Tumors evolve to avoid immune destruction and establish an immunosuppressive microenvironment. Syngeneic mouse tumor models are critical for understanding tumor immune evasion and testing cancer immunotherapy. Derived fr...Tumors evolve to avoid immune destruction and establish an immunosuppressive microenvironment. Syngeneic mouse tumor models are critical for understanding tumor immune evasion and testing cancer immunotherapy. Derived from established mouse tumor cell lines that can already evade the immune system, these models cannot simulate early phases of immunoediting during initial tumorigenesis. We developed a syngeneic mouse teratoma model derived from noncancerous mouse embryonic stem cells and conducted a genome-wide CRISPR screen to identify genes that impact early phases of cancer immunoediting. We found that loss of pro-apoptotic tumor suppressor genes, including Trp53, increased necrosis in teratomas, releasing APOE lipid particles into the extracellular milieu. Infiltrating T cells drawn to tumor necrotic regions accumulated lipids and became dysfunctional. Blocking lipid uptake in T cells or reducing necrosis in teratomas by inactivating the mitochondrial permeability transition pore (mPTP) restored immunosurveillance. Because mouse teratomas were highly enriched for brain tissues, we next examined the tumor-immune interaction in human glioblastoma (GBM). Indeed, infiltrating T cells in TP53-mutated human GBM accumulated APOE and were dysfunctional. Anti-APOE and anti-PDCD1 antibodies synergistically boosted anti-GBM immunity and prolonged survival in mice. Our results link mPTP-mediated tumor necrosis to immune evasion and suggest that targeting the uptake of lipids released by necrotic tumor cells by infiltrating immune cells can enhance cancer immunotherapy.
The brain's hemispheres exhibit profound lateralization, yet the underlying mechanisms remain elusive. Using proteomic and phosphoproteomic analyses of the bilateral striatum - a hub for important brain functions and a c...The brain's hemispheres exhibit profound lateralization, yet the underlying mechanisms remain elusive. Using proteomic and phosphoproteomic analyses of the bilateral striatum - a hub for important brain functions and a common node of autism pathophysiology - we identified significant phosphorylation asymmetries. Particularly, the phosphorylation processes in the left striatum appear more prone to disturbance. Notably, SH3RF2, whose single-copy knockout leads to autism spectrum disorder (ASD)-like behaviors in mice, is uniquely expressed in the striatum, forming a complex with CaMKII (an ASD-associated protein) and PPP1CC. Loss of SH3RF2 disturbs the CaMKII/PP1 "switch", resulting in hyperactivity of CaMKII and increased phosphorylation of its substrate GluR1. In Sh3rf2-deficient mice, heightened GluR1-Ser831 phosphorylation and its aberrant postsynaptic membrane localization in the left striatum may impair the functional lateralization of striatal neurons and contribute to autism-like behaviors. This study unveils the first molecular mechanism governing brain lateralization in mammals, linking its impairment to autism development and treatment strategies.
The γ-carboxylation state of osteocalcin determines its essential functions in bone mineralization or systemic metabolism and serves as a prominent biomarker for bone health and vitamin K nutrition. This post-translation...The γ-carboxylation state of osteocalcin determines its essential functions in bone mineralization or systemic metabolism and serves as a prominent biomarker for bone health and vitamin K nutrition. This post-translational modification of glutamate residues is catalyzed by the membrane-embedded vitamin K-dependent γ-carboxylase (VKGC), which typically recognizes protein substrates through their tightly bound propeptide that triggers γ-carboxylation. However, the osteocalcin propeptide exhibits negligible affinity for VKGC. To understand the underlying molecular mechanism, we determined the cryo-electron microscopy structures of VKGC with osteocalcin carrying a native propeptide or a high-affinity variant at different carboxylation states. The structures reveal a large chamber in VKGC that maintains uncarboxylated and partially carboxylated osteocalcin in partially unfolded conformations, allowing their glutamate-rich region and C-terminal helices to engage with VKGC at multiple sites. Binding of this mature region together with the low-affinity propeptide effectively stimulates VKGC activity, similar to high-affinity propeptides that differ only in closely fitting interactions. However, the low-affinity propeptide renders osteocalcin prone to undercarboxylation at low vitamin K levels, thereby serving as a discernible biomarker. Overall, our studies reveal the unique interaction of osteocalcin with VKGC and provide a framework for designing therapeutic strategies targeting osteocalcin-related bone and metabolic disorders.
Liu K, Yan Z, Bai D
… +27 more, Jiang R, Bi Y, Ma X, Xiang J, Sheng Y, Dong B, Ning Z, Yi S, Liu Y, Lei X, Jia Y, Zhang Y, Zhang Y, Li Y, Wu T, Xi C, Liu S, Liu S, Chen J, Yin J, Kou X, Zhao Y, Wang H, Wang Y, Wei K, Gao S, Liu W
The absence of stem cells capable of efficiently generating both trophoblast and epiblast lineages has hindered precise recapitulation of embryonic development. Through high-content chemical screening, we established an...The absence of stem cells capable of efficiently generating both trophoblast and epiblast lineages has hindered precise recapitulation of embryonic development. Through high-content chemical screening, we established an (AS and LY) AL medium to generate mouse bidirectional pluripotent stem cells (BPSCs) characterized by concurrent expression of OCT4 and CDX2. Mouse BPSCs demonstrated highly plastic differentiation into trophoblast, epiblast and primitive endoderm (PrE) lineages in vitro within 48 h without exogenous inducing factors and efficiently contributed to embryonic and extraembryonic tissues in vivo. Mechanistically, hyperactivation of the Wnt signaling pathway breaks the early lineage differentiation barrier by initiating a Lef1-dependent bypass. Remarkably, integration of BPSCs with PrE induction system enables high-efficiency generation of E8.5-stage embryo models. These advanced models complete gastrulation and recapitulate definitive developmental milestones including brain morphogenesis, neural tube closure, cardiac contraction, somite patterning, and primordial germ cell specification. Moreover, human cells cultured under AL conditions acquire an OCT4 and CDX2 double-positive state and corresponding gene expression profiles, revealing conserved functionality of this culturing platform across species. These findings highlight BPSCs as a powerful tool for investigating early lineage specification and post-gastrulation embryonic development.
Since their first report a decade ago, our understanding of migrasomes - specialized organelles initially identified in migrating cells-has advanced considerably. Researchers have elucidated key aspects of migrasome biol...Since their first report a decade ago, our understanding of migrasomes - specialized organelles initially identified in migrating cells-has advanced considerably. Researchers have elucidated key aspects of migrasome biology, including the mechanisms of their biogenesis, their roles in cellular physiology, and their implications in various diseases. Concurrently, the development of a robust toolkit for migrasome analysis has transformed these structures from mere microscopy curiosities into central players in an emerging field with significant impact on cell biology, developmental biology, immunology, and disease pathology. This review provides a comprehensive summary of current insights into migrasome biology, with a particular focus on the molecular mechanisms governing their formation and their established cellular and physiological functions. In addition, we highlight the current challenges and unresolved questions that continue to shape and propel future research in this exciting area of study.
The vast scope but limited-supporting evidence in sequence databases hinders identification of proteins with specific functionality. Here, we experimentally characterized catalytic efficiency, target site window, motif p...The vast scope but limited-supporting evidence in sequence databases hinders identification of proteins with specific functionality. Here, we experimentally characterized catalytic efficiency, target site window, motif preference, and off-target activity of 1100 apolipoprotein B mRNA-editing enzyme, catalytic polypeptide (APOBEC)-like family cytidine deaminases (CDs) fused with nCas9 in HEK293T cells, thereby generating the largest dataset of experimentally validated functions for a single protein family to date. These data, together with amino acid sequence, three-dimensional structure, and eight additional features, were used to construct a machine learning (ML) model, AlphaCD, which showed high accuracy in predicting catalytic efficiency (0.92) and off-target activity (0.84), as well as target windows (0.73) and catalytic motifs (0.78). We applied the trained model to predict the above catalytic features of 21,335 CDs in Uniprot, and subsampling of 28 CDs further validated its prediction accuracy (0.84, 0.87, 0.75, 0.73, respectively). Alanine scanning-based mutagenesis was then employed to reduce off-targets in one example CD, which produced a remarkably high fidelity, high efficiency cytosine base editor, thus demonstrating AlphaCD application in high-accuracy, high-throughput protein functional characterization, and providing a strategy for accelerated characterization of other proteins.
Roughly 1 billion people are infected by Influenza A viruses (IAVs) worldwide each year, resulting in approximately half a million deaths. Particularly concerning is the threat of IAV spillover from avian and other anima...Roughly 1 billion people are infected by Influenza A viruses (IAVs) worldwide each year, resulting in approximately half a million deaths. Particularly concerning is the threat of IAV spillover from avian and other animal reservoirs. The recent outbreak of highly pathogenic avian influenza H5N1 in US dairy cows highlights this concern. While viruses that enter human populations from such zoonotic transmission typically lack the ability to transmit effectively between humans, they may be only a few mutations from acquiring this capacity. These newly adapted viruses have the potential to be significantly more virulent than seasonal strains. A major contributor to influenza pathology is the over-exuberant immune response to the virus, particularly when the infection is present in distal pulmonary tissues. Maladaptive immune pathway over-activation can drive tissue damage and pathology, often independently of effective viral control. Anti-inflammatories targeting host-initiated pathological processes hold promise, but these avenues require a thorough understanding of virus-triggered lung inflammation before they can be fully exploited. In this review, we will discuss recent advances in our understanding of the cell types that are targeted by IAV, the consequences of IAV infection on the biology of these cells, and their contribution to lung pathology in influenza. We will also discuss how virus-induced hyper-inflammatory responses present new entry-points for therapeutic intervention, showcasing Z-form nucleic acid-binding protein 1 (ZBP1)-initiated necroptosis as an example of one such pathway.
Huang M, Chen M, Yuan G
… +16 more, Cui Y, Shen B, Liu Z, Zhang B, Chen J, Chen D, Qiu S, Zhang Y, Liu L, Qin L, Zhu Y, Liu J, Zhang H, Wu J, Yuan Y, Sha J
Pluripotent stem cells (PSCs) have been derived from various species, but most culture systems stabilize only a single PSC type. By contrast, epiblast cells in vivo exist along a continuum and interact dynamically with b...Pluripotent stem cells (PSCs) have been derived from various species, but most culture systems stabilize only a single PSC type. By contrast, epiblast cells in vivo exist along a continuum and interact dynamically with both embryonic and extraembryonic cells, interactions missing in standard PSC cultures. This absence limits the self-organizing potential of PSCs and leads to disorganized tissue formation in teratomas. To address this, we developed a unified culture system that supports the stable differentiation of epiblast-like cells into multiple key human gastrulating cell types, collectively called human gastrulating stem cells (hGaSCs). hGaSCs, composed of endoderm-like, mesoderm-like, ectoderm-like, amnion ectoderm-like, and primordial germ cell-like cells, maintain a stable balance during long-term culture. In 3D culture, hGaSCs self-assemble into gastruloid-like structures (hGaSC-gastruloids) that model aspects of a Carnegie Stage 7 human embryo, including gastrulation and germ layer specification. Using hGaSC-gastruloids, we modeled the effects of valproic acid (VPA) on human gastrulation and uncovered molecular pathways underlying VPA-induced malformations. When transplanted into the seminiferous tubules, hGaSCs formed embryo-like structures, progressing through fetal tissue and organ development, unlike the disorganized growth seen in teratomas. In conclusion, hGaSCs provide a versatile platform to study human gastrulation, early organogenesis, developmental defects, and drug teratogenicity, with promising applications in tissue and organ generation from cultured stem cells.
Engram cells storing episodic memories are allocated to separate neuronal ensembles. However, how these ensembles maintain their stability to drive precise memory expression, and whether their destabilization contributes...Engram cells storing episodic memories are allocated to separate neuronal ensembles. However, how these ensembles maintain their stability to drive precise memory expression, and whether their destabilization contributes to aging-related memory deficits, remain elusive. Here, we show that during contextual fear memory consolidation, neuronal pentraxin 1 (NPTX1) in Fos transcription-dependent ensemble (F-RAM) of the dentate gyrus (DG) promotes memory expression in the fear context. NPTX1 facilitates K7.2 channel-mediated inhibition of engram cell hyperexcitability, thereby restricting the response of these cells to excitatory inputs from medial entorhinal cortex. Meanwhile, NPTX2 enhances the perisomatic inhibition of Npas4 transcription-dependent ensemble (N-RAM) by parvalbumin (PV) interneurons, thereby preventing fear memory overgeneralization. Pharmacological activation of K7.2 channels or chemogenetic activation of PV interneurons repaired memory deficits caused by engram-specific NPTX depletion. Contextual fear memory precision and NPTX expression in DG engram cells were decreased in aged mice. Overexpressing NPTX1 in F-RAM ensemble or the AMPAR-binding domain of NPTX2 in N-RAM ensemble rescued contextual fear memory deficits. These findings elucidate that the coordination of NPTX1 and NPTX2 prevents engram ensembles from becoming hyperactive and provide a causal link between engram network destabilization and aging-related contextual fear memory deficits.
KCNQ1 potassium channels are essential for physiological processes such as cardiac rhythm and intestinal chloride secretion. KCNE family subunits (KCNE1-5) associate with KCNQ1, conferring distinct properties across vari...KCNQ1 potassium channels are essential for physiological processes such as cardiac rhythm and intestinal chloride secretion. KCNE family subunits (KCNE1-5) associate with KCNQ1, conferring distinct properties across various tissues. KCNQ1 activation requires membrane depolarization and phosphatidylinositol 4,5-bisphosphate (PIP2) whose cellular levels are controlled by Gαq-coupled GPCR activation. While modulation of KCNQ1's voltage-dependent activation by KCNE1/3 is well-characterized, their effects on PIP2-dependent gating of KCNQ1 via GPCR signaling remain less understood. Here we resolved structures of KCNQ1-KCNE1 and reassessed the reported KCNQ1-KCNE3 structures with and without PIP2. We revealed that KCNQ1-KCNE1/3 complexes feature two PIP2-binding sites, with KCNE1/3 contributing to a previously overlooked, uncharacterized site involving residues critical for coupling voltage sensor and pore domains. Via this site, KCNE1 and KCNE3 distinctly modulate the PIP2-dependent gating, in addition to the voltage sensitivity, of KCNQ1. Consequently, KCNE3 converts KCNQ1 into a voltage-insensitive PIP2-gated channel governed by GPCR signaling to maintain ion homeostasis in non-excitable cells. KCNE1, by significantly enhancing KCNQ1's PIP2 affinity and resistance to GPCR regulation, forms predominantly voltage-gated channels with KCNQ1 for conducting the slow-delayed rectifier current in excitable cardiac cells. Our study highlights how KCNE1/3 modulates KCNQ1 gating in different cellular contexts, providing insights into tissue-specifically targeting multi-functional channels.