Maf Transcription Factors Research Articles

Pancreatic expression of CPT1A is essential for whole body glucose homeostasis by supporting glucose-stimulated insulin secretion.

Pancreatic islet β-cells express the Cpt1a gene, which encodes the enzyme carnitine palmitoyltransferase 1A (CPT1A), an enzyme that facilitates entry of long chain fatty acids into the mitochondria. Because fatty acids are required for glucose-stimulated insulin secretion, we tested the hypothesis that CPT1A is essential to support islet β-cell function and mass. In this study, we describe genetic deletion of Cpt1a in pancreatic tissue (Cpt1aPdx1-/-) using C57BL/6J mice. Islet morphology, β-cell transcription factor abundance, islet ATP levels, GLUT2 abundance, and expression of the de-differentiation marker ALDH1A3 were analyzed by immunofluorescent staining. Glucose and insulin tolerance were assessed to investigate the metabolic status of genetic reductions in Cpt1a. Glucose-stimulated insulin secretion was evaluated in vivo and in isolated islets ex vivo by perifusion. Pancreatic deletion of Cpt1a reduced glucose tolerance but did not alter insulin sensitivity. Glucose-stimulated insulin secretion was reduced both in vivo and in islets isolated from Cpt1aPdx1-/- mice relative to control islets. Pancreatic islets from Cpt1aPdx1-/- mice displayed elevations in ALDH1A3, a marker of de-differentiation, but no reduction in nuclear abundance of the β-cell transcription factors MafA and Nkx6.1 or the GLUT2 glucose transporter. However, intracellular ATP abundance was markedly decreased in islets isolated from Cpt1aPdx1-/- relative to littermate control mice. We conclude that there is an important physiological role for pancreatic CPT1A to maintain whole-body glucose homeostasis by supporting glucose-stimulated insulin secretion and maintaining intracellular ATP levels in male mice.

Pancreatic Differentiation of Oral Minor Salivary Gland Stem Cells.

Stem cells from various sources including major salivary glands have been used to establish pancreatic differentiation in an attempt to provide new treatment options for patients with diabetes mellitus. In contrast, the potential of using the more easily accessible intraoral minor salivary glands has not been evaluated so far. Salivary stem cells were isolated from normal labial minor salivary glands that were removed during the excision of a mucocele and were attempted to differentiate into pancreatic cell lines using a culture medium enriched with activin A, retinoic acid and GLP-1.Real time RT-PCR was used to evaluate the expression of the genes of pancreatic transcription factors MafA, Ptf1a, Hb9 and Arx. Complementary, 22 labial minor salivary gland paraffin-embedded specimens were examined using immunohistochemistry for the presence of the relevant gene products of the pancreatic transcription factors Arx, MafA, Ptf1a and Pdx1. The differentiated salivary stem cells(cells of passage 3) expressed the genes of the pancreatic transcription factors MafA, Ptf1a, Hb9 and Arx even on the first day of the experiment while immunohistochemistry also confirmed the presence of the protein products of Arx, MafA, Ptf1a as well as Pdx1[> 50% of the specimens for Arx(5/8) and MafA(7/8), < 50% for Ptf1a(5/11) and Pdx1(5/11)] in ducts, mesenchymal connective tissue and acinar cells. Labial minor salivary glands may share gene and protein characteristics with pancreas suggesting a possible usefulness for pancreatic regeneration or substitution in cases of deficiency.

The small MAF transcription factor MAFG co-opts MITF to promote melanoma progression.

Transcription factor deregulation potently drives melanoma progression by dynamically and reversibly controlling gene expression programs. We previously identified the small MAF family transcription factor MAFG as a putative driver of melanoma progression, prompting an in-depth evaluation of its role in melanoma. MAFG expression increases with human melanoma stages and ectopic MAFG expression enhances the malignant behavior of human melanoma cells in vitro, xenograft models, and genetic mouse models of spontaneous melanoma. Moreover, MAFG induces a melanoma phenotype switch from a melanocytic state to a more dedifferentiated state. Mechanistically, MAFG interacts with the lineage transcription factor MITF which is required for the pro-tumorigenic effects of MAFG. MAFG and MITF co-occupy numerous genomic sites and MAFG overexpression influences the expression of genes harboring binding sites for the MAFG~MITF complex. These results establish MAFG as a potent driver of melanomagenesis through dimerization with MITF and uncover an unappreciated mechanism of MITF regulation.

Abstract 3041: The small MAF transcription factor MAFG is a potent driver of melanoma

Abstract While common mutations in potent proto-oncogenes and tumor suppressors induce melanoma formation, no recurrent mutations associated with the late stages of melanomagenesis have been identified. Instead, non-mutational mechanisms such as the deregulation of transcriptional or epigenetic programs typically promote the malignant progression and metastasis of melanoma. We have previously identified the small MAF transcription factor MAFG as a critical target of the melanoma suppressor miRNA miR-29, prompting an in-depth evaluation of the role of MAFG in melanoma. MAFG expression is elevated in melanoma compared to melanocytes and increases with tumor stage. Ectopic expression of MAFG in human melanocytes and melanoma cells enhances proliferation in vitro and promotes melanoma growth in vivo. Moreover, MAFG overexpression in BrafV600E; PtenFL/+ mice accelerated melanomagenesis, significantly shortening overall survival. Despite being a critical NRF2 dimerization partner, NRF2 was dispensable for the effects of MAFG. Moreover, MAFG overexpression in melanoma cells had no effect on the activity of NRF2 transcriptional reporters, nor did it induce transcriptional programs associated with NRF2. RNA sequencing and pathway analysis revealed MAFG-mediated effects on melanoma-associated processes such hypoxia, the PI3K/AKT pathway, and pigmentation. Indeed, MAFG induced HIF1a under normoxic conditions, enhanced AKT phosphorylation, and downregulated MITF and its target genes in the pigmentation pathway in melanoma cells. Interestingly, MAFG also potently enhanced AXL expression, suggesting that MAFG may promote melanoma, at least in part, by affecting the MITF/AXL balance that is associated with melanoma phenotype switching. We next analyzed whether MAFG is required for melanoma. Interestingly, silencing MAFG in human melanoma cells robustly inhibited growth in vitro. Moreover, the CRISPR/Cas9-mediated knockout of MAFG in an aggressive BrafV600E; PtenFL/FL model potently prevented melanoma formation. Given the mild phenotype of MAFG whole-body knockout mice, our results suggest targeting of MAFG or its downstream pathways as a viable therapeutic approach in melanoma. In conclusion, our study establishes MAFG as a potent driver of melanoma development and nominates it as a therapeutic target. Citation Format: Olga Vera, Michael Martinez, Zulaida Soto-Vargas, Xiaonan Xu, Florian Karreth. The small MAF transcription factor MAFG is a potent driver of melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3041.

Integrated single-cell multiomics uncovers foundational regulatory mechanisms of lens development and pathology.

Ocular lens development entails epithelial to fiber cell differentiation, defects in which cause congenital cataracts. We report the first single-cell multiomic atlas of lens development, leveraging snRNA-seq, snATAC-seq and CUT&RUN-seq to discover previously unreported mechanisms of cell fate determination and cataract-linked regulatory networks. A comprehensive profile of cis- and trans-regulatory interactions, including for the cataract-linked transcription factor MAF, is established across a temporal trajectory of fiber cell differentiation. Furthermore, we identify an epigenetic paradigm of cellular differentiation, defined by progressive loss of the H3K27 methylation writer Polycomb repressive complex 2 (PRC2). PRC2 localizes to heterochromatin domains across master-regulator transcription factor gene bodies, suggesting it safeguards epithelial cell fate. Moreover, we demonstrate that FGF hyper-stimulation in vivo leads to MAF network activation and the emergence of novel lens cell states. Collectively, these data depict a comprehensive portrait of lens fiber cell differentiation, while defining regulatory effectors of cell identity and cataract formation.

Identifying a locus in super-enhancer and its resident NFE2L1/MAFG as transcriptional factors that drive PD-L1 expression and immune evasion

Although the transcriptional regulation of the programmed death ligand 1 (PD-L1) promoter has been extensively studied, the transcription factor residing in the PD-L1 super-enhancer has not been comprehensively explored. Through saturated CRISPR-Cas9 screening of the core region of the PD-L1 super-enhancer, we have identified a crucial genetic locus, referred to as locus 22, which is essential for PD-L1 expression. Locus 22 is a potential binding site for NFE2:MAF transcription factors. Although genetic silencing of NRF2 (NFE2L2) did not result in a reduction of PD-L1 expression, further analysis reveals that MAFG and NFE2L1 (NRF1) play a critical role in the expression of PD-L1. Importantly, lipopolysaccharides (LPS) as the major component of intratumoral bacteria could greatly induce PD-L1 expression, which is dependent on the PD-L1 super-enhancer, locus 22, and NFE2L1/MAFG. Mechanistically, genetic modification of locus 22 and silencing of MAFG greatly reduce BRD4 binding and loop formation but have minimal effects on H3K27Ac modification. Unlike control cells, cells with genetic modification of locus 22 and silencing of NFE2L1/MAFG failed to escape T cell-mediated killing. In breast cancer, the expression of MAFG is positively correlated with the expression of PD-L1. Taken together, our findings demonstrate the critical role of locus 22 and its associated transcription factor NFE2L1/MAFG in super-enhancer– and LPS-induced PD-L1 expression. Our findings provide new insight into understanding the regulation of PD-L1 transcription and intratumoral bacteria-mediated immune evasion.

Comprehensive Search for GPCR Compounds which Can Enhance MafA and/or PDX-1 Expression Levels Using a Small Molecule Compound Library.

It has been shown that chronic hyperglycemia gradually decreases insulin biosynthesis and secretion which is accompanied by reduced expression of very important insulin gene transcription factors MafA and PDX-1. Such phenomena are well known as β-cell glucose toxicity. It has been shown that the downregulation of MafA and/or PDX-1 expression considerably explains the molecular mechanism for glucose toxicity. However, it remained unknown which molecules can enhance MafA and/or PDX-1 expression levels. In this study, we comprehensively searched for G protein-coupled receptor (GPCR) compounds which can enhance MafA and/or PDX-1 expression levels using a small molecule compound library in pancreatic β-cell line MIN6 cells and islets isolated from nondiabetic C57BL/6 J mice and obese type 2 diabetic C57BL/KsJ-db/db mice. We found that fulvestrant and dexmedetomidine hydrochloride increased MafA, PDX-1, or insulin expression levels in MIN6 cells. We confirmed that fulvestrant and dexmedetomidine hydrochloride increased MafA, PDX-1, or insulin expression levels in islets from nondiabetic mice as well. Furthermore, these reagents more clearly enhanced MafA, PDX-1, or insulin expression levels in islets from obese type 2 diabetic db/db mice in which MafA and PDX-1 expression levels are reduced due to glucose toxicity. In conclusion, fulvestrant and dexmedetomidine hydrochloride increased MafA, PDX-1, or insulin expression levels in MIN6 cells and islets from nondiabetic mice and obese type 2 diabetic db/db mice. To the best of our knowledge, this is the first report showing some molecule which can enhance MafA and/or PDX-1 expression levels. Therefore, although further extensive study is necessary, we think that the information in this study could be, at least in part, useful at some point such as in the development of new antidiabetes medicine based on the molecular mechanism of β-cell glucose toxicity in the future.

Opposing gene regulatory programs governing myofiber development and maturation revealed at single nucleus resolution

Skeletal muscle fibers express distinct gene programs during development and maturation, but the underlying gene regulatory networks that confer stage-specific myofiber properties remain unknown. To decipher these distinctive gene programs and how they respond to neural activity, we generated a combined multi-omic single-nucleus RNA-seq and ATAC-seq atlas of mouse skeletal muscle development at multiple stages of embryonic, fetal, and postnatal life. We found that Myogenin, Klf5, and Tead4 form a transcriptional complex that synergistically activates the expression of muscle genes in developing myofibers. During myofiber maturation, the transcription factor Maf acts as a transcriptional switch to activate the mature fast muscle gene program. In skeletal muscles of mutant mice lacking voltage-gated L-type Ca2+ channels (Cav1.1), Maf expression and myofiber maturation are impaired. These findings provide a transcriptional atlas of muscle development and reveal genetic links between myofiber formation, maturation, and contraction.

Therapeutic Potential of Amniotic Fluid-Derived Stem Cells into Pancreatic Lineage in an Animal Model of Diabetes

Diabetes is a major health concern and it is estimated that India became diabetic capital in the future. The therapeutic potential of various medications, cell therapy, and islet transplantation is explored in recent years. The therapeutic potential of Mesenchymal Stem cells (MSCs) from bone marrow and from other sources has been studied forthe treatment of diabetes. MSCsfrom bone marrow ameliorate hyperglycemia but are unable to restore normoglycaemia in diabetic animals when injected with a single dose. The therapeutic potential of another more primitive source of MSCs from human amniotic fluid-derived stem cells (hAFSCs) has not been explored in diabetes. In the present study, we have assessed the effect of hAFSCs in a rat model of diabetes. hAFSCs transplantation has been seen to control blood glucose levels, and promote weight gain and other physiological parameters. Immuno-histochemical results suggested an increase in islet mass and number after hAFSCs transplantation. Q-RTPCR results showed transcription factors Ins-1, Oct-4, MAF-A, and PDX-1 were markedly upregulated in the hAFSCs transplanted group in comparison with diabetes and control groups. These data suggest that hAFSCs possess considerable therapeutic potential for diabetes through the restoration of insulin mass and up regulations of transcription factors of pancreatic lineage. Keyword : Amniotic fluid stem cells, diabetic mellitus, rat

Stapled Peptides as Direct Inhibitors of Nrf2-sMAF Transcription Factors.

Nuclear factor erythroid-related 2-factor 2 (Nrf2) is a transcription factor traditionally thought of as a cellular protector. However, in many cancers, Nrf2 is constitutively activated and correlated with therapeutic resistance. Nrf2 heterodimerizes with small musculoaponeurotic fibrosarcoma Maf (sMAF) transcription factors, allowing binding to the antioxidant responsive element (ARE) and induction of transcription of Nrf2 target genes. While transcription factors are historically challenging to target, stapled peptides have shown great promise for inhibiting these protein-protein interactions. Herein, we describe the first direct cell-permeable inhibitor of Nrf2/sMAF heterodimerization. N1S is a stapled peptide designed based on AlphaFold predictions of the interactions between Nrf2 and sMAF MafG. A cell-based reporter assay combined with in vitro biophysical assays demonstrates that N1S directly inhibits Nrf2/MafG heterodimerization. N1S treatment decreases the transcription of Nrf2-dependent genes and sensitizes Nrf2-dependent cancer cells to cisplatin. Overall, N1S is a promising lead for the sensitization of Nrf2-addicted cancers.

CLEC11A improves insulin secretion and promotes cell proliferation in human beta-cells.

Beta-cell dysfunction is a hallmark of disease progression in patients with diabetes. Research has been focused on maintaining and restoring beta-cell function during diabetes development. The aims of this study were to explore the expression of C-type lectin domain containing 11A (CLEC11A), a secreted sulphated glycoprotein, in human islets and to evaluate the effects of CLEC11A on beta-cell function and proliferation in vitro. To test these hypotheses, human islets and human EndoC-βH1 cell line were used in this study. We identified that CLEC11A was expressed in beta-cells and alpha-cells in human islets but not in EndoC-βH1 cells, whereas the receptor of CLEC11A called integrin subunit alpha 11 was found in both human islets and EndoC-βH1 cells. Long-term treatment with exogenous recombinant human CLEC11A (rhCLEC11A) accentuated glucose-stimulated insulin secretion, insulin content, and proliferation from human islets and EndoC-βH1 cells, which was partially due to the accentuated expression levels of transcription factors MAFA and PDX1. However, the impaired beta-cell function and reduced mRNA expression of INS and MAFA in EndoC-βH1 cells that were caused by chronic palmitate exposure could only be partially improved by the introduction of rhCLEC11A. Based on these results, we conclude that rhCLEC11A promotes insulin secretion, insulin content, and proliferation in human beta-cells, which are associated with the accentuated expression levels of transcription factors MAFA and PDX1. CLEC11A, therefore, may provide a novel therapeutic target for maintaining beta-cell function in patients with diabetes.

The ubiquitin ligase HERC4 suppresses MafA transcriptional activity triggered by GSK3β in myeloma by atypical K63-linked polyubiquitination

MafA and c-Maf are close members of the Maf transcription factor family and indicators of poor prognosis of multiple myeloma (MM). Our previous study finds that the ubiquitin ligase HERC4 induces c-Maf degradation but stabilizes MafA, and the mechanism is elusive. In the present study we find that HERC4 interacts with MafA and mediates its K63-linked polyubiquitination at K33. Moreover, HERC4 inhibits MafA phosphorylation and its transcriptional activity triggered by glycogen synthase kinase 3β (GSK3β). The K33R MafA variant prevents HERC4 from inhibiting MafA phosphorylation and increases MafA transcriptional activity. Further analyses reveal that MafA can also activate the STAT3 signaling but it is suppressed by HERC4. Lastly, we demonstrate that lithium chloride, a GSK3β inhibitor, can upregulate HERC4 and synergizes dexamethasone, a typical anti-MM drug, in inhibiting MM cell proliferation and xenograft growth in nude mice. These findings thus highlight a novel regulation of MafA oncogenic activity in MM and provide the rationale by targeting HERC4/GSK3β/MafA for the treatment of MM.

Large Maf transcription factor family is a major regulator of fast type IIb myofiber determination

Myofibers are broadly characterized as fatigue-resistant slow-twitch (type I) fibers and rapidly fatiguing fast-twitch (type IIa/IIx/IIb) fibers. However, the molecular regulation of myofiber type is not entirely understood; particularly, information on regulators of fast-twitch muscle is scarce. Here, we demonstrate that the large Maf transcription factor family dictates fast type IIb myofiber specification in mice. Remarkably, the ablation of three large Mafs leads to the drastic loss of type IIb myofibers, resulting in enhanced endurance capacity and the reduction of muscle force. Conversely, the overexpression of each large Maf in the type I soleus muscle induces type IIb myofibers. Mechanistically, a large Maf directly binds to the Maf recognition element on the promoter of myosin heavy chain 4, which encodes the type IIb myosin heavy chain, driving its expression. This work identifies the large Maf transcription factor family as a major regulator for fast type IIb muscle determination.

MafB-dependent neurotransmitter signaling promotes β cell migration in the developing pancreas.

Hormone secretion from pancreatic islets is essential for glucose homeostasis and loss or dysfunction of islet cells is a hallmark of type 2 diabetes. Maf transcription factors are critical for establishing and maintaining adult endocrine cell function. However, during pancreas development, MafB is not only expressed in insulin- and glucagon-producing cells, but also Neurog3+ endocrine progenitor cells suggesting additional functions in cell differentiation and islet formation. Here we report that MafB deficiency impairs β cell clustering and islet formation, but also coincides with loss of neurotransmitter and axon guidance receptor gene expression. Moreover, the observed loss of nicotinic receptor gene expression in human and mouse β cells implied that signaling through these receptors contributes to islet cell migration/formation. Inhibition of nicotinic receptor activity resulted in reduced β cell migration towards autonomic nerves and impaired β cell clustering. These findings highlight a novel function of MafB in controlling neuronal-directed signaling events required for islet formation.

Expression Silencing of Mitogen-Activated Protein Kinase 8 Interacting Protein-1 Conferred Its Role in Pancreatic β-Cell Physiology and Insulin Secretion.

Mitogen-activated protein kinase 8 interacting protein-1 (MAPK8IP1) gene has been recognized as a susceptibility gene for diabetes. However, its action in the physiology of pancreatic β-cells is not fully understood. Herein, bioinformatics and genetic analyses on the publicly available database were performed to map the expression of the MAPK8IP1 gene in human pancreatic islets and to explore whether this gene contains any genetic variants associated with type 2 diabetes (T2D). Moreover, a series of functional experiments were executed in a rat insulinoma cell line (INS-1 832/13) to investigate the role of the Mapk8ip1 gene in β-cell function. Metabolic engineering using RNA-sequencing (RNA-seq) data confirmed higher expression levels of MAPK8IP1 in human islets compared to other metabolic tissues. Additionally, comparable expression of MAPK8IP1 expression was detected in sorted human endocrine cells. However, β-cells exhibited higher expression of MAPK8IP1 than ductal and PSC cells. Notably, MAPK8IP1 expression was reduced in diabetic islets, and the expression was positively correlated with insulin and the β-cell transcription factor PDX1 and MAFA. Using the TIGER portal, we found that one genetic variant, "rs7115753," in the proximity of MAPK8IP1, passes the genome-wide significance for the association with T2D. Expression silencing of Mapk8ip1 by small interfering RNA (siRNA) in INS-1 cells reduced insulin secretion, glucose uptake rate, and reactive oxygen species (ROS) production. In contrast, insulin content, cell viability, and apoptosis without cytokines were unaffected. However, silencing of Mapk8ip1 reduced cytokines-induced apoptosis and downregulated the expression of several pancreatic β-cell functional markers including, Ins1, Ins2, Pdx1, MafA, Glut2, Gck, Insr, Vamp2, Syt5, and Cacna1a at mRNA and/or protein levels. Finally, we reported that siRNA silencing of Pdx1 resulted in the downregulation of MAPK8IP1 expression in INS-1 cells. In conclusion, our findings confirmed that MAPK8IP1 is an important component of pancreatic β-cell physiology and insulin secretion.

Specific reprogramming of alpha cells to insulin-producing cells by short glucagon promoter-driven Pdx1 and MafA

Endogenous reprogramming of pancreas-derived non-beta cells into insulin-producing cells is a promising approach to treat type 1 diabetes (T1D). One strategy that has yet to be explored is the specific delivery of insulin-producing essential genes, Pdx1 and MafA, to pancreatic alpha cells to reprogram the cells into insulin-producing cells in an adult pancreas. In this study, we used an alpha cell-specific glucagon (GCG) promoter to drive Pdx1 and MafA transcription factors to reprogram alpha cells to insulin-producing cells in chemically induced and autoimmune diabetic mice. Our results showed that a combination of a short glucagon-specific promoter with AAV serotype 8 (AAV8) can be used to successfully deliver Pdx1 and MafA to pancreatic alpha cells in the mouse pancreas. Pdx1 and MafA expression specifically in alpha cells were also able to correct hyperglycemia in both induced and autoimmune diabetic mice. With this technology, targeted gene specificity and reprogramming were accomplished with an alpha-specific promotor combined with an AAV-specific serotype and provide an initial basis to develop a novel therapy for the treatment of T1D.

The transcription factors ISL-1 and MAFA, but not NKX6-1 Characterize a Stem-cell Derived Population of Endocrine Pancreatic Cells Capable of Controlling blood Glucose in Rodent Models of Diabetes

A stem cell line, derived by reprogramming native human islet cells, consistently generates pure populations of endocrine pancreatic clusters following differentiation cues. Two protocols were developed directing the differentiation of that cell line to pancreatic cells that expressed or lacked expression of the key beta cell maturation-associated factor NKX6-1. The population of stem cell derived endocrine pancreatic clusters that was most consistently capable of regulating blood glucose in rodent models of diabetes lacked NKX6-1 but did manifest high expression of other key drivers of endocrine cell specification and maturation, ISL1 and MAFA. These data support the hypothesis that MAFA and ISL-1, but not NKX6-1, are reliable in vitro markers of committed endocrine pancreatic cells. The population with low NKX6-1 and high in vivo potency was further characterized by transcriptome profiling as an endocrine-committed population progressively maturing in vitro to a state proximal to the native islet.

GSK3β Inhibition Prevents Macrophage Reprogramming by High-Dose Methotrexate

Methotrexate (MTX) is an antifolate drug used as a chemotherapeutic agent for acute lymphoblastic leukemia, where MTX improves patients’ prognosis. Macrophage reprogramming is being increasingly assessed as an antitumor therapeutic strategy. However, and although MTX limits the pathogenic action of macrophages in chronic inflammatory diseases, its effects on tumor-promoting macrophages have not been previously explored. We now report that MTX shapes the transcriptional and functional profile of M-CSF-dependent human macrophages, whose transcriptome is highly enriched in the gene signature that defines pathogenic tumor-associated macrophages (“large TAM”). Specifically, MTX prompted the acquisition of the gene signature of antitumoral “small TAM” and skewed macrophages toward an IL-6high IFNβ1high IL-10low phenotype upon subsequent stimulation. Mechanistically, the MTX-induced macrophage reprogramming effect correlated with a reduction of the M-CSF receptor CSF1R expression and function, as well as a diminished expression of MAF and MAFB transcription factors, primary determinants of pro-tumoral macrophages whose transcriptional activity is dependent on GSK3β. Indeed, the ability of MTX to transcriptionally reprogram macrophages toward an antitumoral phenotype was abrogated by inhibition of GSK3β. Globally, our results establish MTX as a macrophage reprogramming drug and indicate that its ability to modulate macrophage polarization may also underlie its therapeutic benefits. Since GSK3β inhibition abrogates the reprogramming action of MTX, our results suggest that the GSK3β-MAFB/MAF axis constitutes a target for the macrophage-centered antitumor strategies.

C1Q labels a highly aggressive macrophage-like leukemia population indicating extramedullary infiltration and relapse

AbstractExtramedullary infiltration (EMI) is a concomitant manifestation that may indicate poor outcome of acute myeloid leukemia (AML). The underlying mechanism remains poorly understood and therapeutic options are limited. Here, we employed single-cell RNA sequencing on bone marrow (BM) and EMI samples from a patient with AML presenting pervasive leukemia cutis. A complement C1Q+ macrophage-like leukemia subset, which was enriched within cutis and existed in BM before EMI manifestations, was identified and further verified in multiple patients with AML. Genomic and transcriptional profiling disclosed mutation and gene expression signatures of patients with EMI that expressed high levels of C1Q. RNA sequencing and quantitative proteomic analysis revealed expression dynamics of C1Q from primary to relapse. Univariate and multivariate analysis demonstrated adverse prognosis significance of C1Q expression. Mechanistically, C1Q expression, which was modulated by transcription factor MAF BZIP transcription factor B, endowed leukemia cells with tissue infiltration ability, which could establish prominent cutaneous or gastrointestinal EMI nodules in patient-derived xenograft and cell line–derived xenograft models. Fibroblasts attracted migration of the C1Q+ leukemia cells through C1Q–globular C1Q receptor recognition and subsequent stimulation of transforming growth factor β1. This cell-to-cell communication also contributed to survival of C1Q+ leukemia cells under chemotherapy stress. Thus, C1Q served as a marker for AML with adverse prognosis, orchestrating cancer infiltration pathways through communicating with fibroblasts and represents a compelling therapeutic target for EMI.

Mafa-dependent GABAergic activity promotes mouse neonatal apneas

While apneas are associated with multiple pathological and fatal conditions, the underlying molecular mechanisms remain elusive. We report that a mutated form of the transcription factor Mafa (Mafa4A) that prevents phosphorylation of the Mafa protein leads to an abnormally high incidence of breath holding apneas and death in newborn Mafa4A/4A mutant mice. This apneic breathing is phenocopied by restricting the mutation to central GABAergic inhibitory neurons and by activation of inhibitory Mafa neurons while reversed by inhibiting GABAergic transmission centrally. We find that Mafa activates the Gad2 promoter in vitro and that this activation is enhanced by the mutation that likely results in increased inhibitory drives onto target neurons. We also find that Mafa inhibitory neurons are absent from respiratory, sensory (primary and secondary) and pontine structures but are present in the vicinity of the hypoglossal motor nucleus including premotor neurons that innervate the geniohyoid muscle, to control upper airway patency. Altogether, our data reveal a role for Mafa phosphorylation in regulation of GABAergic drives and suggest a mechanism whereby reduced premotor drives to upper airway muscles may cause apneic breathing at birth.