Xiaoqing Zhang

Organ transplantation is the last-resort option to treat organ failure. However, less than 10% of patients benefit from this only option due to lack of major histocompatibility complex (MHC)-matched donor organs and 25%–80% of donated organs could not find MHC-matched recipients. T cell allorecognition is the principal mechanism for allogeneic graf...

Organ transplantation is the last-resort option to treat organ failure. However, less than 10% of patients benefit from this only option due to lack of major histocompatibility complex (MHC)-matched donor organs and 25-80% of donated organs could not find MHC-matched recipients. T cell allorecognition is the principal mechanism for allogeneic graft...

Hypoimmunogenic human pluripotent stem cells (hPSCs) are expected to serve as an unlimited cell source for generating universally compatible “off-the-shelf” cell grafts. However, whether the engineered hypoimmunogenic hPSCs still preserve their advantages of unlimited self-renewal and multilineage differentiation to yield functional tissue cells re...

Cellular diversification is critical for specialized functions of the brain including learning and memory1. Single-cell RNA sequencing facilitates transcriptomic profiling of distinct major types of neuron2–4, but the divergence of transcriptomic profiles within a neuronal population and their link to function remain poorly understood. Here we isol...

Hypoimmunogenic human pluripotent stem cells (hPSCs) are expected to serve as an unlimited cell source for generating universally compatible “off-the-shelf” cell grafts. However, whether the engineered hypoimmunogenic hPSCs still preserve their advantages of unlimited self-renewal and multilineage differentiation to yield functional tissue cells re...

The dorsal and ventral human telencephalons contain different neuronal subtypes, including glutamatergic, GABAergic, and cholinergic neurons, and how these neurons are generated during early development is not well understood. Using scRNA-seq and stringent validations, we reveal here a developmental roadmap for human telencephalic neurons. Both dor...

Congenital hydrocephalus is a major neurological disorder with high rates of morbidity and mortality; however, the underlying cellular and molecular mechanisms remain largely unknown. Reproducible animal models mirroring both embryonic and postnatal hydrocephalus are also limited. Here, we describe a new mouse model of congenital hydrocephalus thro...

PAX6 is a key determinant of human neuroectoderm cell fate. Here, we describe a protocol for genome-scale CRISPR screening for use in genetically engineered human pluripotent stem cells (hPSCs). Using the germ layer reporter PAX6 and an inducible CRISPR/Cas9 knockout system, we describe how to identify lineage-specific preventing genes. This protoc...

Understanding the biological processes that determine three germ layer-entry of human pluripotent stem cells (hPSCs) is a central question in developmental and stem cell biology. Here, we genetically engineered hPSCs with the germ layer reporter and inducible CRISPR/Cas9 knockout system, and a genome-scale screening was performed to define pathways...

Allogeneic immune rejection is a major barrier for the application of human pluripotent stem cells (hPSCs) in regenerative medicine. A broad spectrum of immune cells, including T cells, natural killer (NK) cells, and antigen-presenting cells, which either cause direct cell killing or constitute an immunogenic environment, are involved in allograft...

Medial ganglionic eminence (MGE)-like cells yielded from human pluripotent stem cells (hPSCs)hold great potentials for cell therapies of related neurological disorders. However, cues that orchestrate the maintenance versus differentiation of human MGE progenitors, and ways for large-scale expansion of these cells have not been investigated. Here, w...

It remains largely unknown how Zika virus (ZIKV) infection causes severe microcephaly in human newborns. We examined an Asian lineage ZIKV, SZ01, which similarly infected and demonstrated comparable growth arrest and apoptotic pathological changes in human neuroprogenitors (NPCs) from forebrain dorsal, forebrain ventral as well as hindbrain and spi...

Spinal muscular atrophy (SMA), the leading genetic cause of infant mortality, is caused by reduced levels of functional survival motor neuron (SMN) protein. To identify therapeutic agents for SMA, we established a versatile SMN2-GFP reporter line by targeting the human SMN2 gene. We then screened a compound library and identified Z-FA-FMK as a pote...

It is highly desirable to specify human developmental principles in an appropriate human model with advanced genetic tools. However, genetically engineering human cells with lineage-tracing systems has not been achieved. Here we introduce strategies to construct lineage-tracing systems in human embryonic stem cells (hESCs). The AAVS1 locus was suit...

Most of the brain tumors are malignant with an extremely poor survival rate. Recent progress in identifying cancer stem cells (CSCs) within the brain tumors is starting to revolutionize our understanding in the imitation and progression of tumors as well as relapse and the development of therapeutic strategies. Suspension spheroid body culture para...

Stem cell-based therapies have been used for repairing damaged brain tissue and helping functional recovery after brain injury. Aberrance neurogenesis is related with brain injury, and multipotential neural stem cells from human embryonic stem (hES) cells provide a great promise for cell replacement therapies. Optimized protocols for neural differe...

Smad5 accumulates in cytoplasm at low temperature.

Transcriptome analysis of WT and Smad5 KO hES.

Temperature, pHe and osmolarity fluctuations can induce pHi changes and Smad5 MH1 domain is responsible for pHi sensing.

Chronic metabolic adaptation of Smad5 KO hES.

Oxidative stress, membrane potential and signaling modifiers could not obviously affect Smad5, Smad1 or Smad8 subcellular distribution under normal or low temperature culture conditions.

Smad5 responses to pHe and osmolarity fluctuations.

Transcriptome analysis of WT and LDN treated hES.

Smad5 KO reduces glycolysis.

Smad5 subjects to accelerated CRM1-dependent nuclear export at low temperature and three NESs are involved in its rapid nuclear export.

Environmental cues triggered Smad5 translocation is BMP signaling-independent.

Timelapse video recording of nuclear export of GFP-Smad5 at basic extracellular pH. GFP-Smad5 HEK293 cells were cultured at 37°C, then the cells were placed at basic extracellular pH conditions to record the video.

Smad5 shuttling could not regulate metabolism-related gene expression.

Smad5 KO induces irreversible mitochondrial morphology changes.

Timelapse video recording of nuclear export of GFP-Smad5 at 25°C. GFP-Smad5 HEK293 cells were cultured at 37°C, then the cells were placed at 25°C to record the video.

Smad5 KO impaired mitochondrial respiration.

Increased pHi during neural differentiation.

Smad5 interacts with HK1.

Smad5 (cytosolic) pull down proteins

Timelapse video recording of nuclear export of GFP-Smad5 at hypertonic conditions. GFP-Smad5 HEK293 cells were cultured at 37°C, then the cells were placed at hypertonic conditions to record the video.

Both environmental cues and intracellular bioenergetic states profoundly affect intracellular pH (pHi). How a cell responds to pHi changes to maintain bioenergetic homeostasis remains elusive. Here we show that Smad5, a well-characterized downstream component of bone morphogenetic protein (BMP) signaling responds to pHi changes. Cold, basic or hype...

Neuroectoderm is an important neural precursor. However, chromatin remodeling and its epigenetic regulatory roles during the differentiation of human neuroectodermal cells (hNECs) from human embryonic stem cells (hESCs) remain largely unexplored. Here, we obtained hNECs through directed differentiation from hESCs, and determined chromatin states in...

Environmental stresses are increasingly acknowledged as core causes of abnormal neural induction leading to neural tube defects (NTDs). However, the mechanism responsible for environmental stress-triggered neural induction defects remains unknown. Here, we report that a spectrum of environmental stresses, including oxidative stress, starvation, and...

Embryoid body (EB) formation and adherent culture (AD) paradigms are equivalently thought to be applicable for neural specification of human pluripotent stem cells. Here, we report that sonic hedgehog-induced ventral neuroprogenitors under EB conditions are fated to medial ganglionic eminence (MGE), while the AD cells mostly adopt a floor-plate (FP...

Loss-of-function studies in human pluripotent stem cells (hPSCs) require efficient methodologies for lesion of genes of interest. Here, we introduce a donor-free paired gRNA-guided CRISPR/Cas9 knockout strategy (paired-KO) for efficient and rapid gene ablation in hPSCs. Through paired-KO, we succeeded in targeting all genes of interest with high bi...

The anteroposterior patterning of the central nervous system follows an activation/transformation model, which proposes that a prospective telencephalic fate will be activated by default during the neural induction stage, while this anterior fate could be transformed posteriorly according to caudalization morphogens. Although both extrinsic signals...

The transforming growth factor beta (TGFβ) related signaling is one of the most important signaling pathways regulating early developmental events. Smad2 and Smad3 are structurally similar and it is mostly considered that they are equally important in mediating TGFβ signals. Here, we show that Smad3 is an insensitive TGFβ transducer as compared wit...

We cloned a new splicing variant of Otx2 gene, Otx2c. Otx2c lacks entire exon 4, most of the region encoding the homeodomain. More importantly, Otx2c harbors an early premature stop codon and bioinformatics analysis prefers it to be a non-protein coding RNA. In addition, this splicing variant is not simply a noise during mRNA processing, since it i...