Cell Type-specific Functions Research Articles

PPARs and Myocardial Infarction.

Peroxisome proliferator-activated receptors (PPARs) belong to the nuclear hormone receptor family. They are ligand-activated transcription factors and exist in three different isoforms, PPARα (NR1C1), PPARβ/δ (NR1C2), and PPARγ (NR1C3). PPARs regulate a variety of functions, including glucose and lipid homeostasis, inflammation, and development. They exhibit tissue and cell type-specific expression patterns and functions. Besides the established notion of the therapeutic potential of PPAR agonists for the treatment of glucose and lipid disorders, more recent data propose specific PPAR ligands as potential therapies for cardiovascular diseases. In this review, we focus on the knowledge of PPAR function in myocardial infarction, a severe pathological condition for which therapeutic use of PPAR modulation has been suggested.

Transforming growth factor β in breast cancer: another new trick for the old dog.

Two recent papers from the laboratory of Professor Ming Li demonstrate that inhibition of transforming growth factor β specifically in CD4+ T cells can suppress tumor growth through an unanticipated mechanism.

Nuclear pore complexes in development and tissue homeostasis.

Nuclear pore complexes are multiprotein channels that span the nuclear envelope, which connects the nucleus to the cytoplasm. In addition to their main role in the regulation of nucleocytoplasmic molecule exchange, it has become evident that nuclear pore complexes and their components also have multiple transport-independent functions. In recent years, an increasing number of studies have reported the involvement of nuclear pore complex components in embryogenesis, cell differentiation and tissue-specific processes. Here, we review the findings that highlight the dynamic nature of nuclear pore complexes and their roles in many cell type-specific functions during development and tissue homeostasis.

The Cell Type-Specific Functions of miR-21 in Cardiovascular Diseases.

Cardiovascular diseases are one of the prime reasons for disability and death worldwide. Diseases and conditions, such as hypoxia, pressure overload, infection, and hyperglycemia, might initiate cardiac remodeling and dysfunction by inducing hypertrophy or apoptosis in cardiomyocytes and by promoting proliferation in cardiac fibroblasts. In the vascular system, injuries decrease the endothelial nitric oxide levels and affect the phenotype of vascular smooth muscle cells. Understanding the underlying mechanisms will be helpful for the development of a precise therapeutic approach. Various microRNAs are involved in mediating multiple pathological and physiological processes in the heart. A cardiac enriched microRNA, miR-21, which is essential for cardiac homeostasis, has been demonstrated to act as a cell–cell messenger with diverse functions. This review describes the cell type–specific functions of miR-21 in different cardiovascular diseases and its prospects in clinical therapy.

DmPFC-vlPAG projection neurons contribute to pain threshold maintenance and antianxiety behaviors.

The dorsal medial prefrontal cortex (dmPFC) has been recognized as a key cortical area for nociceptive modulation. However, the underlying neural pathway and the function of specific cell types remain largely unclear. Here, we show that lesions in the dmPFC induced an algesic and anxious state. Using multiple tracing methods including a rabies-based transsynaptic tracing method, we outlined an excitatory descending neural pathway from the dmPFC to the ventrolateral periaqueductal gray (vlPAG). Specific activation of the dmPFC/vlPAG neural pathway by optogenetic manipulation produced analgesic and antianxiety effects in a mouse model of chronic pain. Inhibitory neurons in the dmPFC were specifically activated using a chemogenetic approach, which logically produced an algesic and anxious state under both normal and chronic pain conditions. Antagonists of the GABAA receptor (GABAAR) or mGluR1 were applied to the dmPFC, which produced analgesic and antianxiety effects. In summary, the results of our study suggest that the dmPFC/vlPAG neural pathway might participate in the maintenance of pain thresholds and antianxiety behaviors under normal conditions, while silencing or suppressing the dmPFC/vlPAG pathway might be involved in the initial stages and maintenance of chronic pain and the emergence of anxiety-like behaviors.

Mechanisms of neuronal survival safeguarded by endocytosis and autophagy.

Multiple aspects of neuronal physiology crucially depend on two cellular pathways, autophagy and endocytosis. During endocytosis, extracellular components either unbound or recognized by membrane-localized receptors (termed "cargo") become internalized into plasma membrane-derived vesicles. These can serve to either recycle the material back to the plasma membrane or send it for degradation to lysosomes. Autophagy also uses lysosomes as a terminal degradation point, although instead of degrading the plasma membrane-derived cargo, autophagy eliminates detrimental cytosolic material and intracellular organelles, which are transported to lysosomes by means of double-membrane vesicles, referred to as autophagosomes. Neurons, like all non-neuronal cells, capitalize on autophagy and endocytosis to communicate with the environment and maintain protein and organelle homeostasis. Additionally, the highly polarized, post-mitotic nature of neurons made them adopt these two pathways for cell-specific functions. These include the maintenance of the synaptic vesicle pool in the pre-synaptic terminal and the long-distance transport of signaling molecules. Originally discovered independently from each other, it is now clear that autophagy and endocytosis are closely interconnected and share several common participating molecules. Considering the crucial role of autophagy and endocytosis in cell type-specific functions in neurons, it is not surprising that defects in both pathways have been linked to the pathology of numerous neurodegenerative diseases. In this review, we highlight the recent knowledge of the role of endocytosis and autophagy in neurons with a special focus on synaptic physiology and discuss how impairments in genes coding for autophagy and endocytosis proteins can cause neurodegeneration.

Targeted expression of the arsenate reductase HAC1 identifies cell type specificity of arsenic metabolism and transport in plant roots.

High Arsenic Concentration 1 (HAC1), an Arabidopsis thaliana arsenate reductase, plays a key role in arsenate [As(V)] tolerance. Through conversion of As(V) to arsenite [As(III)], HAC1 enables As(III) export from roots, and restricts translocation of As(V) to shoots. To probe the ability of different root tissues to detoxify As(III) produced by HAC1, we generated A. thaliana lines expressing HAC1 in different cell types. We investigated the As(V) tolerance phenotypes: root growth, As(III) efflux, As translocation, and As chemical speciation. We showed that HAC1 can function in the outer tissues of the root (epidermis, cortex, and endodermis) to confer As(V) tolerance, As(III) efflux, and limit As accumulation in shoots. HAC1 is less effective in the stele at conferring As(V) tolerance phenotypes. The exception is HAC1 activity in the protoxylem, which we found to be sufficient to restrict As translocation, but not to confer As(V) tolerance. In conclusion, we describe cell type-specific functions of HAC1 that spatially separate the control of As(V) tolerance and As translocation. Further, we identify a key function of protoxylem cells in As(V) translocation, consistent with the model where endodermal passage cells, above protoxylem pericycle cells, form a ‘funnel’ loading nutrients and potentially toxic elements into the vasculature.

Regulation of Expression and Latency in BLV and HTLV.

Human T-lymphotrophic virus type 1 (HTLV-1) and Bovine leukemia virus (BLV) belong to the Deltaretrovirus genus. HTLV-1 is the etiologic agent of the highly aggressive and currently incurable cancer adult T-cell leukemia (ATL) and a neurological disease HTLV-1-associated myelopathy (HAM)/tropical spastic paraparesis (TSP). BLV causes neoplastic proliferation of B cells in cattle: enzootic bovine leucosis (EBL). Despite the severity of these conditions, infection by HTLV-1 and BLV appear in most cases clinically asymptomatic. These viruses can undergo latency in their hosts. The silencing of proviral gene expression and maintenance of latency are central for the establishment of persistent infection, as well as for pathogenesis in vivo. In this review, we will present the mechanisms that control proviral activation and retroviral latency in deltaretroviruses, in comparison with other exogenous retroviruses. The 5′ long terminal repeats (5′-LTRs) play a main role in controlling viral gene expression. While the regulation of transcription initiation is a major mechanism of silencing, we discuss topics that include (i) the epigenetic control of the provirus, (ii) the cis-elements present in the LTR, (iii) enhancers with cell-type specific regulatory functions, (iv) the role of virally-encoded transactivator proteins, (v) the role of repressors in transcription and silencing, (vi) the effect of hormonal signaling, (vii) implications of LTR variability on transcription and latency, and (viii) the regulatory role of non-coding RNAs. Finally, we discuss how a better understanding of these mechanisms may allow for the development of more effective treatments against Deltaretroviruses.

Rho GTPases: Big Players in Breast Cancer Initiation, Metastasis and Therapeutic Responses.

Rho GTPases, a family of the Ras GTPase superfamily, are key regulators of the actin cytoskeleton. They were originally thought to primarily affect cell migration and invasion; however, recent advances in our understanding of the biology and function of Rho GTPases have demonstrated their diverse roles within the cell, including membrane trafficking, gene transcription, migration, invasion, adhesion, survival and growth. As these processes are critically involved in cancer initiation, metastasis and therapeutic responses, it is not surprising that studies have demonstrated important roles of Rho GTPases in cancer. Although the majority of data indicates an oncogenic role of Rho GTPases, tumor suppressor functions of Rho GTPases have also been revealed, suggesting a context and cell-type specific function for Rho GTPases in cancer. This review aims to summarize recent progresses in our understanding of the regulation and functions of Rho GTPases, specifically in the context of breast cancer. The potential of Rho GTPases as therapeutic targets and prognostic tools for breast cancer patients are also discussed.

PPARs and Angiogenesis-Implications in Pathology.

Peroxisome proliferator-activated receptors (PPARs) belong to the family of ligand-activated nuclear receptors. The PPAR family consists of three subtypes encoded by three separate genes: PPARα (NR1C1), PPARβ/δ (NR1C2), and PPARγ (NR1C3). PPARs are critical regulators of metabolism and exhibit tissue and cell type-specific expression patterns and functions. Specific PPAR ligands have been proposed as potential therapies for a variety of diseases such as metabolic syndrome, cancer, neurogenerative disorders, diabetes, cardiovascular diseases, endometriosis, and retinopathies. In this review, we focus on the knowledge of PPAR function in angiogenesis, a complex process that plays important roles in numerous pathological conditions for which therapeutic use of PPAR modulation has been suggested.

Abstract MP143: Harnessing Glycomics to Understand Cardiac Biology and Disease

Cell surface glycoproteins play critical roles in maintaining cardiac structure and function, and the glycan-moiety attached to a protein is critical for proper protein folding, stability, and signaling. Despite mounting evidence that glycan structures are key modulators of heart function and must be considered when developing cardiac biomarkers, we currently do not have a comprehensive view of the glycans present in the normal human heart. Here, we used an innovative mass spectrometry approach to generate the first glycan structure libraries for primary human heart tissue, cardiomyocytes (CM) enriched from human heart tissue, and human induced pluripotent stem cell derived CM (hiPSC-CM), containing >260 N- and O- glycans. Comparing the glycome of CM enriched from primary heart tissue to that of heart tissue homogenate, 21 structures significantly differed, and the high mannose class is increased in enriched CM. Moreover, >30% of the glycome significantly changed across 20-100 days of in vitro differentiation, and only 23% of the N -glycan structures were shared between hiPSC-CM and primary CM. Overall, these observations are an important complement to genomic, transcriptomic, and proteomic profiling and reveal new considerations for the use and interpretation of hiPSC-CM models for studies of human development, disease, and drug testing. These data are also expected to aid in the evaluation of the immunogenic potential of hiPSC-CM for transplantation. Finally, harnessing differences observed between immature, proliferative hiPSC-CM and adult primary CM may be exploited to drive in vitro differentiation towards a more mature phenotype. Building on these data, current efforts are underway to develop chamber- and cell-type specific views ( e.g. cardiomyocytes, fibroblasts) of the glycome in the healthy and failing human heart. Such analyses provide a key link to understand the role glycosylation plays in cell-type specific functions and cardiac disease. The structural differences observed here, either among cell types or stages of differentiation, require complex regulation of multiple enzymes in the biosynthetic pathway, and therefore would be challenging to measure with antibody arrays, RNAseq, or proteomics. Therefore, continued application of structure-based glycomics approaches, such as the method used here, will be essential for elucidating the roles that glycans and glycoproteins play during developmental and disease processes in the human heart.

Conditional Gene Targeting Reveals Cell Type-Specific Roles of the Lysosomal Protease Cathepsin L in Mammary Tumor Progression.

Background: Cathepsin L (Ctsl) is a cysteine protease mainly located within the endosomal/lysosomal cell compartment. High expression of Ctsl indicates poor prognosis in human breast cancer. However, the cell type-specific Ctsl functions responsible for this association remain elusive. Methods: Because constitutive Ctsl−/− mice develop a complex phenotype, we developed a conditional model allowing for cell type-specific inactivation of Ctsl in mammary epithelium or myeloid cells in the transgenic mouse mammary tumor virus (MMTV)-polyoma middle T (PyMT) breast cancer model. Results: Ctsl ablation in mammary epithelial cells resulted in delayed initiation and end-stage of cancers. The latter displayed large dead cell areas. Inducible in vitro deletion of Ctsl in MMTV-PyMT-derived breast cancer cells revealed expansion of the acidic cell compartment, alteration of intracellular amino acid levels, and impaired mTOR signaling. In consequence, Ctsl-deficient cells exhibited slow growth rates and high apoptosis susceptibility. In contrast to Ctsl-deficient mammary epithelium, selective knockout of Ctsl in myeloid cells had no effects on primary tumors, but promoted lung metastasis formation. Conclusions: Our cell type-specific in vivo analysis provides strong evidence for a cancer cell-intrinsic, tumor-promoting role of Ctsl in primary breast cancer, whereas metastasis is negatively regulated by Ctsl expressed by bone marrow-derived cells.

Investigation of Isoform Specific Functions of the V-ATPase a Subunit During Drosophila Wing Development.

The vacuolar ATPases (V-ATPases) are ATP-dependent proton pumps that play vital roles in eukaryotic cells. Insect V-ATPases are required in nearly all epithelial tissues to regulate a multiplicity of processes including receptor-mediated endocytosis, protein degradation, fluid secretion, and neurotransmission. Composed of fourteen different subunits, several V-ATPase subunits exist in distinct isoforms to perform cell type specific functions. The 100 kD a subunit (Vha100) of V-ATPases are encoded by a family of five genes in Drosophila, but their assignments are not fully understood. Here we report an experimental survey of the Vha100 gene family during Drosophila wing development. A combination of CRISPR-Cas9 mutagenesis, somatic clonal analysis and in vivo RNAi assays is used to characterize the requirement of Vha100 isoforms, and mutants of Vha100-2, Vha100-3, Vha100-4, and Vha100-5 genes were generated. We show that Vha100-3 and Vha100-5 are dispensable for fly development, while Vha100-1 is not critically required in the wing. As for the other two isoforms, we find that Vha100-2 regulates wing cuticle maturation, while Vha100-4 is the single isoform involved in developmental patterning. More specifically, Vha100-4 is required for proper activation of the Wingless signaling pathway during fly wing development. Interestingly, we also find a specific genetic interaction between Vha100-1 and Vha100-4 during wing development. Our results revealed the distinct roles of Vha100 isoforms during insect wing development, providing a rationale for understanding the diverse roles of V-ATPases.

CRISPRi-based radiation modifier screen identifies long non-coding RNA therapeutic targets in glioma

BackgroundLong non-coding RNAs (lncRNAs) exhibit highly cell type-specific expression and function, making this class of transcript attractive for targeted cancer therapy. However, the vast majority of lncRNAs have not been tested as potential therapeutic targets, particularly in the context of currently used cancer treatments. Malignant glioma is rapidly fatal, and ionizing radiation is part of the current standard-of-care used to slow tumor growth in both adult and pediatric patients.ResultsWe use CRISPR interference (CRISPRi) to screen 5689 lncRNA loci in human glioblastoma (GBM) cells, identifying 467 hits that modify cell growth in the presence of clinically relevant doses of fractionated radiation. Thirty-three of these lncRNA hits sensitize cells to radiation, and based on their expression in adult and pediatric gliomas, nine of these hits are prioritized as lncRNA Glioma Radiation Sensitizers (lncGRS). Knockdown of lncGRS-1, a primate-conserved, nuclear-enriched lncRNA, inhibits the growth and proliferation of primary adult and pediatric glioma cells, but not the viability of normal brain cells. Using human brain organoids comprised of mature neural cell types as a three-dimensional tissue substrate to model the invasive growth of glioma, we find that antisense oligonucleotides targeting lncGRS-1 selectively decrease tumor growth and sensitize glioma cells to radiation therapy.ConclusionsThese studies identify lncGRS-1 as a glioma-specific therapeutic target and establish a generalizable approach to rapidly identify novel therapeutic targets in the vast non-coding genome to enhance radiation therapy.

SerpinB2 Regulates Immune Response in Kidney Injury and Aging.

Expression of SerpinB2, a regulator of inflammatory processes, has been described in the context of macrophage activation and cellular senescence. Given that mechanisms for these processes interact and can shape kidney disease, it seems plausible that SerpinB2 might play a role in renal aging, injury, and repair. We subjected SerpinB2 knockout mice to ischemia-reperfusion injury or unilateral ureteral obstruction. We performed phagocyte depletion to study SerpinB2's role beyond the effects of macrophages and transplanted bone marrow from knockout mice to wild-type mice and vice versa to dissect cell type-dependent effects. Primary tubular cells and macrophages from SerpinB2 knockout and wild-type mice were used for functional studies and transcriptional profiling. Cultured senescent tubular cells, kidneys of aged mice, and renal stress models exhibited upregulation of SerpinB2 expression. Functionally, lack of SerpinB2 in aged knockout mice had no effect on the magnitude of senescence markers but associated with enhanced kidney damage and fibrosis. In stress models, inflammatory cell infiltration was initially lower in knockout mice but later increased, leading to an accumulation of significantly more macrophages. SerpinB2 knockout tubular cells showed significantly reduced expression of the chemokine CCL2. Macrophages from knockout mice exhibited reduced phagocytosis and enhanced migration. Macrophage depletion and bone marrow transplantation experiments validated the functional relevance of these cell type-specific functions of SerpinB2. SerpinB2 influences tubule-macrophage crosstalk by supporting tubular CCL2 expression and regulating macrophage phagocytosis and migration. In mice, SerpinB2 expression seems to be needed for coordination and timely resolution of inflammation, successful repair, and kidney homeostasis during aging. Implications of SerpinB2 in human kidney disease deserve further exploration.

The conserved microRNA miR-34 regulates synaptogenesis via coordination of distinct mechanisms in presynaptic and postsynaptic cells

Micro(mi)RNA-based post-transcriptional regulatory mechanisms have been broadly implicated in the assembly and modulation of synaptic connections required to shape neural circuits, however, relatively few specific miRNAs have been identified that control synapse formation. Using a conditional transgenic toolkit for competitive inhibition of miRNA function in Drosophila, we performed an unbiased screen for novel regulators of synapse morphogenesis at the larval neuromuscular junction (NMJ). From a set of ten new validated regulators of NMJ growth, we discovered that miR-34 mutants display synaptic phenotypes and cell type-specific functions suggesting distinct downstream mechanisms in the presynaptic and postsynaptic compartments. A search for conserved downstream targets for miR-34 identified the junctional receptor CNTNAP4/Neurexin-IV (Nrx-IV) and the membrane cytoskeletal effector Adducin/Hu-li tai shao (Hts) as proteins whose synaptic expression is restricted by miR-34. Manipulation of miR-34, Nrx-IV or Hts-M function in motor neurons or muscle supports a model where presynaptic miR-34 inhibits Nrx-IV to influence active zone formation, whereas, postsynaptic miR-34 inhibits Hts to regulate the initiation of bouton formation from presynaptic terminals.

PD-1 Imposes Qualitative Control of Cellular Transcriptomes in Response to T Cell Activation.

Targeted blockade of programmed cell death 1 (PD-1), an immune-checkpoint receptor that inhibits Tcell activation, provides clinical benefits in various cancers. However, how PD-1 modulates gene expression in Tcells remains enigmatic. Here weinvestigated how PD-1 affects transcriptome changes induced by Tcell receptor (TCR) activation. Intriguingly, we identified a huge variance in PD-1 sensitivity among TCR-inducible genes. When we quantified the half maximal effective concentration (EC50) as the relationship between change in gene expression and TCR signal strength, we found that genes associated with survival and proliferation were efficiently expressed upon TCR activation and resistant to PD-1-mediated inhibition. Conversely, genes encoding cytokines and effector molecules were expressed less efficiently and sensitive to PD-1-mediated inhibition. We further demonstrated that transcription factor binding motifs and CpG frequency in the promoter region affect EC50 and thus the PD-1 sensitivity of genes. Our findings explain how PD-1, dependent on the TCR signal strength, calibrates cellular transcriptomes to shape functional properties of Tcell populations.

In vivo single-cell lineage tracing in zebrafish using high-resolution infrared laser-mediated gene induction microscopy.

Heterogeneity broadly exists in various cell types both during development and at homeostasis. Investigating heterogeneity is crucial for comprehensively understanding the complexity of ontogeny, dynamics, and function of specific cell types. Traditional bulk-labeling techniques are incompetent to dissect heterogeneity within cell population, while the new single-cell lineage tracing methodologies invented in the last decade can hardly achieve high-fidelity single-cell labeling and long-term in-vivo observation simultaneously. In this work, we developed a high-precision infrared laser-evoked gene operator heat-shock system, which uses laser-induced CreERT2 combined with loxP-DsRedx-loxP-GFP reporter to achieve precise single-cell labeling and tracing. In vivo study indicated that this system can precisely label single cell in brain, muscle and hematopoietic system in zebrafish embryo. Using this system, we traced the hematopoietic potential of hemogenic endothelium (HE) in the posterior blood island (PBI) of zebrafish embryo and found that HEs in the PBI are heterogeneous, which contains at least myeloid unipotent and myeloid-lymphoid bipotent subtypes.

An integrated hypothesis for miR-126 in vascular disease.

microRNA miR-126 was among the early discovered miRNAs that are expressed specifically in the vasculature and have critical functions in vascular development. Recent studies have started to unveil potentially important function of miR-126 in vascular diseases, including atherosclerosis, coronary artery disease, stroke and diabetic vasculopathy. The action of miR-126 reflects its function in angiogenesis and inflammation. The expression of miR-126 is downregulated in a variety of vascular diseases, and miR-126 overexpression appears to beneficial for most vascular disease models. In the minireview, we summarize the historic and current research regarding miR-126 function and mechanisms in the vascular system, its link to long noncoding RNAs (lncRNA), as well as the potential of miR-126-based therapeutics for vascular diseases. To explain the seemingly conflicting function of miR-126 from different studies, an integrated hypothesis is proposed that miR-126 has strand- and cell type-specific functions in angiogenesis and inflammation, making it beneficial in many different vascular disease models.

Cell-specific and athero-protective roles for RIPK3 in a murine model of atherosclerosis.

ABSTRACTReceptor-interacting protein kinase 3 (RIPK3) was recently implicated in promoting atherosclerosis progression through a proposed role in macrophage necroptosis. However, RIPK3 has been connected to numerous other cellular pathways, which raises questions about its actual role in atherosclerosis. Furthermore, RIPK3 is expressed in a multitude of cell types, suggesting that it may be physiologically relevant to more than just macrophages in atherosclerosis. In this study, Ripk3 was deleted in macrophages, endothelial cells, vascular smooth muscle cells or globally on the Apoe−/− background using Cre-lox technology. To induce atherosclerosis progression, male and female mice were fed a Western diet for three months before tissue collection and analysis. Surprisingly, necroptosis markers were nearly undetectable in atherosclerotic aortas. Furthermore, en face lesion area was increased in macrophage- and endothelial-specific deletions of Ripk3 in the descending and abdominal regions of the aorta. Analysis of bone-marrow-derived macrophages and cultured endothelial cells revealed that Ripk3 deletion promotes expression of monocyte chemoattractant protein 1 (MCP-1) and E-selectin in these cell types, respectively. Western blot analysis showed upregulation of MCP-1 in aortas with Ripk3-deficient macrophages. Altogether, these data suggest that RIPK3 in macrophages and endothelial cells protects against atherosclerosis through a mechanism that likely does not involve necroptosis. This protection may be due to RIPK3-mediated suppression of pro-inflammatory MCP-1 expression in macrophages and E-selectin expression in endothelial cells. These findings suggest a novel and unexpected cell-type specific and athero-protective function for RIPK3.This article has an associated First Person interview with the first author of the paper.