Subject(s)
COVID-19/virology , Receptors, Virus/genetics , SARS-CoV-2/genetics , Transcription Factors/genetics , Angiotensin-Converting Enzyme 2/metabolism , Animals , CDX2 Transcription Factor/genetics , CDX2 Transcription Factor/metabolism , COVID-19/genetics , COVID-19/metabolism , Disease Models, Animal , Erythroid-Specific DNA-Binding Factors/genetics , Hepatocyte Nuclear Factor 4/genetics , Humans , Intestinal Mucosa/metabolism , Intestinal Mucosa/virology , Intestines/virology , Mice , SARS-CoV-2/metabolism , Serine Endopeptidases/metabolism , Smad4 Protein/genetics , Spike Glycoprotein, Coronavirus/genetics , Transcription Factors/metabolism , Virus InternalizationABSTRACT
Anteroposterior (AP) axis extension during gastrulation requires embryonic patterning and morphogenesis to be spatiotemporally coordinated, but the underlying genetic mechanisms remain poorly understood. Here we define a role for the conserved chromatin factor Gon4l, encoded by ugly duckling (udu), in coordinating tissue patterning and axis extension during zebrafish gastrulation through direct positive and negative regulation of gene expression. Although identified as a recessive enhancer of impaired axis extension in planar cell polarity (PCP) mutants, udu functions in a genetically independent, partially overlapping fashion with PCP signaling to regulate mediolateral cell polarity underlying axis extension in part by promoting notochord boundary formation. Gon4l limits expression of the cell-cell and cell-matrix adhesion molecules EpCAM and Integrinα3b, excesses of which perturb the notochord boundary via tension-dependent and -independent mechanisms, respectively. By promoting formation of this AP-aligned boundary and associated cell polarity, Gon4l cooperates with PCP signaling to coordinate morphogenesis along the AP embryonic axis.
Subject(s)
Erythroid-Specific DNA-Binding Factors/genetics , Erythroid-Specific DNA-Binding Factors/physiology , Gene Expression Regulation, Developmental , Zebrafish Proteins/genetics , Zebrafish Proteins/physiology , Animals , Body Patterning , Cell Adhesion , Cell Communication , Chromatin/chemistry , Crosses, Genetic , Membrane Glycoproteins/physiology , Mutation , Notochord/physiology , Sequence Analysis, RNA , Signal Transduction , Xenopus , ZebrafishABSTRACT
Udu has been shown to play an essential role during blood cell development; however, its roles in other cellular processes remain largely unexplored. In addition, ugly duckling (udu) mutants exhibited somite and myotome boundary defects. Our fluorescence-activated cell sorting analysis also showed that the loss of udu function resulted in defective cell cycle progression and comet assay indicated the presence of increased DNA damage in udu(tu24) mutants. We further showed that the extensive p53-dependent apoptosis in udu(tu24) mutants is a consequence of activation in the Atm-Chk2 pathway. Udu seems not to be required for DNA repair, because both wild-type and udu embryos similarly respond to and recover from UV treatment. Yeast two-hybrid and coimmunoprecipitation data demonstrated that PAH-L repeats and SANT-L domain of Udu interacts with MCM3 and MCM4. Furthermore, Udu is colocalized with 5-bromo-2'-deoxyuridine and heterochromatin during DNA replication, suggesting a role in maintaining genome integrity.
Subject(s)
DNA Damage , Embryo, Nonmammalian/metabolism , Erythroid-Specific DNA-Binding Factors/genetics , Mutation , Zebrafish Proteins/genetics , Animals , Apoptosis/genetics , Body Patterning/genetics , Bromodeoxyuridine/metabolism , COS Cells , Cell Cycle/genetics , Checkpoint Kinase 2 , Chlorocebus aethiops , DNA Replication/drug effects , Embryo, Nonmammalian/embryology , Erythroid-Specific DNA-Binding Factors/deficiency , Erythroid-Specific DNA-Binding Factors/metabolism , Flow Cytometry , Gene Expression Regulation, Developmental , Heterochromatin/metabolism , Immunohistochemistry , In Situ Hybridization , Microscopy, Fluorescence , Protein Binding , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , Two-Hybrid System Techniques , Zebrafish/embryology , Zebrafish/genetics , Zebrafish Proteins/deficiency , Zebrafish Proteins/metabolismABSTRACT
OBJECTIVE: To characterize how interleukin-3 and erythropoietin regulate cell fate by modulating the expression of lineage-specific transcription factors. METHODS: This study analyzed mRNA and protein levels, gene transcription rates, and mRNA and protein stabilities of erythroid-specific transcription factors in lineage-restricted cells derived from the 32D cell line cultured either in interleukin-3 or erythropoietin. RESULTS: Erythroid 32D subclones expressed levels of Idl, Gata-2, and Scl comparable and levels of Eklf and Gata-1 higher than those expressed by myeloid subclones. While maintained in interleukin-3, erythroid cells remained immature despite their high expression of Gata-1, Gata-2, Scl, Eklf, and Idl. Switching the erythroid cells to erythropoietin induced cell maturation (within 48 hours) and reduced expression of Gata-2 and Idl (in 24 hours) but did not alter the expression of Gata-1. The effects of interleukin-3 were mostly mediated by increases in transcription rates (Scl and Gata-2), and that of erythropoietin was apparently due to increased mRNA and protein (Gata-1, Scl, and Eklf) stability. In particular, erythropoietin increased the stability of the processed and transcriptionally more active form of GATA-1 protein. CONCLUSIONS: These results suggest that interleukin-3 and erythropoietin cooperate to establish the lineage-specific transcription factor milieu of erythroid cells: interleukin-3 regulates mainly gene transcription and erythropoietin consistently increases mRNA and protein stability.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Erythroid Cells/drug effects , Erythroid-Specific DNA-Binding Factors/metabolism , Erythropoietin/physiology , Inhibitor of Differentiation Protein 1/metabolism , Interleukin-3/physiology , Proto-Oncogene Proteins/metabolism , Animals , Basic Helix-Loop-Helix Transcription Factors/drug effects , Basic Helix-Loop-Helix Transcription Factors/genetics , Cell Differentiation/drug effects , Cell Differentiation/genetics , Cell Line , Erythroid Cells/cytology , Erythroid-Specific DNA-Binding Factors/drug effects , Erythroid-Specific DNA-Binding Factors/genetics , Erythropoietin/pharmacology , Gene Expression Profiling , Inhibitor of Differentiation Protein 1/drug effects , Inhibitor of Differentiation Protein 1/genetics , Interleukin-3/pharmacology , Mice , Phenotype , Proto-Oncogene Proteins/drug effects , Proto-Oncogene Proteins/genetics , RNA, Messenger/drug effects , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Structure-Activity Relationship , T-Cell Acute Lymphocytic Leukemia Protein 1 , Time FactorsABSTRACT
Hematopoiesis is a complex process which gives rise to all blood lineages in the course of an organism's lifespan. However, the underlying molecular mechanism governing this process is not fully understood. Here we report the isolation and detailed study of a newly identified zebrafish ugly duckling (Udu) mutant allele, Udu(sq1). We show that loss-of-function mutation in the udu gene disrupts primitive erythroid cell proliferation and differentiation in a cell-autonomous manner, resulting in red blood cell (RBC) hypoplasia. Positional cloning reveals that the Udu gene encodes a novel factor that contains 2 paired amphipathic alpha-helix-like (PAH-L) repeats and a putative SANT-L (SW13, ADA2, N-Cor, and TFIIIB-like) domain. We further show that the Udu protein is predominantly localized in the nucleus and deletion of the putative SANT-L domain abolishes its function. Our study indicates that the Udu protein is very likely to function as a transcription modulator essential for the proliferation and differentiation of erythroid lineage.
Subject(s)
Erythroid Precursor Cells/cytology , Erythroid-Specific DNA-Binding Factors/physiology , Erythropoiesis , Transcription Factors/physiology , Zebrafish Proteins/physiology , Animals , Cell Differentiation , Cell Proliferation , Erythrocytes, Abnormal/pathology , Erythroid Cells/cytology , Erythroid-Specific DNA-Binding Factors/genetics , Mutation , Zebrafish , Zebrafish Proteins/geneticsABSTRACT
Sex determination is the process deciding the sex of a developing embryo. This is usually determined genetically; however it is a delicate process, which in many cases can be influenced by environmental factors. The mechanisms controlling zebrafish sex determination and differentiation are not known. To date no sex linked genes have been identified in zebrafish and no sex chromosomes have been identified. However, a number of genes, as presented here, have been linked to the process of sex determination or differentiation in zebrafish. The zebrafish FTZ-F1 genes are of central interest as they are involved in regulating interrenal development and thereby steroid biosynthesis, as well as that they show expression patterns congruent with reproductive tissue differentiation and function. Zebrafish can be sex reversed by exposure to estrogens, suggesting that the estrogen levels are crucial during sex differentiation. The Cyp19 gene product aromatase converts testosterone into 17 beta-estradiol, and when inhibited leads to male to female sex reversal. FTZ-F1 genes are strongly linked to steroid biosynthesis and the regulatory region of Cyp19 contains binding sites for FTZ-F1 genes, further linking FTZ-F1 to this process. The role of FTZ-F1 and other candidates for zebrafish sex determination and differentiation is in focus of this review.
