In vertebrates, six GATA transcription factors, GATA1 through GATA6, have been identified and GATA1-3 is known to be involved in hematopoietic developments, while GATA4-6 play roles in cardiac and endoderm developments. Recently, we and others have found that GATA2 and GATA3 found in the trophectoderm plays a role in gene expression specific to this cell type, but GATA4-6 have not been well characterized in early embryonic developments. Using quantitative polymerase chain reaction (qPCR) and in situ hybridization, we examined the expression of GATA4, 5 and 6 messenger RNAs (mRNAs) in ovine conceptuses and uteri during the peri-implantation period. In ovine conceptuses, GATA4, 5 and 6 transcripts were present on days 15, 17 and 21 (day 0 = day of mating), and high GATA5 and 6 mRNAs were found on day 21, most of which were localized in the trophectoderm and endoderm. Moreover, minute and substantial GATA4 and 5 mRNAs were found in days 15 and 21 uterine endometria, respectively. Increase in GATA4-6 transcripts in day 21 uteri indicates that in addition to GATA1-3, GATA4-6 may also play a potentially novel role in the development of ovine trophectoderm, endoderm and/or uterine endometria following conceptus attachment to the uterine epithelium.

Previously, we reported that although the HSPC frequency in bone marrow cells (BMC) was comparable between β2-/- and β2+/+ mice, transplantation of β2-/- BMC into lethally irradiated CD45.1 recipient resulted in more myeloid cell production than β2+/+ BMC. The objective of this study is to address if integrin β2 deficiency skews granulocyte/macrophage progenitor (GMP) proliferation. FACS analysis demonstrated that GMP frequency and cell number were higher and megakaryocyte/erythrocyte progenitor frequency and cell number were lower in β2-/- mice than β2+/+ mice. However, the common myeloid progenitors (CMP) frequency and cell number were similar between the two groups. The increased GMP number was due to GMP proliferation as evidenced by the percentage of BrdU-incorporating GMP. Whole genome transcriptome analysis identified increased FcεRIα expression in β2-/- CMP compared to β2+/+ CMP. FcεRIα expression on β2-/- GMP was detected increased in β2-/- mice by qRT-PCR and FACS. Although transplantation of FcεRIα^hi GMP or FcεRIα^lo GMP into lethally irradiated CD45.1 recipient resulted in comparable myeloid cell production, transplantation of β2 deficient FcεRIα^hi GMP generated more myeloid cells than β2+/+ FcεRIα^hi GMP. GATA2 expression was increased in β2-/- GMP. Using a luciferase reporter assay, we demonstrated that mutation of the GATA2 binding site in the FcεRIα promoter region diminished FcεRIα transcription. In vitro, the addition of IgE, the ligand of FcεRIα, promoted GMP expansion, which was abrogated by inhibition of JNK phosphorylation. Integrin β2 deficiency promoted GMP proliferation and myeloid cell production, which was mediated via FcεRIα/IgE-induced JNK phosphorylation in GMP. Stem Cells 2019;37:430-440.

Purpose: Mesenchymal stem cells (MSCs) release hematopoietic cytokines, growth factors, and Microvesicles (MVs) supporting the hematopoietic stem cells (HSCs). MVs released from various cells, playing a crucial role in biological functions of their parental cells. MSC-derived MVs contain microRNAs and proteins with key roles in the regulation of hematopoiesis. Umbilical cord blood (UCB) is a source for transplantation but the long-term recovery of platelets is a main problem. Therefore, we intend to show that MSC-MVs are able to improve the differentiation of UCB-derived CD34⁺ cells to megakaryocyte lineage. Methods: In this descriptive study, MSCs were cultured in DMEM to collect the culture supernatant, which was ultracentrifuged for the isolation of MVs. HSCs were isolated from UCB using MACS method and cultured in IMDM supplemented with cytokines and MVs in three different conditions. Megakaryocyte differentiation was evaluated through the expression of specific markers and genes after 72 hours, and the data was analyzed by t test (P<0.05). Results: The expression of specific megakaryocyte markers (CD41 and CD61) in the presence of different concentrations of MSC-MVs did not show any significant difference. Also, the expression of specific genes of megakaryocyte lineage was compared with control group. The expression of GATA2 and c-Mpl was significantly increased, GATA1 was not significantly decreased, and FLI1 was significantly decreased. Conclusion: MSC-MVs could improve the expression of specific megakaryocyte genes; however, there was no significant expression of CD markers. Further studies, including the evaluation of late stages of megakaryocyte differentiation, are required to evaluate platelet production and shedding.

The neurobeachin-like 2 protein (Nbeal2) belongs to the family of beige and Chediak-Higashi (BEACH) domain proteins. Loss-of-function mutations in the human NBEAL2 gene or Nbeal2 deficiency in mice cause gray platelet syndrome, a bleeding disorder characterized by macrothrombocytopenia, splenomegaly, and paucity of α-granules in megakaryocytes and platelets. We found that in mast cells, Nbeal2 regulates the activation of the Shp1-STAT5 signaling axis and the composition of the c-Kit/STAT signalosome. Furthermore, Nbeal2 mediates granule formation and restricts the expression of the transcription factors, IRF8, GATA2, and MITF as well as of the cell-cycle inhibitor p27, which are essential for mast cell differentiation, proliferation, and cytokine production. These data demonstrate the relevance of Nbeal2 in mast cells above and beyond granule biosynthesis.

Objective: To analyze the effect of clinical features, routine laboratory examination and related gene mutation on the OS of patients with myelodysplastic syndrome (MDS) after hematopoietic stem cell transplantation (HSCT).

Methods: 121 patients diagnosed as MDS and underwent hematopoietic stem cell transplantation in the First Affiliated Hospital of Soochow University from October 2013 to August 2018 were selected. Basic information of the patients was collected, and blood cells, bone marrow blasts at initial diagnosis, chromosomal karyotypes and gene mutations of the patients were detected.The effect of different factors on overall survival (OS) was analyzed by statistical method.

Results: Kaplan-Meier univariate analysis shows that OS was significanly different among different age groups. The 3-year OS rate of patients aged 0-29 years was (83.3±7.7) %, the 3-year OS rate in patients aged 30-49 years was (58.1±7.7 %), and the 3-year OS rate of patients aged 50-69 years was (31.0±22.6) %, which was statistically different (P＜0.05) between different groups. There were also significant differences in OS among patients with different transplantation types. 3-year OS rate: HLA-matched sibling HSCT＞unrelated HLA-matched HSCT＞haploidentical HSCT＞micro HSCT. The OS rate of patients with bone marrow blasts≥10% seems lower than blasts＜10%, but there was no statistical difference.The 3-year OS rate of patients with chromosomal karyotype complex abnormality was (47.7±11.5) %, and that of patients without complex abnormality was (80±4.2) % which was statistical difference (P＜0.05). Patients with DNMT3A, NRAS, TP53 and GATA2 mutations had shorter OS time compared with patients without mutation of these genes, which shows statistically significant (P＜0.05). COX multivariate analysis showed that age, chromosome karyotype, DNMT3A, TET2, GATA2 and NRAS were the independent factors influencing OS of patients after HSCT, with statistically significant difference.

Conclusion: age of patients, donor selection of HSCT, chromosome karyotype, DNMT3A, NRAS, TP53, GATA2 and TET2 gene mutations are all independent factors affecting the OS of patients after HSCT. Therefore, the assessment of the OS of MDS patients with transplantation requires comprehensive consideration.

题目: 影响造血干细胞移植的骨髓增生异常综合征患者 生存期的因素分析.

目的: 分析进行造血干细胞移植的骨髓增生异常综合征（myelodysplastic syndrome，MDS）患者的临床特点、常规实验室检查及相关基因突变对总生存期（overall survival，OS）的影响.

方法: 选取并回顾性分析2013年10月至2018年8月在苏州大学附属第一医院诊断MDS并进行造血干细胞移植（hematopoietic stem cell transplantation，HSCT）的患者121例，收集患者基本信息，并检测患者血细胞、初诊骨髓原始细胞、染色体核型及基因突变情况。用统计学方法分析不同因素对生存期的影响.

结果: Kaplan-Meier单因素分析发现，不同年龄段之间OS有明显差异，患者年龄越高，3年OS率越低，0-29岁患者3年OS率为（83.3±7.7）% ；30-49岁患者3年OS率为（58.1±7.7）%；50-69岁患者均3年OS率为（31.0±22.6）%, 且各组之间OS率统计学差异显著（P＜0.05）。不同移植类型对患者OS也有显著影响，3年OS率：同胞全相合HSCT＞非血缘全相合HSCT＞单倍体相合HSCT＞微移植。骨髓原始细胞≥10%的患者OS率低于＜10%的患者，但差异没有统计学意义。染色体核型复杂异常患者3年OS率为（47.7±11.5）%，非复杂异常患者的3年OS率为（80±4.2）%（P＜0.05）。DNMT3A、NRAS、TP53、GATA2基因非突变患者比基因突变的患者移植后OS期长，差异有统计学意义（P＜0.05）。COX多因素分析结果显示，年龄、染色体核型、DNMT3A、TET2、GATA2、NRAS对MDS移植后OS有独立影响，且差异具有统计学意义.

结论: 患者年龄，造血干细胞移植供体选择，患者初诊染色体核型，DNMT3A、NRAS、TP53、GATA2、TET2基因突变均是影响MDS移植后生存期的独立因素。因此对MDS移植患者生存期的评估需要综合多方面考虑.

The development of macrophages is a highly regulated process requiring coordination amongst transcription factors. The presence/absence, relative levels, antagonism, or synergy of all transcription factors involved is critical to directing lineage cell fate and differentiation. While relative levels of many key myeloid transcription factors have been determined in mammalian macrophage differentiation, a similar set of studies have yet to be conducted in a teleost system. In this study, we report on the mRNA levels of transcription factors (cebpa, cjun, cmyb, egr1, gata1, gata2, gata3, lmo2, mafb, pax5, pu.1 and runx1) in sorted goldfish progenitor cells, monocytes, and macrophages from primary kidney macrophage cultures. The mRNA levels of runx1 and pu.1 were significantly higher, gata3 and pax5 mRNA levels were lower, in monocytes compared to progenitors, and the mRNA levels of cjun, egr1, gata2, gata3, mafb and pax5 were significantly decreased in macrophages compared to progenitor cells. The relative mRNA levels of the interferon regulatory factor family of transcription factors, irf1, irf2, irf5, irf7, irf8 and irf9 in sorted progenitors, monocytes and macrophages were also measured. In contrast to other irf family transcription factors examined, irf8 mRNA levels were increased in monocytes compared to progenitors by greater than three-fold, suggesting that irf8 is important for monopoiesis. Lastly, we show the differential regulation of myeloid transcription factor mRNA levels in sorted progenitor cells from 1, 2, or 3-day old cultures in response to the recombinant goldfish growth factors, rgCSF-1 and rgKITLA.

We describe a patient, who developed during the first week of life petechiae and hematomas caused by severe thrombocytopenia and gastrointestinal bleeding due to multiple small gastric hemangiomata. Bone marrow examination showed hypermegakaryocytosis and dysmegakaryopoiesis. Alloimmune thrombocytopenia was excluded. Only 3 y later, platelet counts normalized and bleedings disappeared but small skin hemangiomata remained. Electron microscopy showed enlarged round platelets with a paucity of alpha granules similar as in GATA1-deficient patients but no GATA1 mutation was found. Immunoblot analysis showed a strong interaction between patient Igs and recombinant GATA1, GATA2, and the N finger (Nf) of GATA1. The lymphocyte transformation test with recombinant GATA1Nf was positive. In vitro culturing of normal CD34 cells with purified patient Igs showed a decreased number of megakaryocyte colonies but an increased overall size of the colonies compared with control Igs. Mice injected with patient Igs showed a reduced platelet count compared with mice injected with control Igs. Thrombopoiesis was also reduced after injection of patient Igs in transgenic zebrafish compared with control Igs. In conclusion, this study is the first report of an anti-GATA1 autoantibody leading to severe thrombocytopenia and gastrointestinal bleeding from multiple pinpoint hemangiomata.

The GATA family of transcription factors are zinc finger (ZF) DNA-binding proteins that regulate transcription during development and cell differentiation. GATA2 plays an essential role in the regulation of hematopoiesis. As a result, mutations in this gene or alterations in its expression level or function have been linked to a variety of human hematologic disorders. In this review, we summarize the findings and developments over the recent years regarding the clinical correlations and functional properties of distinct GATA2 mutations in hematopoietic malignancies, with particular focus on the mutational hotspots in the ZF domains.

The cellular and molecular mechanisms underlying specification of human hematopoietic stem cells (HSCs) remain elusive. Strategies to recapitulate human HSC emergence in vitro are required to overcome limitations in studying this complex developmental process. Here, we describe a protocol to generate hematopoietic stem and progenitor-like cells from human dermal fibroblasts employing a direct cell reprogramming approach. These cells transit through a hemogenic intermediate cell-type, resembling the endothelial-to-hematopoietic transition (EHT) characteristic of HSC specification. Fibroblasts were reprogrammed to hemogenic cells via transduction with GATA2, GFI1B and FOS transcription factors. This combination of three factors induced morphological changes, expression of hemogenic and hematopoietic markers and dynamic EHT transcriptional programs. Reprogrammed cells generate hematopoietic progeny and repopulate immunodeficient mice for three months. This protocol can be adapted towards the mechanistic dissection of the human EHT process as exemplified here by defining GATA2 targets during the early phases of reprogramming. Thus, human hemogenic reprogramming provides a simple and tractable approach to identify novel markers and regulators of human HSC emergence. In the future, faithful induction of hemogenic fate in fibroblasts may lead to the generation of patient-specific HSCs for transplantation.

OBJECTIVE

Peroxisome-proliferator activated receptor γ (PPARγ) plays a pivotal role in inhibition of hepatic stellate cell (HSC) activation, a key step for liver fibrogenesis. Adipocyte-derived hormone leptin has been shown to promote liver fibrosis in murine and human. PPARγ includes two subtypes, PPARγ1 and PPARγ2. Our previous study indicated that leptin down-regulated PPARγ1 expression in HSCs. The aim of this study was to investigate the effect of leptin on PPARγ2 expression and the underlying mechanisms in HSCs.

METHODS

Real-time PCR and western blot analyses were used to examine gene expression. The promoter activities were detected by luciferase assay.

RESULTS

Leptin reduced PPARγ2 expressions at promoter level, mRNA level, and protein level in HSCs, which required β-catenin, p38 mitogen-activated protein kinase, and delta-like homolog1 (DLK1) signaling pathways. Leptin induced GATA binding protein 2 (GATA2) expression through DLK1 pathway and GATA2 reduced PPARγ2 expression. Ectopic expression of PPARγ2 reduced the protein levels of α-smooth muscle actin and α1(I)collagen in HSCs.

CONCLUSIONS

Since obese patients, often accompanied by hyperleptinemia, are more prone to liver fibrosis, the data from this study might have potential implications for clarifying the mechanisms for liver fibrogenesis in obese patients with hyperleptinemia.

Mycobacterium abscessus (M. abscessus) infection is resistant to multi-antibacterial treatment, and surgical resection is often recommended. We report a case of M. abscessus infection in a young patient suspected of having a GATA2 mutation. A 19-year-old woman with a medical history of severe sinusitis and a family history of non-tuberculous mycobacteriosis presented at our hospital. M. abscessus was confirmed by sputum culture. The patient received multidrug therapy, including clarithromycin. CT scan demonstrated bronchodilation and capacity decrease due to non-obstructive atelectasis in the middle lobe. We performed thoracoscopic resection without complications. Congenital immunodeficiency was suspected given the patient's past medical and family history. The result of lymphocyte subset analysis revealed a GATA2 mutation, but no genetic mutation was detected by a next-generation sequencer. The patient followed a good clinical course. This paper reports the successful treatment of an M. abscessus infection and the importance of checking the genetic background of young patients.

Keywords: GATA2 mutation; Mycobacterium abscessus; surgical resection.

Background/objective: The role of gene expression changes in acne patients treated by oral isotretinoin (ISO) and in influencing the ISO therapeutic effects is still unclear. In this study, we investigated the gene profiles of patients with severe acne who responded variously to ISO therapy.

Methods: The peripheral blood of 113 acne vulgaris patients (Pillsbury IV grade) was collected before treatment. After 8 weeks of oral ISO, nine acne patients were selected and divided into the following groups. A: effectively treated by ISO, group B: ineffectively treated by ISO, group C: ISO-induced acne flare-up, and 3 healthy subjects were included as control group D. The peripheral blood of patients pre- and post-treatment was subjected to high-throughput RNA sequencing technology and bioinformatics analysis of the separate groups (n = 3). The candidate genes were validated by qRT-PCR.

Results: Comparing pre- and post-oral ISO treatment, gene expression was changed as 39 genes in ISO-effective group, 345 genes in ISO-ineffective group, and 57 genes in ISO-induced acne flare-up group. Comparing the ISO-induced acne flare-up group with healthy control subjects revealed 34 upregulated genes and 23 downregulated genes, while comparing the ISO-induced acne flare-up group with ISO-ineffective patients identified 1835 changed genes. Expression of GATA2 (2.73 fold, P=0.024512), C4BPA (35.87 folds, P=0.038073), and CCR5 (2.48 folds, P=0.004681) increased in the ISO-induced acne flare-up patients. Meanwhile, the expression of DEFA3 (0.18 fold, P=0.041934), ELANE (0.14 fold, P=0.030767), MMP9 (0.41 fold, P=0.013383), and RPS4Y1 (0.00018 fold, P=0.000986) decreased when compared with ISO-ineffective patients.

Conclusion: Oral ISO treatment could temporarily alter gene expression in acne patients. ISO therapeutic mechanisms were involved, not only in regulating the inflammatory reaction but also in the process of DNA repair. GATA2, C4BPA, CCR5, DEFA3, ELANE, MMP9, and RPS4Y1 might be susceptible to genes that could participate in the ISO-induced aggravation of acne.

Keywords: acne vulgaris; gene profile; isotretinoin; mechanisms; treatment.

Congenital neutropenia (CN) is a rare inherited disease that results in recurrent, life-threatening bacterial infections due to a deficiency of mature neutrophils. They are usually caused by heterozygous ELANE mutations although mutations in other genes like HAX-1, G6PC3 and GFI1 have also been reported. Identifying the causative mutation aids in the establishment of diagnosis and rules out other secondary causes of neutropenia like autoimmune cytopenia and evolving aplasia. We aimed to identify the molecular defects in CN patients who had no mutations in ELANE gene, by next generation sequencing (NGS) targeting a customised panel of genes.

DNA samples were sequenced with an Illumina NextSeq sequencer using an in-house customised panel of genes at ≥100× depth. Bioinformatics analysis was carried out and the pathogenic variants were identified using a stepwise filtering and analysis strategy. Specific mutations identified were subsequently validated by Sanger sequencing.

RESULTS

The pathogenic variants identified in the study includes previously reported variants in SBDS (compound heterozygous c.258+2T>C and c.1A>T), GATA2 (heterozygous c.1186C>T) and novel variants in WAS (hemizygous c.812T>C), JAGN1 (homozygous c.70G>A) and RTEL1 (heterozygous c.2893G>C) genes.

CONCLUSION

This study highlights that the absence of ELANE mutations does not rule out the diagnosis of CN and this NGS based approach with a customised panel will help in diagnostic confirmation in such patients. The early onset of the disease, clinical severity and associated high risk of malignant transformation in CN strongly suggests the need for early diagnosis and therapeutic intervention.

Embryonic stem cells (ESCs) have been shown to have an ability to form a large number of functional endothelial cells in vitro, but generating organ-specific endothelial cells remains a challenge. Sonic hedgehog (SHH) pathway is one of the crucial developmental pathways that control differentiation of many embryonic cell types such as neuroectodermal, primitive gut tube and developing limb buds; SHH pathway is important for functioning of adult cell of skin, bone, liver as well as it regulates haematopoiesis. Misregulation of SHH pathway leads to cancers such as hepatic, pancreatic, basal cell carcinoma, medulloblastoma, etc. However, its role in differentiation of human ESCs into endothelial cells has not been completely elucidated. Here, we examined the role of SHH signalling pathway in endothelial differentiation of hESCs by growing them in the presence of an SHH agonist (purmorphamine) and an SHH antagonist (SANT-1) for a period of 6 days. Interestingly, we found that activation of SHH pathway led to a higher expression of set of transcription factors such as BRACHYURY, GATA2 and RUNX1, thus favouring hemogenic endothelium; whereas inhibition of SHH pathway led to a reduced expression of set of markers such as RUNX1 and BRACHURY, and an increased expression of set of markers - NFATC1, c-KIT, GATA4, CD31 & CD34, thus favouring endocardiogenic endothelium. The results of this study have revealed the previously unreported deterministic role of SHH pathway in specification of endothelial cells differentiated from human ESCs into hemogenic vs. endocardiogenic lineage; this finding could have major implications for clinical applications.

Keywords: Differentiation; Endocardiogenic endothelium; Endothelial cells; Hemogenic endothelium; Human embryonic stem; Sonic hedgehog pathway.

Neurological diseases (NDs) are progressive disorders, the progression of which can be significantly affected by a range of common diseases that present as comorbidities. Clinical studies, including epidemiological and neuropathological analyses, indicate that patients with type 2 diabetes (T2D) have worse progression of NDs, suggesting pathogenic links between NDs and T2D. However, finding causal or predisposing factors that link T2D and NDs remains challenging. To address these problems, we developed a high-throughput network-based quantitative pipeline using agnostic approaches to identify genes expressed abnormally in both T2D and NDs, to identify some of the shared molecular pathways that may underpin T2D and ND interaction. We employed gene expression transcriptomic datasets from control and disease-affected individuals and identified differentially expressed genes (DEGs) in tissues of patients with T2D and ND when compared to unaffected control individuals. One hundred and ninety seven DEGs (99 up-regulated and 98 down-regulated in affected individuals) that were common to both the T2D and the ND datasets were identified. Functional annotation of these identified DEGs revealed the involvement of significant cell signaling associated molecular pathways. The overlapping DEGs (i.e., seen in both T2D and ND datasets) were then used to extract the most significant GO terms. We performed validation of these results with gold benchmark databases and literature searching, which identified which genes and pathways had been previously linked to NDs or T2D and which are novel. Hub proteins in the pathways were identified (including DNM2, DNM1, MYH14, PACSIN2, TFRC, PDE4D, ENTPD1, PLK4, CDC20B, and CDC14A) using protein-protein interaction analysis which have not previously been described as playing a role in these diseases. To reveal the transcriptional and post-transcriptional regulators of the DEGs we used transcription factor (TF) interactions analysis and DEG-microRNAs (miRNAs) interaction analysis, respectively. We thus identified the following TFs as important in driving expression of our T2D/ND common genes: FOXC1, GATA2, FOXL1, YY1, E2F1, NFIC, NFYA, USF2, HINFP, MEF2A, SRF, NFKB1, USF2, HINFP, MEF2A, SRF, NFKB1, PDE4D, CREB1, SP1, HOXA5, SREBF1, TFAP2A, STAT3, POU2F2, TP53, PPARG, and JUN. MicroRNAs that affect expression of these genes include mir-335-5p, mir-16-5p, mir-93-5p, mir-17-5p, mir-124-3p. Thus, our transcriptomic data analysis identifies novel potential links between NDs and T2D pathologies that may underlie comorbidity interactions, links that may include potential targets for therapeutic intervention. In sum, our neighborhood-based benchmarking and multilayer network topology methods identified novel putative biomarkers that indicate how type 2 diabetes (T2D) and these neurological diseases interact and pathways that, in the future, may be targeted for treatment.

Lim domain only 2 (LMO2) is a transcriptional co-factor required for angiogenesis and the specification of haematopoietic cells during development. LMO2 is widely expressed within haematopoiesis with the exception of T-cells. Failure to downregulate LMO2 during T-cell maturation leads to leukaemia, thus underlining the critical nature of context-dependent regulation of LMO2 expression. We previously identified a distal regulatory element of LMO2 (element -25) that cooperates with the proximal promoter in directing haematopoietic expression. Here we dissected the functional activity of element -25 and showed it to consist of two modules that conferred independent and cell-type specific activities: a 3' myeloid enhancer and a 5' T-cell repressor. The myeloid enhancer was bound by GATA2 in progenitors and its activity depended on a highly conserved GATA motif, whereas the T-cell repressor moiety of element -25 was bound by the Core Binding Factor in T-cells and its repressive activity depended on a highly conserved RUNT motif. Since the myeloid enhancer and nearby downstream region is recurrently involved in oncogenic translocations, our data suggest that the -25 enhancer region provides an open chromatin environment prone to translocations, which in turn cause aberrant LMO2 expression in T-cells due to the removal of the adjacent T-cell repressor.

Hemocytes, the cellular component of invertebrate hemolymph, are essential for invertebrate immunity, but the hematopoiesis and regulation mechanism are still largely unknown. In the present study, a conserved hematopoietic transcription factor Cg-GATA2/3 was identified in Pacific oyster Crassotrea gigas, which was evolutionarily close to the vertebrate GATA1/2/3. Cg-GATA2/3 was mainly distributed in the immune organs, such as gill, hemocytes, and mantle. After Cg-GATA2/3 was interferenced by dsRNA, the mRNA expressions of hemocytes specific gene (EcSOD) and hematopoietic transcription factor (C-Myb) were all significant down-regulated, and the hemocyte renewal rates also decreased both in hemolymph and gill. During the larval developmental stages, the mRNA transcripts of Cg-GATA2/3 increased immediately after fertilization and kept a high level during blastula and early trochophore larvae stage (4-10 hpf, hours post fertilization), then decreased sharply in early D-veliger larvae stage (15 hpf). Whole-mount immunofluorescence assay further revealed that the abundant immunoreactivity of Cg-GATA2/3 was distributed in the whole body of blastula and gastrula embryos, while specialized gradually to a ring structure around the dorsal region in trochophore larvae. In the D-veliger and umbo larvae, scattered positive signals appeared in the specific sinus structure on the dorsal side and velum region. These results demonstrated that Cg-GATA2/3 was a hematopoietic lineage-specific transcription factor to regulate the hemocyte production, and it could also be used as hematopoietic specific marker to trace potential developmental events of hematopoiesis during ontogenesis of oyster.

Background. Following the two rounds of whole-genome duplication that occurred during deuterostome evolution, a third genome duplication occurred in the lineage of teleost fish and is considered to be responsible for much of the biological diversification within the lineage. GATA2, a member of GATA family of transcription factors, is an important regulator of gene expression in hematopoietic cell in mammals, yet the role of this gene or its putative paralogs in ray-finned fishes remains relatively unknown. Methods. In this study, we attempted to identify GATA2 sequences from the transcriptomes and genomes of multiple teleosts using the bioinformatic tools MrBayes, MEME, and PAML. Following identification, comparative analysis of genome structure, molecular evolution rate, and expression by real-time qPCR were used to predict functional divergence of GATA2 paralogs and their relative transcription in organs of female and male tongue soles (Cynoglossus semilaevis). Results. Two teleost GATA2 genes were identified in the transcriptomes of tongue sole and Japanese flounder (Paralichthysolivaceus). Synteny and phylogenetic analysis confirmed that the two genes likely originated from the teleost-specific genome duplication . Additionally, selection pressure analysis predicted these gene duplicates to have undergone purifying selection and possible divergent new functions. This was supported by differential expression pattern of GATA2a and GATA2b observed in organs of female and male tongue soles. Discussion. Our results indicate that two GATA2 genes originating from the first teleost-specific genome duplication have remained transcriptionally active in some fish species and have likely undergone neofunctionalization. This knowledge provides novel insights into the evolution of the teleost GATA2 genes and constituted important groundwork for further research on the GATA gene family.

We aimed to identify QTL for serum electrolyte traits by performing a GWAS of calcium, chloride, sodium, potassium and magnesium ion concentrations in Chinese Holstein cattle. We detected five SNPs that had significant associations with calcium ion concentrations and identified GATA2 from Bos taurus chromosome (BTA)22 as having the highest significance. Among the genes with significant results, we speculate that TMEM123 might be related to calcium channel proteins according to the functions of the TMEM family.

Music-listening and performance have been shown to affect human gene expression. In order to further elucidate the biological basis of the effects of music on the human body, we studied the effects of music-listening on gene regulation by sequencing microRNAs of the listeners (Music Group) and their controls (Control Group) without music exposure. We identified upregulation of six microRNAs (hsa-miR-132-3p, hsa-miR-361-5p, hsa-miR-421, hsa-miR-23a-3p, hsa-miR-23b-3p, hsa-miR-25-3p) and downregulation of two microRNAs (hsa-miR-378a-3p, hsa-miR-16-2-3p) in Music Group with high musical aptitude. Some upregulated microRNAs were reported to be responsive to neuronal activity (miR-132, miR-23a, miR-23b) and modulators of neuronal plasticity, CNS myelination and cognitive functions like long-term potentiation and memory. miR-132 plays a critical role in regulating TAU protein levels and is important for preventing tau protein aggregation that causes Alzheimer's disease. miR-132 and DICER, upregulated after music-listening, protect dopaminergic neurons and are important for retaining striatal dopamine levels. Some of the transcriptional regulators (FOS, CREB1, JUN, EGR1 and BDNF) of the upregulated microRNAs were immediate early genes and top candidates associated with musical traits. BDNF and SNCA, co-expressed and upregulated in music-listening and music-performance, are both are activated by GATA2, which is associated with musical aptitude. Several miRNAs were associated with song-learning, singing and seasonal plasticity networks in songbirds. We did not detect any significant changes in microRNA expressions associated with music education or low musical aptitude. Our data thereby show the importance of inherent musical aptitude for music appreciation and for eliciting the human microRNA response to music-listening.

Keywords: Music-listening; gene regulation; microRNA; music education; musical aptitude; sequencing.

In vivo hematopoietic generation occurs in waves of primitive and definitive cell emergence. Differentiation cultures of pluripotent embryonic stem cells (ESCs) offer an accessible source of hematopoietic cells for blood-related research and therapeutic strategies. However, despite many approaches, it remains a goal to robustly generate hematopoietic progenitor and stem cells (HP/SCs) in vitro from ESCs. This is partly due to the inability to efficiently promote, enrich, and/or molecularly direct hematopoietic emergence. Here, we use Gata2Venus (G2V) and Ly6a(SCA1)GFP (LG) reporter ESCs, derived from well-characterized mouse models of HP/SC emergence, to show that during in vitro differentiation they report emergent waves of primitive hematopoietic progenitor cells (HPCs), definitive HPCs, and B-lymphoid cell potential. These results, facilitated by enrichment of single and double reporter cells with HPC properties, demonstrate that in vitro ESC differentiation approximates the waves of hematopoietic cell generation found in vivo, thus raising possibilities for enrichment of rare ESC-derived HP/SCs.

Delta-like ligand 4 (DLL4) expression in endothelial cells is intimately associated with angiogenic sprouting and vascular remodeling, but the precise mechanism of transcriptional regulation of DLL4 remains incompletely understood. Here, we showed that LIM-domain binding protein 2 (LDB2) plays an important role in regulating basal DLL4 and VEGF-induced DLL4 expression. Knockdown of LDB2 using siRNA enhanced endothelial sprouting and tubular network formation in vitro. Injection of ldb2-morpholino resulted in defective development of intersegmental vessels in zebrafish. Reduction or overexpression of LDB2 in endothelial cells decreased or increased DLL4 expression. LDB2 regulated DLL4 promoter activity by binding to its promoter region and the same promoter region was occupied and regulated by the LMO2/TAL1/GATA2 complex. Interestingly, LDB2 also mediated VEGF-induced DLL4 expression in endothelial cells. The regulation of DLL4 by the LDB2 complex provides a novel mechanism of DLL4 transcriptional control that may be exploited to develop therapeutics for aberrant vascular remodeling. [BMB Reports 2018; 51(1): 21-26].

Hormonal regulation of pituitary gonadotropin gene expression has been attributed to gonadotropin-releasing hormone (GnRH)-mediated stimulation of immediate early gene expression and gonadal steroid interactions with their respective nuclear receptors. A number of orphan nuclear receptors including steroidogenic factor 1, liver receptor homologue 1, dosage-sensitive sex reversal, adrenal hypoplasia critical region, on chromosome X, gene 1, and chicken ovalbumin upstream promoter-transcription factors I/II as well as the GATA family members, GATA2 and GATA4, have also been implicated in transcriptional regulation of the gonadotropin genes. We hypothesized that hormonally mediated changes in these latter transcription factors may provide an additional mechanism for mediating hormonal effects beyond the more classically appreciated pathways. In these studies, we demonstrate significant regulation of orphan nuclear receptor and GATA messenger RNA levels by GnRH, dihydrotestosterone, estradiol, and progesterone in both cultured primary pituitary cells and gonadotrope-derived cell line, LβT2. These results advance our understanding of the complex mechanisms by which GnRH and steroid hormones achieve precise regulation of anterior pituitary function.