Hua Huang

Stemness-associated cell states are linked to chemotherapy resistance in acute myeloid leukemia (AML). We uncovered a direct mechanistic link between expression of the stem cell transcription factor GATA2 and drug resistance. The GATA-binding protein 2 (GATA2) plays a central role in blood stem cell generation and maintenance. We find substantial intrapatient and interpatient variability in GATA2 expression across samples from patients with AML. GATA2 expression varies by molecular subtype and has been linked to outcome. In a murine model, KMT2A-MLL3–driven AML originating from a stem cell or immature progenitor cell population has higher Gata2 expression and is more resistant to the standard AML chemotherapy agent doxorubicin. Deletion of Gata2 resulted in a more robust induction of p53 after exposure to doxorubicin. Chromatin immunoprecipitation sequencing, RNA sequencing, and functional studies revealed that GATA2 regulates the expression of RASSF4, a modulator of the p53 inhibitor MDM2 (mouse double minute 2). GATA2 and RASSF4 are anticorrelated in human cell lines and in bulk and single-cell expression data sets from patients with AML. Knockdown of Rassf4 in Gata2-low cells resulted in doxorubicin or nutlin-3 resistance. Conversely, overexpression of Rassf4 results in sensitization of cells expressing high levels of Gata2. Finally, doxorubicin and nutlin-3 are synergistic in Gata2-high murine AML and in samples from patients with AML. We discovered a previously unappreciated role for GATA2 in dampening p53-mediated apoptosis via transcriptional regulation of RASSF4, a modulator of MDM2. This role for GATA2 directly links the expression of a stemness-associated transcription factor to chemotherapy resistance.

Mast cells (MCs) play a critical role in allergic inflammation, anaphylaxis, and chronic inflammatory diseases such as asthma, COPD, and osteoarthritis. Dysregulated MC activation can lead to MC activation syndrome (MACS), which is observed in patients with long COVID. MCs express the high‐affinity receptor for IgE and, upon activation, release mediators and cytokines that trigger anaphylactic shock and promote allergic inflammation. They also interact with epithelial and nerve cells, which are crucial in forming a complex network of cell–cell and gene–gene interactions driving chronic inflammation that can confer resistance to treatment. In this review, in the context of the literature, we focus on experiments conducted in our laboratory investigating how transcription factors and enhancers regulate genes critical in mouse MC differentiation and function related to human lung inflammation.

Background: Antigenic stimulation through cross-linking the IgE receptor and epithelial cell-derived cytokine IL-33 are potent stimuli of mast cell (MC) activation. Moreover, IL-33 primes a variety of cell types, including MCs to respond more vigorously to external stimuli. However, target genes induced by the combined IL-33 priming and antigenic stimulation have not been investigated in human skin mast cells (HSMCs) in a genome-wide manner. Furthermore, epigenetic changes induced by the combined IL-33 priming and antigenic stimulation have not been evaluated. Results: We found that IL-33 priming of HSMCs enhanced their capacity to promote transcriptional synergy of the IL1B and CXCL8 genes by 16- and 3-fold, respectively, in response to combined IL-33 and antigen stimulation compared to without IL-33 priming. We identified the target genes in IL-33-primed HSMCs in response to the combined IL-33 and antigenic stimulation using RNA sequencing (RNA-seq). We found that the majority of genes synergistically upregulated in the IL-33-primed HSMCs in response to the combined IL-33 and antigenic stimulation were predominantly proinflammatory cytokine and chemokine genes. Moreover, the combined IL-33 priming and antigenic stimulation increase chromatin accessibility in the synergy target genes but not synergistically. Transcription factor binding motif analysis revealed more binding sites for NF-κB, AP-1, GABPA, and RAP1 in the induced or increased chromatin accessible regions of the synergy target genes. Conclusions: Our study demonstrates that IL-33 priming greatly potentiates MCs’ ability to transcribe proinflammatory cytokine and chemokine genes in response to antigenic stimulation, shining light on how epithelial cell-derived cytokine IL-33 can cause exacerbation of skin MC-mediated allergic inflammation.

Background: Histamine is a critical mediator of anaphylaxis, a neurotransmitter, and a regulator of gastric acid secretion. Histidine decarboxylase is a rate-limiting enzyme for histamine synthesis. However, in vivo regulation of Hdc, the gene that encodes histidine decarboxylase is poorly understood. Objective: We sought to investigate how enhancers regulate Hdc gene transcription and histamine synthesis in resting conditions and in a mouse model of anaphylaxis. Methods: H3K27 acetylation histone modification and chromatin accessibility were used to identify candidate enhancers; The enhancer activity of candidate enhancers was measured in a reporter gene assay; and the function enhancers were validated using CRISPR deletion. Results: Deletion of the GC box, which binds to zinc finger transcription factors, in the proximal Hdc enhancer, reduced Hdc gene transcription and histamine synthesis in the mouse and human mast cell lines. Mast cells, basophils, brain cells, and stomach cells from GC box-deficient mice transcribed the Hdc gene much less than similar cells from wild-type mice and Hdc GC box-deficient mice failed to develop anaphylaxis. Conclusion: Our results demonstrate that the HDC GC box within the proximal enhancer in the mouse and human HDC gene is essential for Hdc gene transcription, histamine synthesis, and histamine-mediated anaphylaxis in vitro and in vivo.

Macrophages play a critical role in inflammatory responses during infections. These cells are activated by infections through stimulation of TLRs expressed on their cell surface and produce pro-inflammatory cytokines, including TNF. However, distal enhancers that regulate TNF gene transcription in human macrophages have not been investigated. This study used an unbiased genomic approach to identify six candidate enhancers in human primary alveolar macrophages within a 131 kb region from the transcription start site (TSS) of the TNF gene, covering 13 genes. Of these candidate enhancers, five showed enhancer activity, with three targeting the TNF gene and two targeting neighboring genes. Deletion of the distal TNF E-16 enhancer led to a 73% reduction in TNF gene transcription in response to poly (I:C) stimulation in the THP-1 human leukemia monocytic cell line. Additionally, deletion of the E-7.1/hHS-8 enhancer resulted in a 41% reduction in TNF mRNA, while deletion of the PE enhancer had a lesser effect, resulting in a 52% reduction in TNF gene transcription. Massively parallel reporter assays (MPRA) indicated that the transcription factor AP-1 and EGR1-binding sites at the distal TNF E-16 enhancer were crucial in mediating enhancer activity. This study shows that both distal and proximal enhancers work together to fully transcribe the TNF gene in human macrophages in response to TLR ligand poly (I:C) stimulation.

Background: Antigenic stimulation through cross-linking the IgE receptor and epithelial cell-derived cytokine IL-33 are potent stimuli of mast cell (MC) activation. Moreover, IL-33 primes a variety of cell types, including MCs to respond more vigorously to external stimuli. However, target genes induced by the combined IL-33 priming and antigenic stimulation have not been investigated in human skin mast cells (HSMCs) in a genome-wide manner. Furthermore, epigenetic changes induced by the combined IL-33 priming and antigenic stimulation have not been evaluated. Results: We found that IL-33 priming of HSMCs enhanced their capacity to promote transcriptional synergy of the IL1B and CXCL8 genes by 16- and 3-fold, respectively, in response to combined IL-33 and antigen stimulation compared to without IL-33 priming. We identified the target genes in IL-33-primed HSMCs in response to the combined IL-33 and antigenic stimulation using RNA sequencing (RNA-seq). We found that the majority of genes synergistically upregulated in the IL-33-primed HSMCs in response to the combined IL-33 and antigenic stimulation were predominantly proinflammatory cytokine and chemokine genes. Moreover, the combined IL-33 priming and antigenic stimulation increase chromatin accessibility in the synergy target genes but not synergistically. Transcription factor binding motif analysis revealed more binding sites for NF-κB, AP-1, GABPA, and RAP1 in the induced or increased chromatin accessible regions of the synergy target genes. Conclusions: Our study demonstrates that IL-33 priming greatly potentiates MCs’ ability to transcribe proinflammatory cytokine and chemokine genes in response to antigenic stimulation, shining light on how epithelial cell-derived cytokine IL-33 can cause exacerbation of skin MC-mediated allergic inflammation.

Herein, a semianalytical model is presented for solitary wave diffraction by a surface-piercing concentric dual-arc thin wall. The two arc-shaped walls are porous and thin, and they are placed on a flat seafloor. As a key element, two imaginary closed cylindrical structures with two arc-shaped walls of different radii are introduced such that the entire computation domain can be partitioned into three subdomains, two bounded and one unbounded, based on common interfaces, within which the analytical solution is obtained using eigenfunction expression matching. Furthermore, a system of linear algebraic equations to determine the unknown coefficients is derived by satisfying the boundary and matching conditions. The numerical results obtained for the limiting cases are exactly the same as published results for a solid and porous cylindrical structure. Meanwhile, the effects of the wave incident angle, opening angle, annular spacing, and porous-effect parameter on wave loads and wave elevations are investigated. In addition, as a particular case, a concentric structure with an arc-shaped porous exterior wall is investigated as an alternative to the two-cylinder structure in practical engineering. It is discovered that the arc-shaped exterior wall can effectively reduce the wave force and wave height around the interior cylinder, compared with a single cylinder. By contrast, the wave forces and wave runup on the interior cylinder are similar to those of the interior cylinder of a concentric two-cylinder system.