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Host responses – autophagy, cell death, and inflammation – limit the growth of bacterial pathogens while minimizing tissue damage. During the early stages of infection, Mycobacterium tuberculosis ( Mtb ) thwarts these and other innate immune defense mechanisms in alveolar macrophages (AMs) derived from the yolk sac; in later stages, it circumvents defenses in recruited mononuclear cells (MNCs) and survives within them despite additional cytokine stimulation from recruited T cells. The mechanisms that drive variable rates of Mtb growth in different macrophage subtypes and how Mtb manipulates inflammatory responses to grow within innate immune cells remain obscure. Here we explored the role of the host factor, Tax-1 binding protein 1 (Tax1bp1), an autophagy receptor that targets pathogens for degradation through selective autophagy and terminates pro-inflammatory cytokine responses. Unexpectedly, we found that Tax1bp1-deficient mice were less susceptible to Mtb infection, and generated reduced inflammatory cytokine responses, compared to wild-type mice; the same mutant mice exhibited decreased growth of, and inflammatory cytokine responses to, Listeria monocytogenes , suggesting that Tax1bp1 plays a role in host responses to multiple intracellular pathogens. Contrary to our previous ex vivo findings in bone marrow-derived macrophages (BMDMs), in vivo growth of Mtb in AMs and a subset of recruited MNCs was more limited in mice lacking Tax1bp1 relative to wild-type mice. To better understand these differences, we performed global protein abundance measurements in mock- and Mtb- infected AM samples ex vivo from wild-type mice. These experiments revealed that Tax1bp1 protein abundance does not significantly change early after infection in AMs but does in BMDMs; moreover, early after infection, Tax1bp1-deficiency reduced necrotic-like cell death -- an outcome that favors Mtb replication -- in AMs but not BMDMs. Together, these results show that deficiency of Tax1bp1 plays a crucial, cell type-specific role in linking the regulation of autophagy, cell death, and anti-inflammatory host responses and overall reducing bacterial growth.

The COVID-19 pandemic has been driven by SARS-CoV-2 variants with enhanced transmission and immune escape. Apart from extensive evolution in the Spike protein, non-Spike mutations are accumulating across the entire viral genome and their functional impact is not well understood. To address the contribution of these mutations, we reconstructed genomes of recent Omicron variants with disabled Spike expression (replicons) to systematically compare their RNA replication capabilities independently from Spike. We also used a single reference replicon and complemented it with various Omicron variant Spike proteins to quantify viral entry capabilities in single-round infection assays. Viral entry and RNA replication were negatively correlated, suggesting that as variants evolve reduced entry functions under growing immune pressure on Spike, RNA replication increases as a compensatory mechanism. We identified multiple mutations across the viral genome that enhanced viral RNA replication. NSP6 emerged as a hotspot with a distinct L260F mutation independently arising in the BQ.1.1 and XBB.1.16 variants. Using mutant and revertant NSP6 viral clones, the L260F mutation was validated to enhance viral replication in cells and increase pathogenesis in mice. Notably, this mutation reduced host lipid droplet content by NSP6. Collectively, a systematic analysis of RNA replication of recent Omicron variants defined NSP6’s key role in viral RNA replication that provides insight into evolutionary trajectories of recent variants with possible therapeutic implications.

Mechanisms specifying cancer cell states and response to therapy are incompletely understood. Here we show epigenetic reprogramming shapes the cellular landscape of schwannomas, the most common tumors of the peripheral nervous system. We find schwannomas are comprised of 2 molecular groups that are distinguished by activation of neural crest or nerve injury pathways that specify tumor cell states and the architecture of the tumor immune microenvironment. Moreover, we find radiotherapy is sufficient for interconversion of neural crest schwannomas to immune-enriched schwannomas through epigenetic and metabolic reprogramming. To define mechanisms underlying schwannoma groups, we develop a technique for simultaneous interrogation of chromatin accessibility and gene expression coupled with genetic and therapeutic perturbations in single-nuclei. Our results elucidate a framework for understanding epigenetic drivers of tumor evolution and establish a paradigm of epigenetic and metabolic reprograming of cancer cells that shapes the immune microenvironment in response to radiotherapy. Schwannomas are regularly treated with radiotherapy, but the molecular effects on these tumours and their microenvironment remain unclear. Here, the authors show that radiotherapy can induce epigenetic reprogramming and immune infiltration in schwannomas, and develop the snARC-seq approach to analyse the epigenomic evolution at the single-cell level.

Meningiomas are associated with inactivation of NF2 /Merlin, but approximately one-third of meningiomas with favorable clinical outcomes retain Merlin expression. Biochemical mechanisms underlying Merlin-intact meningioma growth are incompletely understood, and non-invasive biomarkers that may be used to guide treatment de-escalation or imaging surveillance are lacking. Here, we use single-cell RNA sequencing, proximity-labeling proteomic mass spectrometry, mechanistic and functional approaches, and magnetic resonance imaging (MRI) across meningioma xenografts and patients to define biochemical mechanisms and an imaging biomarker that underlie Merlin-intact meningiomas. We find Merlin serine 13 (S13) dephosphorylation drives meningioma Wnt signaling and tumor growth by attenuating inhibitory interactions with β-catenin and activating the Wnt pathway. MRI analyses show Merlin-intact meningiomas with S13 phosphorylation and favorable clinical outcomes are associated with high apparent diffusion coefficient (ADC). These results define mechanisms underlying a potential imaging biomarker that could be used to guide treatment de-escalation or imaging surveillance for patients with Merlin-intact meningiomas. The molecular mechanisms underlying merlin-intact meningioma growth remain to be explored. Here, the authors show that merlin activates Wnt signalling and tumour growth through its dephosphorylation on serine 13 attenuating the inhibitory interactions with β-catenin.

The S. cerevisiae ISR1 gene encodes a putative kinase with no ascribed function. Here, we show that Isr1 acts as a negative regulator of the highly-conserved hexosamine biosynthesis pathway (HBP), which converts glucose into uridine diphosphate N-acetylglucosamine (UDP-GlcNAc), the carbohydrate precursor to protein glycosylation, GPI-anchor formation, and chitin biosynthesis. Overexpression of ISR1 is lethal and, at lower levels, causes sensitivity to tunicamycin and resistance to calcofluor white, implying impaired protein glycosylation and reduced chitin deposition. Gfa1 is the first enzyme in the HBP and is conserved from bacteria and yeast to humans. The lethality caused by ISR1 overexpression is rescued by co-overexpression of GFA1 or exogenous glucosamine, which bypasses GFA1's essential function. Gfa1 is phosphorylated in an Isr1-dependent fashion and mutation of Isr1-dependent sites ameliorates the lethality associated with ISR1 overexpression. Isr1 contains a phosphodegron that is phosphorylated by Pho85 and subsequently ubiquitinated by the SCF-Cdc4 complex, largely confining Isr1 protein levels to the time of bud emergence. Mutation of this phosphodegron stabilizes Isr1 and recapitulates the overexpression phenotypes. As Pho85 is a cell cycle and nutrient responsive kinase, this tight regulation of Isr1 may serve to dynamically regulate flux through the HBP and modulate how the cell's energy resources are converted into structural carbohydrates in response to changing cellular needs.

HIV, the virus that causes AIDS, heavily relies on the machinery of human cells to infect and replicate. Our study focuses on the host cell’s ubiquitination system which is crucial for numerous cellular processes. Many pathogens, including HIV, exploit this system to enhance their own replication and survival. E3 proteins are part of the ubiquitination pathway that are useful drug targets for host-directed therapies. We interrogated the 116 E3s found in human immune cells known as CD4+ T cells, since these are the target cells infected by HIV. Using CRISPR, a gene-editing tool, we individually removed each of these enzymes and observed the impact on HIV infection in human CD4+ T cells isolated from healthy donors. We discovered that 10 of the E3 enzymes had a significant effect on HIV infection. Two of them, TRAF2 and UHRF1, modulated HIV activity within the cells and triggered an increased release of HIV from previously dormant or “latent” cells in a new primary T cell assay. This finding could guide strategies to perturb hidden HIV reservoirs, a major hurdle to curing HIV. Our study offers insights into HIV-host interactions, identifies new factors that influence HIV infection in immune cells, and introduces a novel methodology for studying HIV infection and latency in human immune cells.

Background The neuropeptide arginine vasopressin (Avp) modulates social behaviors via its two centrally expressed receptors, the Avp 1a receptor and the Avp 1b receptor (Avpr1b). Recent work suggests that, at least in mice, Avp signaling through Avpr1b within the CA2 region of the hippocampus is critical for normal aggressive behaviors and social recognition memory. However, this brain area is just one part of a larger neural circuit that is likely to be impacted in Avpr1b knockout (−/−) mice. To identify other brain areas that are affected by altered Avpr1b signaling, genotypic differences in immediate early gene activation, i.e. c-FOS and early growth response factor 1 (EGR-1), were quantified using immunocytochemistry following a single exposure to an intruder. Results In females, no genotypic differences in intruder-evoked c-FOS or EGR-1 immunoreactivity were observed in any of the brain areas measured. In males, while there were no intruder-evoked genotypic differences in c-FOS immunoreactivity, genotypic differences were observed in EGR-1 immunoreactivity within the ventral bed nucleus of the stria terminalis and the anterior hypothalamus; with Avpr1b −/− males having less EGR-1 immunoreactivity in these regions than controls. Conclusions These data are the first to identify specific brain areas that may be a part of a neural circuit that includes Avpr1b-expressing cells in the CA2 region of the hippocampus. It is thought that this circuit, when working properly, plays a role in how an animal evaluates its social context.

Physical literacy (PL) is gaining more attention from educational policy-makers, practitioners, and researchers as a way to improve health and wellness outcomes for children and youth. While the development of PL is important for early years children, there is limited attention in the literature that explores the political, cultural, and social discourses imbued in colonialism that implicate how PL is actualized in Indigenous early childhood education (ECE) contexts. This case assemblage explores how the culturally rooted, interdisciplinary, and community-based PL initiative, Nature’s Way–Our Way (NWOW), negotiated movement with three early childhood educators in the pilot project with an early childhood education centre (ECEC) in Saskatchewan, Canada. Through postqualitative approaches to research, this case assemblage adopts new materialist methodologies to show how the natural order of knowing in movement was disrupted through moments of rupture generating stories of PL to encompass radical relationality with land. As land becomes a vital and lively part of PL storying, it can function as an important protective factor for Indigenous preschool-aged children’s wholistic wellness.

As multicellular organisms grow, spatial and temporal patterns of gene expression are strictly regulated to ensure that developmental programs are invoked at appropriate stages. In this work, we describe a putative transcriptional regulator in Arabidopsis, TACO LEAF (TCO), whose overexpression results in the ectopic activation of reproductive genes during vegetative growth. Isolated as an activation-tagged allele, tco-1D displays gene misexpression and phenotypic abnormalities, such as curled leaves and early flowering, characteristic of chromatin regulatory mutants. A role for TCO in this mode of transcriptional regulation is further supported by the subnuclear accumulation patterns of TCO protein and genetic interactions between tco-1D and chromatin modifier mutants. The endogenous expression pattern of TCO and gene misregulation in tco loss-of-function mutants indicate that this factor is involved in seed development. We also demonstrate that specific serine residues of TCO protein are targeted by the ubiquitous kinase CK2. Collectively, these results identify TCO as a novel regulator of gene expression whose activity is likely influenced by phosphorylation, as is the case with many chromatin regulators.

Background Fibroblast growth factors (FGFs) are crucial signaling molecules that direct the development of the vertebrate brain. FGF8 gene signaling in particular, may be important for the development of the hypothalamus–pituitary–adrenal (HPA)-axis. Indeed, newborn Fgf8 hypomorphic mice harbor a major reduction in the number of vasopressin (VP) neurons in the paraventricular nucleus (PVN), the central output component of the HPA-axis. Additionally, recent studies indicated that adult heterozygous ( +/neo ) Fgf8 hypomorphic mice exhibit more anxiety-like behaviors than wildtype (WT) mice. These studies led us to investigate whether Fgf8 hypomorphy abrogated VP and/or corticotropin-releasing hormone (CRH) neuronal development in the postnatal day (PN) 21 and adult mouse PVN. Furthermore, we studied whether Fgf8 hypomorphy disrupted HPA responsiveness in these mice. Methods Using immunohistochemistry, we examined the development of VP and CRH neurons located in the PVN of PN 21 and adult Fgf8 +/neo mice. Moreover, we used a restraint stress (RS) paradigm and measured corticosterone levels with enzyme immunoassays in order to assess HPA axis activation. Results The number of VP neurons in the PVN did not differ between WT and Fgf8 +/neo mice on PN 21 and in adulthood. In contrast, CRH immunoreactivity was much higher in Fgf8 +/neo mice than in WT mice on PN 21, this difference was no longer shown in adult mice. RS caused a higher increase in corticosterone levels in adult Fgf8 +/neo mice than in WT mice after 15 min, but no difference was seen after 45 min. Conclusions First, Fgf8 hypomorphy did not eliminate VP and CRH neurons in the mouse PVN, but rather disrupted the postnatal timing of neuropeptide expression onset in PVN neurons. Second, Fgf8 hypomorphy may, in part, be an explanation for affective disorders involving hyperactivity of the HPA axis, such as anxiety.