Chengqi Lin

The underlying developmental mechanisms of defects in human organogenesis remain poorly defined. Pathogenic variants in AFF3, the super elongation complex subunit, cause a congenital disorder of neural and limb development, yet their developmental basis is unclear. Stem cell-based embryo model gastruloids show that AFF3, enriched in neural progenitors and neuromesodermal progenitors, is required for female neural and musculoskeletal development. AFF3 loss depletes neural and musculoskeletal lineages. Mechanistically, AFF3 restrains premature and excessive activation of the X-inactivation mediator XIST. Abnormal XIST upregulation represses autosomal genes required for neural and musculoskeletal development. Genetic deletion or pharmacological attenuation of XIST restores gastruloid morphogenesis, rescues neural and musculoskeletal development via autosomal genes, and improves cerebral organoid formation and neurogenesis after transplantation. These findings identify AFF3-mediated XIST restraint as an essential safeguard for early human organogenesis, and its failure can be traced back to gastrulation.

Summary The mammalian heart is formed from multiple mesoderm-derived cell lineages. However, it remains largely unknown when and how the specification of mesoderm towards cardiac lineages is determined. Here, we systematically depict the transcriptional trajectories toward cardiomyocyte in early mouse embryo, and characterize the epigenetic landscapes underlying the early mesodermal lineage specification by single-cell multi-omics analyses. The analyses also reveal distinct core regulatory networks (CRN) in controlling specification of mesodermal lineages. We further demonstrate the essential role HAND1 and FOXF1 in driving the earliest cardiac progenitors specification. These key transcription factors occupy at distinct enhancers, but function synergistically and hierarchically to regulate the expression of cardiac-specific genes. In addition, HAND1 is required for exiting from the nascent mesoderm program, while FOXF1 is essential for driving cardiac differentiation during juxta-cardiac field (JCF) specification. Our findings establish transcriptional and epigenetic determinants specifying the early cardiac lineage, providing insights for the investigation of congenital heart defects.

RING finger protein 219 (RNF219) is a co‐factor for the CCR4‐NOT deadenylase complex in mammals. Here, we found that mutations within the C3HC4 scaffold of the RING finger domain in RNF219 are capable of forming condensates via liquid–liquid phase separation (LLPS), though the wild‐type RING finger domain intrinsically suppresses LLPS. We further demonstrated that the adjacent coiled‐coil 1 (CC1) domain promotes the potential of RNF219 to form condensates. Moreover, the mutant RNF219 condensates are able to encapsulate the CCR4‐NOT complex, inhibiting the RNA deadenylation activity of CCR4‐NOT. Additionally, we observed that RNF219 mutations could promote cell proliferation. These findings suggest a pathogenic mechanism whereby RNF219 mutations could induce CCR4‐NOT condensate formation, inhibit deadenylation‐dependent mRNA decay and drive cell proliferation.

Obtaining single-cell spatial information remains a challenge in spatial transcriptomics. Here we develop SEU-TCA, a method that leverages transfer component analysis to improve single-cell spatial mapping accuracy. Application to multiple single-cell and spatial transcriptomic datasets shows superior performance in spatial deconvolution and cell mapping. Using SEU-TCA, we explore spatial gene expression and regulon activity during mouse gastrulation and identify anterior second heart field progenitors regulated by Irx1. Functional experiments reveal that Irx1 deletion disrupts anterior second heart field development and causes ventricular septal defects, underscoring SEU-TCA’s potential for advancing developmental biology research.

Despite significant advances in structural and genetic studies, investigations of early embryonic functions, such as brain activity, have long been constrained by technical challenges. Functional ultrasound (fUS) has emerged as a breakthrough modality, enabling real-time monitoring of brain activity with exceptional spatial and temporal resolution and offering unprecedented opportunities for studying functional embryonic development. In this study, we used fUS to monitor whole-embryo activity in mice from embryonic days E8.5 to E18.5, revealing patterns of neural activity throughout embryogenesis. This approach provides new opportunities to explore brain development dynamically as it unfolds. Moreover, we observed embryo responses to external stimuli, including sound, in both mice and cynomolgus macaques, offering insights into early sensory processing and neural maturation. In summary, our study establishes fUS as a powerful tool for studying embryonic brain functional development, with significant implications for scientific research, especially in non-human primate models, and clinical applications.

SPT5 exhibits versatile functions in RNA Pol II promoter proximal pausing, pause release, and elongation in metazoans. However, the mechanism underlying the functional switch of SPT5 during early elongation has not been fully understood. Here, we report that the phosphorylation site-rich domain (PRD)/CTR1 and the prion-like domain (PLD)/CTR2, which are situated adjacent to each other within the C-terminal repeat (CTR) in SPT5, play pivotal roles in Pol II pausing and elongation, respectively. Our study demonstrates that LEDGF/p75 is highly enriched at promoters, especially paused promoters, and prevents the phosphorylation of SPT5 PRD by the super elongation complex (SEC). Furthermore, deletion of LEDGF IBD leads to increased SEC occupancies and SPT5 PRD phosphorylation at promoters and also increased pause release. In sum, our study reveals that LEDGF and SEC function cooperatively on SPT5 distinct domains to ensure proper transcriptional transition from pausing to elongation.

Up to an estimated 10% of women experience miscarriage in their lifetimes. Embryonic aneuploidy is a leading cause for miscarriage, infertility and congenital defects. Here we identify variants of ELL3, a gene encoding a transcription elongation factor, in couples who experienced consecutive early miscarriages due to embryonic aneuploidy. Maternal ELL3 knockout leads to mouse oocyte aneuploidy, subfertility and miscellaneous embryonic defects. Mechanistically, we find that ELL3 localizes to the spindle during meiosis, and that ELL3 depletion in both mouse and human oocytes increases the incidence of meiotic spindle abnormality. ELL3 coordinates with TPX2 to ensure the proper function of the microtubule motor KIF11. Live imaging analysis shows that ELL3 is paramount for promoting spindle assembly and driving chromosome movement. Together, our findings implicate maternal ELL3 deficiency in causing oocyte aneuploidy and early miscarriage.

Energy deprivation and metabolic rewiring of cardiomyocytes are widely recognized hallmarks of heart failure. Here, we report that HEY2 (a Hairy/Enhancer-of-split-related transcriptional repressor) is upregulated in hearts of patients with dilated cardiomyopathy. Induced Hey2 expression in zebrafish hearts or mammalian cardiomyocytes impairs mitochondrial respiration, accompanied by elevated ROS, resulting in cardiomyocyte apoptosis and heart failure. Conversely, Hey2 depletion in adult mouse hearts and zebrafish enhances the expression of mitochondrial oxidation genes and cardiac function. Multifaceted genome-wide analyses reveal that HEY2 enriches at the promoters of genes known to regulate metabolism (including Ppargc1, Esrra and Cpt1) and colocalizes with HDAC1 to effectuate histone deacetylation and transcriptional repression. Consequently, restoration of PPARGC1A/ESRRA in Hey2- overexpressing zebrafish hearts or human cardiomyocyte-like cells rescues deficits in mitochondrial bioenergetics. Knockdown of Hey2 in adult mouse hearts protects against doxorubicin-induced cardiac dysfunction. These studies reveal an evolutionarily conserved HEY2/HDAC1-Ppargc1/Cpt transcriptional module that controls energy metabolism to preserve cardiac function.

Recently, RNA velocity has driven a paradigmatic change in single-cell RNA sequencing (scRNA-seq) studies, allowing the reconstruction and prediction of directed trajectories in cell differentiation and state transitions. Most existing methods of dynamic modeling use ordinary differential equations (ODE) for individual genes without applying multivariate approaches. However, this modeling strategy inadequately captures the intrinsically stochastic nature of transcriptional dynamics governed by a cell-specific latent time across multiple genes, potentially leading to erroneous results. Here, we present SDEvelo, a generative approach to inferring RNA velocity by modeling the dynamics of unspliced and spliced RNAs via multivariate stochastic differential equations (SDE). Uniquely, SDEvelo explicitly models inherent uncertainty in transcriptional dynamics while estimating a cell-specific latent time across genes. Using both simulated and four scRNA-seq and spatial transcriptomics datasets, we show that SDEvelo can model the random dynamic patterns of mature-state cells while accurately detecting carcinogenesis. Additionally, the estimated gene-shared latent time can facilitate many downstream analyses for biological discovery. We demonstrate that SDEvelo is computationally scalable and applicable to both scRNA-seq and sequencing-based spatial transcriptomics data.