Correction of age-associated defects in dendritic cells enables CD4+ T cells to eradicate tumors.

Defective host defenses later in life are associated with changes in immune cell activities, suggesting that age-specific considerations are needed in immunotherapy approaches. In this study, we found that PD-1 and CTLA4-based cancer immunotherapies are unable to eradicate tumors in elderly mice. This defect in anti-tumor activity correlated with two known age-associated immune defects: diminished abundance of systemic naive CD8+ T cells and weak migratory activities of dendritic cells (DCs). We identified a vaccine adjuvant, referred to as a DC hyperactivator, which corrects DC migratory defects in the elderly. Vaccines containing tumor antigens and DC hyperactivators induced T helper type 1 (TH1) CD4+ T cells with cytolytic activity that drive anti-tumor immunity in elderly mice. When administered early in life, DC hyperactivators were the only adjuvant identified that elicited anti-tumor CD4+ T cells that persisted into old age. These results raise the possibility of correcting age-associated immune defects through DC manipulation.

A multivalent mRNA monkeypox virus vaccine (BNT166) protects mice and macaques from orthopoxvirus disease.

In response to the 2022 outbreak of mpox driven by unprecedented human-to-human monkeypox virus (MPXV) transmission, we designed BNT166, aiming to create a highly immunogenic, safe, accessible, and scalable next-generation vaccine against MPXV and related orthopoxviruses. To address the multiple viral forms and increase the breadth of immune response, two candidate multivalent mRNA vaccines were evaluated pre-clinically: a quadrivalent vaccine (BNT166a; encoding the MPXV antigens A35, B6, M1, H3) and a trivalent vaccine (BNT166c; without H3). Both candidates induced robust T cell responses and IgG antibodies in mice, including neutralizing antibodies to both MPXV and vaccinia virus. In challenge studies, BNT166a and BNT166c provided complete protection from vaccinia, clade I, and clade IIb MPXV. Furthermore, immunization with BNT166a was 100% effective at preventing death and at suppressing lesions in a lethal clade I MPXV challenge in cynomolgus macaques. These findings support the clinical evaluation of BNT166, now underway (NCT05988203).

Host and pathogen response to bacteriophage engineered against Mycobacterium abscessus lung infection.

Two mycobacteriophages were administered intravenously to a male with treatment-refractory Mycobacterium abscessus pulmonary infection and severe cystic fibrosis lung disease. The phages were engineered to enhance their capacity to lyse M. abscessus and were selected specifically as the most effective against the subject's bacterial isolate. In the setting of compassionate use, the evidence of phage-induced lysis was observed using molecular and metabolic assays combined with clinical assessments. M. abscessus isolates pre and post-phage treatment demonstrated genetic stability, with a general decline in diversity and no increased resistance to phage or antibiotics. The anti-phage neutralizing antibody titers to one phage increased with time but did not prevent clinical improvement throughout the course of treatment. The subject received lung transplantation on day 379, and systematic culturing of the explanted lung did not detect M. abscessus. This study describes the course and associated markers of a successful phage treatment of M. abscessus in advanced lung disease.

Control of gasdermin D oligomerization and pyroptosis by the Ragulator-Rag-mTORC1 pathway.

The process of pyroptosis is mediated by inflammasomes and a downstream effector known as gasdermin D (GSDMD). Upon cleavage by inflammasome-associated caspases, the N-terminal domain of GSDMD forms membrane pores that promote cytolysis. Numerous proteins promote GSDMD cleavage, but none are known to be required for pore formation after GSDMD cleavage. Herein, we report a forward genetic screen that identified the Ragulator-Rag complex as being necessary for GSDMD pore formation and pyroptosis in macrophages. Mechanistic analysis revealed that Ragulator-Rag is not required for GSDMD cleavage upon inflammasome activation but rather promotes GSDMD oligomerization in the plasma membrane. Defects in GSDMD oligomerization and pore formation can be rescued by mitochondrial poisons that stimulate reactive oxygen species (ROS) production, and ROS modulation impacts the ability of inflammasome pathways to promote pore formation downstream of GSDMD cleavage. These findings reveal an unexpected link between key regulators of immunity (inflammasome-GSDMD) and metabolism (Ragulator-Rag).

Herceptin: A First Assault on Oncogenes that Launched a Revolution.

This year's Lasker Clinical Research Award honors H. Michael Shepard, Dennis J. Slamon, and Axel Ullrich for their invention of Herceptin, the first monoclonal antibody that blocks a cancer-causing protein, and for its development as a life-saving therapy for women with breast cancer.

Uganda Genome Resource Enables Insights into Population History and Genomic Discovery in Africa.

Genomic studies in African populations provide unique opportunities to understand disease etiology, human diversity, and population history. In the largest study of its kind, comprising genome-wide data from 6,400 individuals and whole-genome sequences from 1,978 individuals from rural Uganda, we find evidence of geographically correlated fine-scale population substructure. Historically, the ancestry of modern Ugandans was best represented by a mixture of ancient East African pastoralists. We demonstrate the value of the largest sequence panel from Africa to date as an imputation resource. Examining 34 cardiometabolic traits, we show systematic differences in trait heritability between European and African populations, probably reflecting the differential impact of genes and environment. In a multi-trait pan-African GWAS of up to 14,126 individuals, we identify novel loci associated with anthropometric, hematological, lipid, and glycemic traits. We find that several functionally important signals are driven by Africa-specific variants, highlighting the value of studying diverse populations across the region.

Gasdermin D permeabilization of mitochondrial inner and outer membranes accelerates and enhances pyroptosis.

Gasdermin D (GSDMD)-activated inflammatory cell death (pyroptosis) causes mitochondrial damage, but its underlying mechanism and functional consequences are largely unknown. Here, we show that the N-terminal pore-forming GSDMD fragment (GSDMD-NT) rapidly damaged both inner and outer mitochondrial membranes (OMMs) leading to reduced mitochondrial numbers, mitophagy, ROS, loss of transmembrane potential, attenuated oxidative phosphorylation (OXPHOS), and release of mitochondrial proteins and DNA from the matrix and intermembrane space. Mitochondrial damage occurred as soon as GSDMD was cleaved prior to plasma membrane damage. Mitochondrial damage was independent of the B-cell lymphoma 2 family and depended on GSDMD-NT binding to cardiolipin. Canonical and noncanonical inflammasome activation of mitochondrial damage, pyroptosis, and inflammatory cytokine release were suppressed by genetic ablation of cardiolipin synthase (Crls1) or the scramblase (Plscr3) that transfers cardiolipin to the OMM. Phospholipid scramblase-3 (PLSCR3) deficiency in a tumor compromised pyroptosis-triggered anti-tumor immunity. Thus, mitochondrial damage plays a critical role in pyroptosis.

A TOG Protein Confers Tension Sensitivity to Kinetochore-Microtubule Attachments.

The development and survival of all organisms depends on equal partitioning of their genomes during cell division. Accurate chromosome segregation requires selective stabilization of kinetochore-microtubule attachments that come under tension due to opposing pulling forces exerted on sister kinetochores by dynamic microtubule tips. Here, we show that the XMAP215 family member, Stu2, makes a major contribution to kinetochore-microtubule coupling. Stu2 and its human ortholog, ch-TOG, exhibit a conserved interaction with the Ndc80 kinetochore complex that strengthens its attachment to microtubule tips. Strikingly, Stu2 can either stabilize or destabilize kinetochore attachments, depending on the level of kinetochore tension and whether the microtubule tip is assembling or disassembling. These dichotomous effects of Stu2 are independent of its previously studied regulation of microtubule dynamics. Altogether, our results demonstrate how a kinetochore-associated factor can confer opposing, tension-dependent effects to selectively stabilize tension-bearing attachments, providing mechanistic insight into the basis for accuracy during chromosome segregation.

Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1.

Nuclear receptor-binding SET-domain protein 1 (NSD1), a methyltransferase that catalyzes H3K36me2, is essential for mammalian development and is frequently dysregulated in diseases, including Sotos syndrome. Despite the impacts of H3K36me2 on H3K27me3 and DNA methylation, the direct role of NSD1 in transcriptional regulation remains largely unknown. Here, we show that NSD1 and H3K36me2 are enriched at cis-regulatory elements, particularly enhancers. NSD1 enhancer association is conferred by a tandem quadruple PHD (qPHD)-PWWP module, which recognizes p300-catalyzed H3K18ac. By combining acute NSD1 depletion with time-resolved epigenomic and nascent transcriptomic analyses, we demonstrate that NSD1 promotes enhancer-dependent gene transcription by facilitating RNA polymerase II (RNA Pol II) pause release. Notably, NSD1 can act as a transcriptional coactivator independent of its catalytic activity. Moreover, NSD1 enables the activation of developmental transcriptional programs associated with Sotos syndrome pathophysiology and controls embryonic stem cell (ESC) multilineage differentiation. Collectively, we have identified NSD1 as an enhancer-acting transcriptional coactivator that contributes to cell fate transition and Sotos syndrome development.

A Second Space Age Spanning Omics, Platforms, and Medicine Across Orbits.

The recent acceleration of commercial, private, and multi-national spaceflight has created an unprecedented level of activity in low Earth orbit (LEO), concomitant with the highest-ever number of crewed missions entering space and preparations for exploration-class (>1 year) missions. Such rapid advancement into space from many new companies, countries, and space-related entities has enabled a"Second Space Age." This new era is also poised to leverage, for the first time, modern tools and methods of molecular biology and precision medicine, thus enabling precision aerospace medicine for the crews. The applications of these biomedical technologies and algorithms are diverse, encompassing multi-omic, single-cell, and spatial biology tools to investigate human and microbial responses to spaceflight. Additionally, they extend to the development of new imaging techniques, real-time cognitive assessments, physiological monitoring, and personalized risk profiles tailored for astronauts. Furthermore, these technologies enable advancements in pharmacogenomics (PGx), as well as the identification of novel spaceflight biomarkers and the development of corresponding countermeasures. In this review, we highlight some of the recent biomedical research from the National Aeronautics and Space Administration (NASA), Japan Aerospace Exploration Agency (JAXA), European Space Agency (ESA), and other space agencies, and also detail the commercial spaceflight sector's (e.g. SpaceX, Blue Origin, Axiom, Sierra Space) entrance into aerospace medicine and space biology, the first aerospace medicine biobank, and the myriad upcoming missions that will utilize these tools to ensure a permanent human presence beyond LEO, venturing out to other planets and moons.

Allosteric interactions prime androgen receptor dimerization and activation.

The androgen receptor (AR) is a nuclear receptor that governs gene expression programs required for prostate development and male phenotype maintenance. Advanced prostate cancers display AR hyperactivation and transcriptome expansion, in part, through AR amplification and interaction with oncoprotein cofactors. Despite its biological importance, how AR domains and cofactors cooperate to bind DNA has remained elusive. Using single-particle cryo-electron microscopy, we isolated three conformations of AR bound to DNA, showing that AR forms a non-obligate dimer, with the buried dimer interface utilized by ancestral steroid receptors repurposed to facilitate cooperative DNA binding. We identify novel allosteric surfaces which are compromised in androgen insensitivity syndrome and reinforced by AR's oncoprotein cofactor, ERG, and by DNA-binding motifs. Finally, we present evidence that this plastic dimer interface may have been adopted for transactivation at the expense of DNA binding. Our work highlights how fine-tuning AR's cooperative interactions translate to consequences in development and disease.

Hierarchy of RNA functional dynamics.

RNA dynamics play a fundamental role in many cellular functions. However, there is no general framework to describe these complex processes, which typically consist of many structural maneuvers that occur over timescales ranging from picoseconds to seconds. Here, we classify RNA dynamics into distinct modes representing transitions between basins on a hierarchical free-energy landscape. These transitions include large-scale secondary-structural transitions at >0.1-s timescales, base-pair/tertiary dynamics at microsecond-to-millisecond timescales, stacking dynamics at timescales ranging from nanoseconds to microseconds, and other "jittering" motions at timescales ranging from picoseconds to nanoseconds. We review various modes within these three different tiers, the different mechanisms by which they are used to regulate function, and how they can be coupled together to achieve greater functional complexity.

The IgM receptor FcµR limits tonic BCR signaling by regulating expression of the IgM BCR.

The FcµR receptor for the crystallizable fragment (Fc) of immunoglobulin M (IgM) can function as a cell-surface receptor for secreted IgM on a variety of cell types. We found here that FcµR was also expressed in the trans-Golgi network of developing B cells, where it constrained transport of the IgM-isotype BCR (IgM-BCR) but not of the IgD-isotype BCR (IgD-BCR). In the absence of FcµR, the surface expression of IgM-BCR was increased, which resulted in enhanced tonic BCR signaling. B-cell-specific deficiency in FcµR enhanced the spontaneous differentiation of B-1 cells, which resulted in increased serum concentrations of natural IgM and dysregulated homeostasis of B-2 cells; this caused the spontaneous formation of germinal centers, increased titers of serum autoantibodies and excessive accumulation of B cells. Thus, FcµR serves as a critical regulator of B cell biology by constraining the transport and cell-surface expression of IgM-BCR.

Identification of multipotent luminal progenitor cells in human prostate organoid cultures.

The prostate gland consists of basal and luminal cells arranged as pseudostratified epithelium. In tissue recombination models, only basal cells reconstitute a complete prostate gland, yet murine lineage-tracing experiments show that luminal cells generate basal cells. It has remained challenging to address the molecular details of these transitions and whether they apply to humans, due to the lack of culture conditions that recapitulate prostate gland architecture. Here, we describe a 3D culture system that supports long-term expansion of primary mouse and human prostate organoids, composed of fully differentiated CK5+ basal and CK8+ luminal cells. Organoids are genetically stable, reconstitute prostate glands in recombination assays, and can be experimentally manipulated. Single human luminal and basal cells give rise to organoids, yet luminal-cell-derived organoids more closely resemble prostate glands. These data support a luminal multilineage progenitor cell model for prostate tissue and establish a robust, scalable system for mechanistic studies.

Organoid cultures derived from patients with advanced prostate cancer.

The lack of in vitro prostate cancer models that recapitulate the diversity of human prostate cancer has hampered progress in understanding disease pathogenesis and therapy response. Using a 3D organoid system, we report success in long-term culture of prostate cancer from biopsy specimens and circulating tumor cells. The first seven fully characterized organoid lines recapitulate the molecular diversity of prostate cancer subtypes, including TMPRSS2-ERG fusion, SPOP mutation, SPINK1 overexpression, and CHD1 loss. Whole-exome sequencing shows a low mutational burden, consistent with genomics studies, but with mutations in FOXA1 and PIK3R1, as well as in DNA repair and chromatin modifier pathways that have been reported in advanced disease. Loss of p53 and RB tumor suppressor pathway function are the most common feature shared across the organoid lines. The methodology described here should enable the generation of a large repertoire of patient-derived prostate cancer lines amenable to genetic and pharmacologic studies.

Inflammasomes: Threat-Assessment Organelles of the Innate Immune System.

Inflammasomes are supramolecular organizing centers that operate to drive interleukin-1 (IL-1)-dependent inflammation. Depending on context, inflammatory caspases act upstream or downstream of inflammasome assembly, serving as the principal enzymes that control activities of these organelles. In this review, we discuss mechanisms of inflammasome assembly and signaling. We posit that upstream regulatory proteins, classically known as pattern-recognition receptors, operate to assess infectious and non-infectious threats to the host. Threat assessment is achieved through two general strategies: (1) direct binding of receptors to microbial or host-derived ligands or (2) indirect detection of changes in cellular homeostasis. Upon activation, these upstream regulatory factors seed the assembly of inflammasomes, leading to IL-1 family cytokine release from living (hyperactive) or dead (pyroptotic) cells. The molecular and physiological consequences of these distinct cell fate decisions are discussed.

Defying Death: The (W)hole Truth about the Fate of GSDMD Pores.

Pyroptosis is an inflammatory cell death response initiated by supramolecular organizing centers known as inflammasomes. In a recent issue of Science, Rühl et al. (2018) challenge the paradigm that inflammasome signaling necessitates pyroptosis by demonstrating that ESCRTIII-dependent membrane repair can delay or prevent gasdermin D-mediated cell death.

Glucocorticoid receptor confers resistance to antiandrogens by bypassing androgen receptor blockade.

The treatment of advanced prostate cancer has been transformed by novel antiandrogen therapies such as enzalutamide. Here, we identify induction of glucocorticoid receptor (GR) expression as a common feature of drug-resistant tumors in a credentialed preclinical model, a finding also confirmed in patient samples. GR substituted for the androgen receptor (AR) to activate a similar but distinguishable set of target genes and was necessary for maintenance of the resistant phenotype. The GR agonist dexamethasone was sufficient to confer enzalutamide resistance, whereas a GR antagonist restored sensitivity. Acute AR inhibition resulted in GR upregulation in a subset of prostate cancer cells due to relief of AR-mediated feedback repression of GR expression. These findings establish a mechanism of escape from AR blockade through expansion of cells primed to drive AR target genes via an alternative nuclear receptor upon drug exposure.

Direct observation of coordinated assembly of individual native centromeric nucleosomes.

Eukaryotic chromosome segregation requires the kinetochore, a megadalton-sized machine that forms on specialized centromeric chromatin containing CENP-A, a histone H3 variant. CENP-A deposition requires a chaperone protein HJURP that targets it to the centromere, but it has remained unclear whether HJURP has additional functions beyond CENP-A targeting and why high AT DNA content, which disfavors nucleosome assembly, is widely conserved at centromeres. To overcome the difficulties of studying nucleosome formation in vivo, we developed a microscopy assay that enables direct observation of de novo centromeric nucleosome recruitment and maintenance with single molecule resolution. Using this assay, we discover that CENP-A can arrive at centromeres without its dedicated centromere-specific chaperone HJURP, but stable incorporation depends on HJURP and additional DNA-binding proteins of the inner kinetochore. We also show that homopolymer AT runs in the yeast centromeres are essential for efficient CENP-A deposition. Together, our findings reveal requirements for stable nucleosome formation and provide a foundation for further studies of the assembly and dynamics of native kinetochore complexes.

DASH/Dam1 complex mutants stabilize ploidy in histone-humanized yeast by weakening kinetochore-microtubule attachments.

Forcing budding yeast to chromatinize their DNA with human histones manifests an abrupt fitness cost. We previously proposed chromosomal aneuploidy and missense mutations as two potential modes of adaptation to histone humanization. Here, we show that aneuploidy in histone-humanized yeasts is specific to a subset of chromosomes that are defined by their centromeric evolutionary origins but that these aneuploidies are not adaptive. Instead, we find that a set of missense mutations in outer kinetochore proteins drives adaptation to human histones. Furthermore, we characterize the molecular mechanism underlying adaptation in two mutants of the outer kinetochore DASH/Dam1 complex, which reduce aneuploidy by suppression of chromosome instability. Molecular modeling and biochemical experiments show that these two mutants likely disrupt a conserved oligomerization interface thereby weakening microtubule attachments. We propose a model through which weakened microtubule attachments promote increased kinetochore-microtubule turnover and thus suppress chromosome instability. In sum, our data show how a set of point mutations evolved in histone-humanized yeasts to counterbalance human histone-induced chromosomal instability through weakening microtubule interactions, eventually promoting a return to euploidy.