTOR is aberrantly overactivated in 70% ovarian cancers, mTOR cascade inhibitors (such as those blocking the kinase activity of mTOR itself or upstream kinases PI3K/AKT) have demonstrated disappointing activity in ovarian cancer clinical trials. These findings indicate that, despite its pivotal role in normal cells, hyperactivated mTOR does not act as a master regulator of metabolism in this cancer context. Surprisingly, we have identified that the RNA binding protein LARP1, a known phospho-target of mTORC1 and activator of ribosomal biogenesis, is responsible for metabolic reprogramming in mTOR-dysregulated cancers. LARP1 post-transcriptionally regulates the expression of several hundred rate-limiting enzymes involved in multiple aspects of metabolism, including glycolysis and oxidative phosphorylation. Through this mechanism LARP1 sustains ATP production and mTORC1 localisation on the lysosome, thereby activating cell proliferation despite the scarcity of extracellular nutrients. Our findings show that, by sustaining global cellular metabolism in response to growth factor signalling, LARP1 has a central post-transcriptional role in controlling mTORC1 localisation and driving cancer progression, a key cancer hallmark.
Occupational stress for employee turnover intention: mediation effect of service climate and emotion regulation
Authors: Gautam Dhruba Kumar , Gautam Prakash Kumar
Published: September 2022

Purpose

This study examines the effect of occupational stress on turnover intention of employees working in the banking industry. The authors examine the mediating effects of service climate and emotional regulations of the employees in the relationship between occupational stress and intention to leave the organization.

Design/methodology/approach

This study followed stratified sampling technique for data collection from employees of ten commercial banks based on the banks' financial performance of top 5 and bottom 5 out of 27 banks. Data were collected at 2 stages, first from 465 employees for occupational stressors and second from 408 employees among the participants in the first stage for turnover intention, service climate and emotional regulation. Harman’s one-factor test was conducted to examine the common method bias. Confirmatory factor analysis (CFA), regression analysis and Preacher and Hayes Process Macro approach were used to examine mediation effect.

Findings

Three factors, namely workload (WL), role ambiguity and growth opportunity expectations were identified as the occupational stressors in the banking industry, predicting a positive relation of overall occupational stress to the intention to turnover. Service climate and the employees’ emotion regulation ability mediate the relationship between stress and turnover intention. Results also revealed no significant role of control variables in predicting occupational stress and turnover intention.

Practical implications

This study implies that the WL, role ambiguity and growth opportunity expectations of the employees cause stress in employees which may lead to have turnover intention. In order to get success in competitive environment, managers of banking industry can address stressors by enhancing service climate and formulating policies and programs to strengthen the emotion regulation which is evidence to strengthen the reciprocity approach of social exchange theory in employees’ commitment.

Originality/value

This study contributes to the social exchange theory and attempts to fulfill the gaps in empirical research on personnel psychology, human capital\ and organization management in developing countries.
Prediction of the failure strain of FRP sheets on confined concrete columns with circular cross-section
Authors: Silva Lobo Paulo , Jesus Mariana , Freitas Paulo S.A.
Published: September 2022

Purpose

In this paper a model for the prediction of the failure strain of fiber reinforced polymer (FRP) sheets is proposed, based on a significant database consisting of results reported in the literature for concrete columns with circular cross-section confined with carbon, glass and aramid fiber reinforced polymers. This study aims to analyze the aforementioned model.

Design/methodology/approach

Significant experimental research on the confinement of columns externally wrapped with FRP has been carried out in the last decades. Several models of confined concrete have been proposed using different approaches for the prediction of the failure strain of FRP sheets. The majority of those proposals were developed to be simple, thus significant differences with experimental results can be found.

Findings

The results of a total of 572 specimens were considered. Statistical analysis was used to derive a regression model, which relates the outcome to a set of predictors of interest using linear assumptions.

Originality/value

The proposed model is shown to be more accurate than other proposals found in the literature.
IsGuanxicritical to the job burnout of ITP and IS/IT users in Chinese society?
Authors: Chang Christina Ling-hsing , Wu Sheng
Published: September 2022

Purpose

The speedy development of information technology (IT) is indeed a significant issue to support the enterprises to spread their products and services to their customers. Although the adoption of IT in the enterprises can help employees' work, it will increase their work exhaustion and job burnout too.

Design/methodology/approach

The survey data of this study was collected from Chinese IT personnel (ITP): 543 and Chinese information systems (IS)/IT users: 525, totaling 1,068 respondents.

Findings

The result shows that the influences of the organizational factors and guanxi on the work exhaustion are different between Chinese ITP and IS/IT users in the research model. Also the implications of the study are discussed and made some contribution to the research and management practice.

Originality/value

By comparison between the Chinese ITP and IS/IT users, this study shows that guanxi can be antecedent factor to some organizational factors, and the organizational factors guanxi have had a different effect on the work exhaustion of Chinese ITP and IS/IT users. Given that the critical successes to the organizations, management should focus on what direction to reduce Chinese ITP and IS/IT users work exhaustion.
Leading for equity: how principals experience professional learning
Authors: Clayton Jennifer , Nganga Christine
Published: September 2022

Purpose

This study examined how principals experienced professional learning that asked them to reflect on their own biases and beliefs, as well as to determine where there are systemic issues of inequity in their schools, and what they planned to do to address those challenges.

Design/methodology/approach

Through interviews, observations, and documents, the authors explored the building and sustaining of equity mindsets in education.

Findings

The authors analysis led to four overarching themes that collectively answered the research questions about how the principals learned individually and in concert with each other. The themes included the power of the personal journey and commitment, the importance of building a network of support, the benefit of practice-oriented activities that disrupt power structures and inequitable practices in schools, and sustainability.

Research limitations/implications

The study focused on a single institute and as such, findings are not generalizable, but may be transferable.

Practical implications

Schools and school districts may use the findings when designing professional learning, especially as it relates to leading with and for equity and in cross-district collaborations.

Originality/value

This contributes to the literature around concrete ways principals learn and reflect on enacting equity in schools.