Biomedical subjects
Publications and source records attributed to Pengcheng Wang.
Systematic comparison of Cas12i gene-editing tools in tomato identifies Cas-SF01 as the most efficient variant, achieving editing efficiencies comparable to conventional Cas9 at some genomic sites and providing a compact, highly specific, and versatile tool for studying gene function, engineering fruit metabolism, and developing improved tomato varieties through precision breeding.
The emergence of SARS-CoV-2 has posed significant threats to global health, particularly for the older population. Similarly, common human coronaviruses, such as HCoV-229E, which typically cause mild cold-like symptoms, can lead to severe diseases, underscoring the need to understand virus-host interactions and identify host factors contributing to viral pathogenesis and disease progression. In this study, we perform a genome-wide CRISPR knockout screen using HCoV-229E and identify UHRF1 as a potent restriction factor. Mechanistically, UHRF1 suppresses HCoV-229E infection by downregulating the expression of its cell entry receptor, APN, through promoter hypermethylation. Focused CRISPR activation screens of UHRF1-downregulated genes confirm the critical role of APN in HCoV-229E infection and identify additional genes (e.g., SIGLEC1, PLAC8, and heparan sulfate biosynthesis genes) contributing to the restrictive functions of UHRF1. Transcriptomic and single-cell RNA sequencing analysis reveal that UHRF1 expression decreases with age, negatively correlating with increased APN expression. This age-related decline in UHRF1 is validated in primary alveolar macrophages from elderly individuals, which exhibit heightened susceptibility to HCoV-229E compared to those from younger individuals. Our findings highlight UHRF1 as a key age-related host defense factor against coronavirus and provide insights into the epigenetic regulation of viral entry receptors.
Epigenetic editing is a promising strategy for modifying gene expression while avoiding the permanent alterations and potential genotoxicity of genome-editing technologies. Here we designed optimized epigenetic regulators (EpiRegs) by testing combinations of transcription activator-like effector (TALE)-based and catalytically deactivated Cas9 (dCas9)-based epigenetic modification effectors and fusion protein structures. TALE-based EpiReg (EpiReg-T) achieved a final efficiency of 98% in mice, surpassing the initial dCas9-based efficiency of 64%. We demonstrated the approach in macaques by introducing DNA methylation and histone modifications to inhibit proprotein convertase subtilisin/kexin type 9 (PCSK9) expression, thereby lowering low-density lipoprotein cholesterol levels. A single dose of EpiReg-T delivered with lipid nanoparticles achieved efficient (>90%) and long-lasting (343 days) silencing of PCSK9 in the liver. Integrative multiomic analyses revealed minimal off-target effects in EpiReg-T-treated monkeys, mice and human-derived cells. EpiReg can be redirected to other genes by reengineering the DNA-binding domain. Our findings represent a step toward the clinical application of epigenetic editing for the treatment of human diseases.
We isolated two T-DNA insertion mutants of Arabidopsis thaliana GLUTATHIONE PEROXIDASE3 (ATGPX3) that exhibited a higher rate of water loss under drought stress, higher sensitivity to H(2)O(2) treatment during seed germination and seedling development, and enhanced production of H(2)O(2) in guard cells. By contrast, lines engineered to overexpress ATGPX3 were less sensitive to drought stress than the wild type and displayed less transpirational water loss, which resulted in higher leaf surface temperature. The atgpx3 mutation also disrupted abscisic acid (ABA) activation of calcium channels and the expression of ABA- and stress-responsive genes. ATGPX3 physically interacted with the 2C-type protein phosphatase ABA INSENSITIVE2 (ABI2) and, to a lesser extent, with ABI1. In addition, the redox states of both ATGPX3 and ABI2 were found to be regulated by H(2)O(2). The phosphatase activity of ABI2, measured in vitro, was reduced approximately fivefold by the addition of oxidized ATGPX3. The reduced form of ABI2 was converted to the oxidized form by the addition of oxidized ATGPX3 in vitro, which might mediate ABA and oxidative signaling. These results suggest that ATGPX3 might play dual and distinctive roles in H(2)O(2) homeostasis, acting as a general scavenger and specifically relaying the H(2)O(2) signal as an oxidative signal transducer in ABA and drought stress signaling.
Alveolar epithelial type II cells secrete lung surfactant via exocytosis. Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) are implicated in this process. Lipid rafts, the cholesterol- and sphingolipid-rich microdomains, may offer a platform for protein organization on the cell membrane. We tested the hypothesis that lipid rafts organize exocytotic proteins in type II cells and are essential for the fusion of lamellar bodies, the secretory granules of type II cells, with the plasma membrane. The lipid rafts, isolated from type II cells using 1% Triton X-100 and a sucrose gradient centrifugation, contained the lipid raft markers, flotillin-1 and -2, whereas they excluded the nonraft marker, Na+-K+ ATPase. SNAP-23, syntaxin 2, and VAMP-2 were enriched in lipid rafts. When type II cells were depleted of cholesterol, the association of SNAREs with the lipid rafts was disrupted and the formation of fusion pore was inhibited. Furthermore, the cholesterol-depleted plasma membrane had less ability to fuse with lamellar bodies, a process mediated by annexin A2. The secretagogue-stimulated secretion of lung surfactant from type II cells was also reduced by methyl-beta-cyclodextrin. When the raft-associated cell surface protein, CD44, was cross-linked using anti-CD44 antibodies, the CD44 clusters were observed. Syntaxin 2, SNAP-23, and annexin A2 co-localized with the CD44 clusters, which were cholesterol dependent. Our results suggested that lipid rafts may form a functional platform for surfactant secretion in alveolar type II cells, and raft integrity was essential for the fusion between lamellar bodies with the plasma membrane.
OBJECTIVE: To find out an ideal method of internal fixation for the treatment of fractures of proximal radius. METHODS: From August 2001 to April 2004, 24 patients suffering from fractures of proximal radius were treated with Ni-Ti shape memory alloy embracing fixator. Of 24 patients, there were 19 males and 5 females, aging from 16 to 48 years; there were 9 cases of wrestling injury, 5 cases of crush injury, 4 cases of traffic accident injury, 3 cases of direct violence impact injury and 3 cases of high falling injury. All patients were given operation according to fix method of the shape memory alloy embracing fixator. All cases were regularly followed up to observe the fracture healing and functional recovery. RESULTS: All cases were followed up postoperatively from 4 months to 21 months with an average of 10 months. No complication occurred during operation and after operation. Healing by first intention was achieved. X-ray showed that all cases achieved clinical union 8-12 weeks after operation. According to Anderson's criteria and the results were excellent in 18 cases, good in 5 cases, and fair in 1 case. The excellent and good rate was 95.83%. CONCLUSION: The internal fixation with shape memory alloy embracing fixator has as follows advantages: less trauma, esay-to-do, stable fixation, better tissue compatibility and less complication in treating fractures of proximal radius. This method can enhance the fracture healing and functional recovery. So it is a better method to treat fractures of proximal radius.
Aimed at the complex community structure and higher fragmentation of urban green space, and based on the investigation of synusia structure and its coverage, this paper studied the vegetation quantity of ornamental green space in the workshop area of Wuhan Iron and Steel Company, with the help of GIS. The results showed that different life forms of ornamental plants in this area had a greater difference in their single leaf area and leaf area index (LAI), and the LAI was not only depended on single leaf area, but also governed by the shape of tree crown and the intensive degree of branches and leaves. The total vegetation quantity was 1 694.2 hm2, with the average LAI being 7.75, and the vegetation quantity of arbor-shrub-herb and arbor-shrub communities accounted for 79.7% and 92.3% of the total, respectively, reflecting that the green space structure was dominated by arbor species and by arbor-shrub-herb and arbor-shrub community types. Single layer-structured lawn had a less percentage, while the vegetation quantity of herb synusia accounted for 22.9% of the total, suggesting an afforestation characteristic of "making use of every bit of space" in the workshop area. The vegetation quantity of urban ornamental green space depended on the area of green space, its synusia structure, and the LAI and coverage of ornamental plants. In enlarging urban green space, ornamental plant species with high LAI should be selected, and community structure should be improved to have a higher vegetation quantity in urban area. To quantify the vegetation quantity of urban ornamental green space more accurately, synusia should be taken as the unit to measure the LAI of typical species, and the synusia structure and its coverage of different community types should be investigated with the help of remote sensing images and GIS.
The phytohormone abscisic acid (ABA) modulates the expression of many genes important to plant growth and development and to stress adaptation. In this study, we found that an APETALA2/EREBP-type transcription factor, AtERF7, plays an important role in ABA responses. AtERF7 interacts with the protein kinase PKS3, which has been shown to be a global regulator of ABA responses. AtERF7 binds to the GCC box and acts as a repressor of gene transcription. AtERF7 interacts with the Arabidopsis thaliana homolog of a human global corepressor of transcription, AtSin3, which in turn may interact with HDA19, a histone deacetylase. The transcriptional repression activity of AtERF7 is enhanced by HDA19 and AtSin3. Arabidopsis plants overexpressing AtERF7 show reduced sensitivity of guard cells to ABA and increased transpirational water loss. By contrast, AtERF7 and AtSin3 RNA interference lines show increased sensitivity to ABA during germination. Together, our results suggest that AtERF7 plays an important role in ABA responses and may be part of a transcriptional repressor complex and be regulated by PKS3.
RNA interference (RNAi) is a sequence-specific post-transcriptional gene silencing process. Although it is widely used in the loss-of-function studies, none of the current RNAi technologies can achieve cell-specific gene silencing. The lack of cell specificity limits its usage in vivo. Here, we report a cell-specific RNAi system using an alveolar epithelial type II cell-specific promoter--the surfactant protein C (SP-C) promoter. We show that the SP-C-driven small hairpin RNAs specifically depress the expression of the exogenous reporter (enhanced green fluorescent protein) and endogenous genes (lamin A/C and annexin A2) in alveolar type II cells, but not other lung cells, using cell and organ culture in vitro as well as in vivo. The present study provides an efficient strategy in silencing a gene in one type of cell without interfering with other cell systems, and may have a significant impact on RNAi therapy.
The secretion of lung surfactant in alveolar type II cells is a complex process involving the fusion of lamellar bodies with the plasma membrane. This process is somewhat different from the exocytosis of hormones and neurotransmitters. For example, it is a relatively slower process, and lamellar bodies are very large vesicles with a diameter of approximately 1 microm. SNARE proteins are the conserved molecular machinery of exocytosis in the majority of secretory cells. However, their involvement in surfactant secretion has not been reported. Here, we showed that syntaxin 2 and SNAP-23 are expressed in alveolar type II cells. Both proteins are associated with the plasma membrane, and to some degree with lamellar bodies. An antisense oligonucleotide complementary to syntaxin 2 decreased its mRNA and protein levels. The same oligonucleotide also inhibited surfactant secretion, independent of secretagogues. A peptide derived from the N-terminus of syntaxin 2 or the C-terminus of SNAP-23 significantly inhibited Ca(2+)- and GTPgammaS-stimulated surfactant secretion from permeabilized type II cells in a dose-dependent manner. Furthermore, introduction of anti-syntaxin 2 or anti-SNAP-23 antibodies into permeabilized type II cells also inhibited surfactant release. Our results suggest that syntaxin 2 and SNAP-23 are required for regulated surfactant secretion.
In this paper, four types of landscape structures and their green-land landscape patterns, including the landscapes of dominant green-land patch pattern, even green-land patch pattern, dominant wooded corridor pattern and building or concrete covering pattern (control) in the central district of Yichang city in Hubei Province, were analyzed respectively on the basis of landscape ecological theory. The atmospheric noise and the contents of SO2, NOx and total suspension particle (TSP) of the landscapes were monitored respectively by comparative method. The results showed that the landscape of building or concrete covering pattern (control) was mostly composed of woodless corridors and building or concrete covering patches with the tiptop green-land fragmentation index (18.125 3 ind x hm(-2)) and only had 1.00% green-land coverage, which had a relatively higher atmospheric noise and the highest TSP content in the landscape. The landscape of dominant green-land patch pattern had the highest green-land coverage (up to 43.59%) dominated by great green-land patches and the least fragmentation index (0.453 9 ind x hm(-2)), in which, the atmospheric noise weakened by 28.12% and the TSP content reduced significantly by 86.42%, comparing to the control. The landscape of even green-land patch pattern had the relatively lower green-land coverage (11.34%) and fragmentation index (2.751 1 ind x hm(-2)), which was mainly composed of the middle or small green-land patches and wooded corridors with a regular distribution. In the landscape, the TSP content reduced obviously by 46.62% of the control, while the effect of dust retention was only 53.95% of that in the landscape of dominant green-land patch pattern. In the landscape of dominant wooded corridor pattern, which was a traffic center and turned into a main pollution resource in the city, there were a relatively higher green-land fragmentation index (6.870 0 ind x hm(-2)) and the highest wooded corridor density (0.844 3 hm x hm(-2)) with only 6.13% green-land coverage, and the atmospheric noise and the TSP and NOx content increased by 21.47%, 5.08% and 9.06%, respectively, comparing to control. It was obvious that the greater the average area of the green-land patch and the lower the fragmentation index of green-land patches, the more effective the green-land on purifying air pollution.
Annexin II has been implicated in membrane fusion during the exocytosis of lamellar bodies from alveolar epithelial type II cells. Most previous studies were based on the fusion assays by using model membranes. In the present study, we investigated annexin II-mediated membrane fusion by using isolated lamellar bodies and plasma membrane as determined by the relief of octadecyl rhodamine B (R18) self-quenching. Immunodepletion of annexin II from type II cell cytosol reduced its fusion activity. Purified annexin II tetramer (AIIt) induced the fusion of lamellar bodies with the plasma membrane in a dose-dependent manner. This fusion is Ca2+-dependent and is highly specific to AIIt because other annexins (I and II monomer, III, IV, V, and VI) were unable to induce the fusion. Modification of the different functional residues of AIIt by N-ethylmaleimide, nitric oxide, or peroxynitrite abolished AIIt-mediated fusion. Arachidonic acid enhanced AIIt-mediated fusion and reduced its Ca2+ requirement to an intracellularly achievable level. This effect is due to membrane-bound arachidonic acid, not free arachidonic acid. Other fatty acids including linolenic acid, palmitoleic acid, myristoleic acid, stearic acid, palmitic acid, and myristic acid had little effect. AIIt-mediated fusion was suppressed by the removal of arachidonic acid from lamellar body and plasma membrane using bovine serum albumin. The addition of arachidonic acid back to the arachidonic acid-depleted membranes restored its fusion activity. Our results suggest that the fusion between lamellar bodies with the plasma membrane is driven by the synergistic action of AIIt and arachidonic acid.
N-ethylmaleimide-sensitive fusion protein (NSF) and soluble NSF attachment protein (alpha-SNAP) are thought to be soluble factors that transiently bind and disassemble SNAP receptor complex during exocytosis in neuronal and endocrine cells. Lung surfactant is secreted via exocytosis of lamellar bodies from alveolar epithelial type II cells. However, the secretion of lung surfactant is a relatively slow process, and involvement of SNAP receptor and its cofactors (NSF and alpha-SNAP) in this process has not been demonstrated. In this study, we investigated a possible role of alpha-SNAP in surfactant secretion. alpha-SNAP was predominantly associated with the membranes in alveolar type II cells as determined by Western blot and immunocytochemical analysis using confocal microscope. Membrane-associated alpha-SNAP was not released from the membrane fraction when the cells were lyzed in the presence of Ca2+ or Mg2+ATP. The alkaline condition (0.1 M Na2CO3, pH 12), known to extract peripheral membrane proteins also failed to release it from the membrane. Phase separation using Triton X-114 showed that alpha-SNAP partitioned into both aqueous and detergent phases. NSF had membrane-bound characteristics similar to alpha-SNAP in type II cells. Permeabilization of type II cells with beta-escin resulted in a partial loss of alpha-SNAP from the cells, but cellular NSF was relatively unchanged. Addition of exogenous alpha-SNAP to the permeabilized cells increased surfactant secretion in a dose-dependent manner, whereas exogenous NSF has much less effects. An alpha-SNAP antisense oligonucleotide decreased its protein level and inhibited surfactant secretion. Our results suggest a role of alpha-SNAP in lung surfactant secretion.