Biology subjects

Akiyama, H.

Publications and source records attributed to Akiyama, H..

At least 19 recordsLinked to original sources

bioRxiv2026

Engineering robust cardiomyocyte differentiation from iPSCs through WNT-mediated process window expansion

Lack of robustness and high variability in cardiomyocyte (CM) differentiation from human induced pluripotent stem cells (hiPSCs) remain critical challenges for cardiac regenerative medicine and biological research. Guided by developmental principles of lineage specification, we investigated whether early WNT inhibition could expand the process window of CM differentiation and thereby improve robustness. To test this, we compared early WNT inhibition from days 1--5 with standard inhibition from days 3--5 following a 1-day CHIR99021 (CHIR) treatment for WNT activation. Focusing on CHIR concentration as a critical parameter, we found that early WNT inhibition substantially broadened the concentration window yielding [≥]80% differentiation efficiency across distinct hiPSC lines. Extending the analysis to a two-dimensional parameter space incorporating cell density revealed a markedly expanded [≥]80% process window. Temporal analysis of lineage-specific transcription factor expression showed that early WNT inhibition suppressed paraxial mesoderm (PM) specification, likely through MSGN1 suppression, in the range of high initial WNT activation that otherwise favored PM fate, thereby extending the process window. It also suppressed definitive endoderm (DE) specification within the expanded window, promoting a more favorable lateral plate mesoderm/cardiac fate. We further demonstrated substantially reduced variability under parameter perturbations and across wells and batches, accompanied by higher differentiation efficiency, without compromising CM phenotypes. These findings demonstrate early WNT inhibition as an effective strategy for expanding the process window of CM differentiation and improving robustness, while exemplifying the process engineering principle that a broader window can support more robust differentiation, with potential relevance to stem cell--based bioprocesses.

bioengineering↗

bioRxiv2026

Osteocyte State Transitions Modulate Bone Remodeling During Early Skeletal Aging

Osteocytes are long-lived cells that play a central role in bone homeostasis, yet age-related changes in their functional states remain poorly understood, particularly because skeletal aging involves multiple processes beyond cellular senescence. We generated an osteocyte-specific MepeCre mouse line and combined osteocyte ablation in young and middle-aged mice with skeletal phenotyping, single-cell transcriptomics, and senolytic treatment. MepeCre-driven recombination was largely confined to osteocytes, with minimal off-target activity. Osteocyte ablation increased bone mass at both ages, indicating that osteocytes constrain bone accrual as part of their role in skeletal homeostasis. However, the accompanying remodeling changes differed with age: enhanced osteoblast activity predominated in young mice, whereas reduced osteoclast-mediated bone resorption predominated in middle-aged mice. Single-cell transcriptomics revealed distinct osteocyte subpopulations whose relative abundance shifted with age, from a predominantly matrix-enriched state in young mice to an expanded aging-transitional state in middle-aged mice. Although this state showed partial enrichment of senescence-associated transcriptional signatures, senolytic treatment failed to recapitulate the increase in bone mass induced by osteocyte ablation. Osteocyte therefore regulate bone mass through age-dependent mechanisms that coincide with shifts in osteocyte-state composition. These changes emerge by middle age and may contribute to early remodeling imbalance before overt cellular senescence during skeletal aging. Graphical AbstractGraphical summary of the findings of this study. AA, amino acids; NA, nucleic acid; UA, uric acid; TCA, tricarboxylic acid.

molecular biology↗

bioRxiv2026

Evaluation of Retinal Safety of Hypoxia-Inducible Factor Prolyl Hydroxylase Inhibitors

Hypoxia-inducible factor prolyl hydroxylase (HIF-PH) inhibitors are widely used for the treatment of renal anemia; however, their effects on intraocular vascular endothelial growth factor (VEGF) expression remain unclear. In this study, we examined the effects of all five HIF-PH inhibitors --roxadustat, daprodustat, vadadustat, enarodustat, and molidustat--on Vegfa expression in the retina in mice. C57BL/6J mice were orally administered each inhibitor. Six hours after administration, the kidney, retina, and liver were collected, and transcription levels were quantified by real-time quantitative reverse transcription PCR. Renal Epo transcription was significantly increased by molidustat (P < 0.01), roxadustat (P < 0.01), and enarodustat (P < 0.05). Retinal Vegfa transcription was significantly increased by four inhibitors (P < 0.01), with molidustat showing no significant effect. In the liver, Vegfa transcription was increased by daprodustat (P < 0.05) and vadadustat (P < 0.01). Furthermore, renal Epo and retinal Vegfa transcription levels showed a moderate positive correlation with a marginal trend toward statistical significance (r = 0.37, P = 0.08). These findings indicate that HIF-PH inhibitors differentially regulate hypoxia-responsive genes across tissues and suggest that retinal VEGF upregulation should be considered when evaluating the safety of these agents.

pharmacology and toxicology↗

bioRxiv2025

Engineering plant tandem kinase immune receptors expands effector recognition profiles

Plant intracellular immune receptors are widely deployed in breeding to protect crops from disease. In addition to nucleotide-binding leucine-rich repeat receptors (NLRs), tandem kinase proteins (TKPs) have recently emerged as an important family of immune receptors within staple cereal food crops, but how TKPs recognize effectors and whether they are amenable to engineering is essentially unknown. Here, we show that the barley and wheat TKPs Rmo2 and Rwt7 recognize different blast fungus effectors via their integrated HMA domains using different protein interfaces with nanomolar binding affinity. Structural analysis pinpointed interface residues that dictate effector recognition and enabled engineering of dual-specificity TKPs. These results establish integrated HMA domains as programmable modules within TKPs for designing new specificities in plant immunity for diseases relevant to global agriculture.

plant biology↗

bioRxiv2025

Hox - Meis-relayed topographical genetic switch underlies cardiopharyngeal neural crest diversification, revealed by multimodal analysis

Neural crest cells (NCCs) are multipotent migratory cells essential for cardiac development, yet the lineage trajectories and gene regulatory networks (GRNs) underlying their differentiation in the cardiopharyngeal region remain unclear. Here, we integrate single-cell RNA-seq, spatial transcriptomics, and multiomic analyses to construct a comprehensive map of NCC lineages in developing mouse cardiopharyngeal tissues. We identify a transition from Hox-positive pharyngeal NCCs to Hox-negative intracardiac populations associated with the outflow tract cushion, accompanied by a shift in Meis transcription factor binding and GRN architecture. By contrast, NCCs forming the aorticopulmonary septum and great vessel smooth muscle retain distinct Hox-codes. A Meis2-Sox9-Scx GRN defines a skeletogenic progenitor-like intermediate state that gives rise to coronary artery smooth muscle and semilunar valves. Our findings suggest that the loss of Hox-dependent regional identity enables pharyngeal NCCs to acquire new fates upon entering the cardiac cushion, providing insight into the developmental origins of coronary and valvular calcification.

developmental biology↗

bioRxiv2025

A scalable proteogenomic framework for dissecting phospho-signaling pathways in primary immune cells

Signaling networks modulated by post-translational modifications orchestrate cellular responses to external cues. Traditional approaches to study these pathways lack the throughput to systematically capture the causal architecture of these signaling pathways at scale. Here, we present an integrated proteogenomic framework that combines saturating genetic perturbations with high-throughput proteomics to systematically map cytokine-induced signaling in primary human T cells. Supporting this framework is simplePhos, a streamlined, low-input phosphoproteomics workflow that enables scalable, time-resolved analysis without the requirement for specialized equipment or robotics. We extensively validate the simplePhos pipeline by applying inflammatory stimuli, including type I and II interferons, lipopolysaccharide, and Sendai virus to primary T cells and myeloid cells, establishing foundational datasets in these treatment contexts. Ultimately, using type I interferon signaling in genetically modified T cells as a model, we demonstrate that combined application of genetic alterations and proteomic analyses can map key signaling nodes in primary immune cells. This represents a powerful strategy to mechanistically interrogate phospho-signaling networks in human immune cells, with broad applications in translational immunology and therapeutic development.

systems biology↗

bioRxiv2025

Self-organized fingering instabilities drive the emergence of tissue morphogenesis in digit organoids.

The emergence of complex anatomical structures -such as the hands- from unstructured tissues remains a fundamental question in developmental biology. Turing-type reaction-diffusion models have provided a molecular explanation for the periodic pre-patterning of digits; however, the physical principles driving 3D morphogenesis remain incompletely understood. To identify the biophysical design principles leading to digit formation, we develop a limb-mesenchymal organoid system that spontaneously forms elongated, digit-like protrusions. Iterations between experiments and agent-based models at the cellular level identify sufficient microscopic mechanisms leading to morphogenesis of digit-like structures: symmetry-breaking and the elongation of digits result from a combination of differential cell adhesion and morphogen-induced chemotaxis and convergent-extension. Lastly, to describe tissue-scale deformations, we perform a coarse-graining analysis of the agent-based model and derive a continuum model that reveals a structural analogy to Cahn-Hilliard-type equations. These equations are typically used to describe fluid phase separation and so-called ''fingering instabilities'' in fluid physics. Here, we show that they also accurately describe organoid morphogenesis. These findings suggest that ''finger'' formation is driven by a mechanical fingering instability acting in concert with chemical patterning, shedding a new light on vertebrate limb morphogenesis.

developmental biology↗

bioRxiv2025

5'' cap sequestration is required for sensing of unspliced HIV-1 RNA by MDA5

Heterogenous transcription start site (TSS) usage dictates the structure and function of unspliced HIV-1 RNAs (usRNA). We and others have previously reported that expression and Rev/CRM1-mediated nuclear export of HIV-1 usRNA in macrophages activates MDA5, MAVS, and innate immune signaling cascades. In this study, we reveal that MDA5 sensing of viral usRNA is strictly determined by TSS, 5 leader structure, and RNA function. We show that cap-sequestered HIV-1 usRNAs (cap1G) destined for viral genome packaging are specifically targeted by MDA5, while translation-destined (cap3G) usRNAs are remarkably immunologically silent. Using mutant viruses which express usRNA with altered 5 cap-exposed leader structure, or inclusion of a retroviral constitutive transport element which drives mRNA-like NXF1-dependent nuclear export of viral usRNA, we show that cap exposure and nuclear export pathway choice are major determinants of both lentiviral RNA immunogenicity and function. In total, we identify innate immune system evasion as a possible rationale for the universal conservation of heterogenous TSS usage among ancestral and extant HIV-1 isolates and shed light on how MDA5 fundamentally discriminates between self and non-self RNAs. SIGNIFICANCEInnate immune activation is critical to both the process of initial infection establishment and ongoing chronic inflammation in HIV-1 infection. While MDA5 has been identified as the sensor which detects unspliced HIV-1 RNA produced in infected cells, it remains unclear how HIV-1 unspliced RNAs, which are generated by cellular transcriptional processes, are recognized as non-self. Here, we reveal that HIV-1 RNA function determines MDA5-driven immunogenicity. We show that only unspliced RNAs which traffic to membrane-associated viral assembly sites are immunogenic, while unspliced RNAs which are ribosomally translated to produce viral proteins are immunologically silent. These findings not only advance our knowledge of how the human innate immune system recognizes HIV-1 unspliced RNAs as foreign but also provide a rationale for the selective advantage to generate two pools of unspliced RNAs during HIV-1 replication.

microbiology↗

bioRxiv2025

Evasion of CARD8 Activation During HIV-1 Assembly

As intracellular parasites, viruses must devise sophisticated mechanisms to produce and assemble viral components while suppressing activation of innate immune effectors. Here, we report that coordination of HIV-1 assembly by the viral polyprotein Gag suppresses inappropriately-timed protease (PR) activity to evade the PR activity sensor, CARD8. Employing mutants of Gag, we show that disruption of domains controlling viral assembly site (MA) or virus particle release (NC and p6) lead to premature activation of PR and the CARD8 inflammasome, resulting in IL-1{beta} secretion and pyroptotic cell death. Further, we demonstrate that previously-observed host-adaptive mutations in HIV-1 MA (M30K) and p6 (PTAP duplication) associated with greater fitness in humans differentially modulate the process of viral assembly and budding to evade CARD8-mediated cell death. Altogether, this work reveals adaptation to human CARD8 by HIV-1 Gag upon zoonotic transmission from chimpanzees and suggests that assembly-regulated CARD8 activation influences the trajectory of HIV-1 evolution and fitness in humans.

microbiology↗

bioRxiv2025

Engineering a Controlled Cardiac Multilineage Co-Differentiation Process Using Statistical Design of Experiments

BackgroundThe heart is a complex organ composed of diverse cell types, whose interplay is crucial for development, tissue homeostasis, and disease progression. A potential approach to recapitulate this heterotypic multicellular environment is the co-differentiation of induced pluripotent stem cells (iPSCs) into multiple cardiac cell types, facilitating the unlocking of their full potential for regenerative medicine and drug development. However, the inherent complexity of co-differentiation, where multiple differentiation factors simultaneously influence the induction of multiple cell types, presents a significant challenge in achieving high controllability over the process toward the desired outcome. Thus, a robust strategy is essential for engineering a controlled co-differentiation process with broader applicability. MethodsGiven the importance of heterotypic cellular proportions in facilitating proper interactions, we present a new strategy to engineer a controlled cardiac co-differentiation process from iPSCs using statistical design of experiments to simultaneously generate cardiomyocytes, mural cells, and endothelial cells, which are major constituents of the heart. ResultsWe divided the process into two stages: progenitor cell induction and the subsequent trilineage co-differentiation, allowing for stage-specific optimization. Given that the performance of progenitor cell induction may critically influence the overall process performance, we carefully optimized activin A and CHIR-99021 using the sequential design of experiments to achieve approximately 95% induction efficiency of KDR+/PDGFR-+ cardiogenic mesoderm cells from iPSCs with minimal batch-to-batch variability. In the trilineage co-differentiation stage, we developed unique multi-response models to delineate trilineage co-differentiation ratios within a defined parameter space of WNT signal inhibitor and vascular endothelial growth factor. This enabled the identification of potential conditions that steer co-differentiation toward desired cellular constitutions, a critical factor of effective cellular interplays. Repeated trilineage co-differentiation experiments confirmed the high process controllability, with a close match between actual and predicted differentiation ratios. Furthermore, cardiomyocytes from trilineage co-differentiation exhibited a more mature sarcomere gene expression profile than those from monolineage differentiation. ConclusionsThese results highlight the effectiveness of our strategy for engineering a stem cell co-differentiation process and its applicability where multicellular interactions are crucial.

bioengineering↗

bioRxiv2024

Expression of Intron-containing HIV-1 RNA Induces NLRP1 Inflammasome Activation in Myeloid Cells

Despite the success of antiretroviral therapy (ART) in suppressing plasma viremia in people living with human immunodeficiency virus type-1 (HIV-1), persistent viral RNA expression in tissue reservoirs is observed and can contribute to HIV-1-induced immunopathology and comorbidities. Infection of long-lived innate immune cells, such as tissue-resident macrophages and microglia may contribute to persistent viral RNA production and chronic inflammation. We recently reported that de-novo cytoplasmic expression of HIV-1 intron-containing RNA (icRNA) in macrophages and microglia leads to MDA5 and MAVS-dependent innate immune sensing and induction of type I IFN responses, demonstrating that HIV icRNA is a pathogen-associated molecular pattern (PAMP). In this report, we show that cytoplasmic expression of HIV-1 icRNA also induces NLRP1 inflammasome activation and IL-1{beta} secretion in macrophages and microglia in a RLR- and endosomal TLR-independent manner. Infection of both macrophages and microglia with either replication-competent or single-cycle HIV-1 induced IL-1{beta} secretion, which was attenuated when cytoplasmic expression of viral icRNA was prevented. While IL-1{beta} secretion was blocked by treatment with caspase-1 inhibitors or knockdown of NLRP1 or caspase-1 expression in HIV- infected macrophages, overexpression of NLRP1 significantly enhanced IL-1{beta} secretion in an HIV-icRNA-dependent manner. Immunoprecipitation analysis revealed interaction of HIV-1 icRNA, but not multiply-spliced HIV-1 RNA, with NLRP1, suggesting that HIV-1 icRNA sensing by NLRP1 is sufficient to trigger inflammasome activation. Together, these findings reveal a pathway of NLRP1 inflammasome activation induced by de novo expressed HIV icRNA in HIV- infected myeloid cells.

microbiology↗

bioRxiv2024

Macrophage-intrinsic MDA5-IRF5 axis drives HIV-1 icRNA-induced inflammatory responses

Despite effective antiretroviral therapy (ART), transcriptionally competent HIV-1 reservoirs persist and contribute to persistent immune activation in people living with HIV (PWH). HIV-1-infected macrophages are important mediators of chronic innate immune activation, though mechanisms remain unclear. We previously reported that nuclear export and cytoplasmic expression of HIV-1 intron-containing RNA (icRNA) activates mitochondrial antiviral signaling protein (MAVS)-mediated type I interferon (IFN) responses in macrophages. In this study, we demonstrate an essential role of melanoma differentiation-associated protein 5 (MDA5) in sensing HIV-1 icRNA and promoting MAVS-dependent IRF5 activation in macrophages. Suppression of MDA5, but not RIG-I expression nor disruption of endosomal TLR pathway, abrogated HIV-1 icRNA-induced type I IFN responses and IP-10 expression in macrophages. Furthermore, induction of IP-10 in macrophages upon HIV-1 icRNA sensing by MDA5 was uniquely dependent on IRF5. Additionally, monocytes and MDMs from older (>50 years) individuals exhibit constitutively higher levels of IRF5 expression compared to younger (<35 years) individuals, and HIV-1 icRNA induced IP-10 expression was significantly enhanced in older macrophages, which was attenuated upon ablation of IRF5 expression suggesting that IRF5 functions as a major mediator of pro-inflammatory response downstream of MDA5-dependent HIV-1 icRNA sensing, dysregulation of which might contribute to chronic inflammation in older PWH.

microbiology↗

bioRxiv2023

Incorporation of 5 methylcytidine alleviates innate immune response to self-amplifying RNA vaccine

In order to improve vaccine effectiveness and safety profile of existing synthetic RNA-based vaccines, we have developed a self-amplifying RNA (saRNA)-based vaccine expressing membrane-anchored receptor binding domain (RBD) of SARS-CoV-2 S protein (S-RBD) and have demonstrated that a minimal dose of this saRNA vaccine elicits robust immune responses. Results from a recent clinical trial with 5-methylcytidine (5mC) incorporating saRNA vaccine demonstrated reduced vaccine-induced adverse effects while maintaining robust humoral responses. In this study, we investigate the mechanisms accounting for induction of efficient innate and adaptive immune responses and attenuated adverse effects induced by the 5mC-incorporated saRNA. We show that the 5mC-incorporating saRNA platform leads to prolonged and robust expression of antigen, while induction of type-I interferon (IFN-I), a key driver of reactogenicity, is attenuated in peripheral blood mononuclear cells (PBMCs), but not in macrophages and dendritic cells. Interestingly, we find that the major cellular source of IFN-I production in PBMCs is plasmacytoid dendritic cells (pDCs), which is attenuated upon 5mC incorporation in saRNA. In addition, we demonstrate that monocytes also play an important role in amplifying proinflammatory responses. Furthermore, we show that the detection of saRNA is mediated by a host cytosolic RNA sensor, RIG-I. Importantly, 5mC-incorporating saRNA vaccine candidate produced robust IgG responses against S-RBD upon injection in mice, thus providing strong support for the potential clinical use of 5mC-incorporating saRNA vaccines.

immunology↗

bioRxiv2022

Cardiac neural crest lineage diversity and underlying gene regulatory networks revealed by multimodal analysis

Neural crest cells (NCCs), a multipotent stem cell population, contribute to cardiac development as a source of the outflow septum, vascular smooth muscle and semilunar valves. However, genetic programs underlying lineage diversification of cardiac NCCs remain largely unknown. Using single-cell (sc) and spatial transcriptomics, we demonstrate multiple NCC subpopulations with distinct gene expression signatures; smooth muscle(-like), non-muscle mesenchymal, and Schwann cell progenitor/melanoblast-like cells. Integrative scRNA-seq and scATAC-seq analyses predict lineage trajectories starting from immature NCCs, which bifurcate into smooth muscle(-like) and non-muscle mesenchymal lineages in association with hierarchical transcription factor networks. Combinatory analyses with Cre-mediated genetic lineage tracing characterize intermediate NCCs at the bifurcation as Sox9+/Scx+ tendon and cartilage progenitor-like cells with genetic programs, some of which are common to skeletal tissues whereas others are unique to cardiac NCCs. These findings provide a basis for understanding the roles of NCCs in cardiac development and pathogenesis particularly associated with calcification.

developmental biology↗

bioRxiv2022

CD169-mediated restrictive SARS-CoV-2 infection of macrophages induces pro-inflammatory responses

Exacerbated and persistent innate immune response marked by pro-inflammatory cytokine expression is thought to be a major driver of chronic COVID-19 pathology. Although macrophages are not the primary target cells of SARS-CoV-2 infection in humans, viral RNA and antigens in activated monocytes and macrophages have been detected in post-mortem samples, and dysfunctional monocytes and macrophages have been hypothesized to contribute to a protracted hyper-inflammatory state in COVID-19 patients. In this study, we demonstrate that CD169, a myeloid cell specific I-type lectin, facilitated ACE2-independent SARS-CoV-2 fusion and entry in macrophages. CD169- mediated SARS-CoV-2 entry in macrophages resulted in expression of viral genomic and sub-genomic (sg) RNAs with minimal viral protein expression and no infectious viral particle release, suggesting a post-entry restriction of the SARS-CoV-2 replication cycle. Intriguingly this post-entry replication block was alleviated by exogenous ACE2 expression in macrophages. Restricted expression of viral gRNA and sgRNA in CD169+ macrophages elicited a pro-inflammatory cytokine expression (TNF, IL-6 and IL-1{beta}) in a RIG-I, MDA-5 and MAVS-dependent manner, which was suppressed by remdesivir pre- treatment. These findings suggest that de novo expression of SARS-CoV-2 RNA in macrophages contributes to the pro-inflammatory cytokine signature and that blocking CD169-mediated ACE2 independent infection and subsequent activation of macrophages by viral RNA might alleviate COVID-19-associated hyperinflammatory response. Author SummaryOver-exuberant production of pro-inflammatory cytokine expression by macrophages has been hypothesized to contribute to severity of COVID-19 disease. Molecular mechanisms that contribute to macrophage-intrinsic immune activation during SARS- CoV-2 infection are not fully understood. Here we show that CD169, a macrophage- specific sialic-acid binding lectin, facilitates abortive SARS-CoV-2 infection of macrophages that results in innate immune sensing of viral replication intermediates and production of proinflammatory responses. We identify an ACE2-independent, CD169- mediated endosomal viral entry mechanism that results in cytoplasmic delivery of viral capsids and initiation of virus replication, but absence of infectious viral production. Restricted viral replication in CD169+ macrophages and detection of viral genomic and sub-genomic RNAs by cytoplasmic RIG-I-like receptor family members, RIG-I and MDA5, and initiation of downstream signaling via the adaptor protein MAVS, was required for innate immune activation. These studies uncover mechanisms important for initiation of innate immune sensing of SARS-CoV-2 infection in macrophages, persistent activation of which might contribute to severe COVID-19 pathophysiology.

immunology↗

bioRxiv2021

C-type natriuretic peptide facilitates autonomic Ca2+ entry in growth plate chondrocytes for stimulating bone growth

The growth plates are cartilage tissues found at both ends of developing bones, and vital proliferation and differentiation of growth plate chondrocytes are primarily responsible for bone growth. C-type natriuretic peptide (CNP) stimulates bone growth by activating natriuretic peptide receptor 2 (NPR2) which is equipped with guanylate cyclase on the cytoplasmic side, but its signaling pathway is unclear in growth plate chondrocytes. We previously reported that transient receptor potential melastatin-like 7 (TRPM7) channels mediate intermissive Ca2+ influx in growth plate chondrocytes, leading to activation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) for promoting bone growth. In this report, we provide experimental evidence indicating a functional link between CNP and TRPM7 channels. Our pharmacological data suggest that CNP-evoked NPR2 activation elevates cellular cGMP content and stimulates big-conductance Ca2+-dependent K+ (BK) channels as a substrate for cGMP-dependent protein kinase (PKG). BK channel-induced hyperpolarization likely enhances the driving force of TRPM7-mediated Ca2+ entry and seems to accordingly activate CaMKII. Indeed, ex vivo organ culture analysis indicates that CNP-facilitated bone growth is abolished by chondrocyte-specific Trpm7 gene ablation. The defined CNP signaling pathway, the NPR2-PKG-BK channel-TRPM7 channel-CaMKII axis, likely pinpoints promising target proteins for developing new therapeutic treatments for divergent growth disorders.

cell biology↗

bioRxiv2021

Drp1 SUMO/deSUMOylation by Senp5 isoforms influences ER tubulation and mitochondrial dynamics to regulate brain development

Brain development is a highly orchestrated process requiring spatiotemporally regulated mitochondrial dynamics. Drp1, a key molecule in the mitochondrial fission machinery, undergoes various post-translational modifications including conjugation to the small ubiquitin-like modifier (SUMO). However, the functional significance of SUMOylation/deSUMOylation on Drp1 remains controversial. SUMO-specific protease 5 (Senp5L) catalyzes the deSUMOylation of Drp1. We revealed that a splicing variant of Senp5L, Senp5S, which lacks peptidase activity, prevents deSUMOylation of Drp1 by competing against other Senps. The altered SUMOylation level of Drp1 induced by Senp5L/5S affects Drp1 ubiquitination and tubulation of the endoplasmic reticulum (ER), thereby influencing mitochondrial morphology. A dynamic SUMOylation/deSUMOylation balance controls neuronal polarization and migration during the development of the cerebral cortex. These findings suggest a novel role of post translational modification, in which a deSUMOylation enzyme isoform competitively regulates mitochondrial dynamics and ER tubulation via Drp1 SUMOylation levels in a tightly controlled process of neuronal differentiation and corticogenesis.

neuroscience↗

bioRxiv2020

Lung lineage transcription factor NKX2-1 epigenetically resolves opposing cell fates in vivo

Differential use of identical DNA sequences leads to distinct tissue lineages and then multiple cell types within a lineage, an epigenetic process central to progenitor and stem cell biology. The associated genomic changes, especially in native tissues, remain insufficiently understood, and are hereby addressed in the mouse lung, where the same lineage transcription factor NKX2-1 promotes the diametrically opposed alveolar type 1 (AT1) and AT2 cell fates. We show that the cell-type-specific function of NKX2-1 is attributed to its differential chromatin binding that is acquired or retained during development in coordination with partner transcriptional factors. Loss of YAP/TAZ redirects NKX2-1 from its AT1-specific to AT2-specific binding sites, leading to transcriptionally exaggerated AT2 cells when deleted in progenitors or AT1-to-AT2 conversion when deleted after fate commitment. Nkx2-1 mutant AT1 and AT2 cells gain distinct accessible sites including those of the opposite fate while adopting the gastrointestinal fate, suggesting an epigenetic plasticity larger than a transcriptional one. Our genomic analysis of single or purified cells, coupled with precision genetics, provides an epigenetic roadmap of alveolar cell fate and potential, and introduces an experimental benchmark for unraveling the in vivo function of lineage transcription factors.

developmental biology↗