KLF4 Research Articles

Antifibrotic factor KLF4 is repressed by the miR-10/TFAP2A/TBX5 axis in dermal fibroblasts: insights from twins discordant for systemic sclerosis

ObjectivesSystemic sclerosis (SSc) is a complex disease of unknown aetiology in which inflammation and fibrosis lead to multiple organ damage. There is currently no effective therapy that can halt the...

RVU120 (SEL120) CDK8/19 Inhibitor - a Drug Candidate for the Treatment of MDS Can Induce Erythroid Differentiation

Background: Myelodysplastic syndromes (MDS) are a group of malignant blood disorders characterized by ineffective hematopoiesis and cytopenias and frequent evolution to acute myeloid leukemia (AML). MDS results from the expansion of genetically and epigenetically changed clones with impaired differentiation and maturation. Primary clinical goals in MDS are to achieve remissions, alleviate symptoms associated with cytopenias, and minimize the transfusion burden. While supportive red blood cell transfusions, erythropoiesis-stimulating agents and novel targeted agents may lead to clinical improvement, an allogeneic bone marrow transplant (BMT) remains the only potential curative option for patients with MDS. RVU120 (SEL120) is a specific, selective inhibitor of CDK8 and its paralog CDK19. A first-in-human Phase Ib clinical trial with RVU120 in patients with AML or high risk (HR)-MDS is currently ongoing. Preclinical studies indicated the strong antileukemic potential of RVU120 that was often associated with the multilineage commitment of CD34+ AML cells. Moreover, RVU120 could improve proliferation and induce erythroid differentiation of CD34+ cells derived from Diamond-Blackfan anemia (DBA) patients.Aims: Primary aim was to evaluate the erythroid differentiation potential of RVU120 in primary MDS and transformed cord blood CD34+ blood cells characterized with an early block in erythroid differentiation.Methods: Efficacy and mechanism of action of RVU120 and other CDK8 inhibitors were investigated in cord blood (CB) cells transduced with TLS-ERG, a fusion gene generated from t(16;21)(p11;q22). Transformed cells displayed an increased capacity for self-renewal, proliferation, and altered erythroid differentiation. Efficacy was further tested in CD34+ cells isolated from BM of MDS patients and established MDS cell lines. Cells were treated with RVU120 and global transcriptional changes and chromatin status were analyzed by RNA-seq, ATAC-seq, and ChIP-seq. Cell cycle, proliferation, and lineage-specific markers were studied by flow cytometry in liquid and semi-solid methylcellulose-based media.Results: RVU120 treatment leads to transcriptional reprogramming of transformed CD34+ CB cells. The most profound changes included decreased CDK8 occupancy followed by increased STAT5 and RNA Polymerase II loading at transcription start site and gene bodies. RVU120 treatment transcriptionally represses multiple genes associated with hematopoietic and leukemia stem cells such as CD34, FLI1, ENG and RGS18 and importantly induce the expression of genes involved in erythroid commitment, including regulators of erythroid/megakaryocytic fate, such as RGS18, KLF1, FLI1, INHBA, GATA1/2 and hemoglobin genes. Detailed analysis of transformed CB and MDS CD34+ cells by flow cytometry at early and late time points reflected sequential changes in the expression of lineage-specific surface markers, leading to erythroid differentiation.Conclusions: Presented results indicate strong erythroid differentiation potential of RVU120 in CD34+ cells that acquired genetic abnormalities leading to arrested erythroid commitment, characteristics of many MDS and AML subtypes. Observed differentiation phenotype strikingly resembles effects of RVU120 in DBA cells caused by disruption of genes encoding ribosomal proteins. Detailed transcriptomic profiling strongly links the differentiation with enrichment of genes representing regulators of erythroid commitment and hemoglobin metabolism. Further studies are warranted to investigate the efficacy of RVU120 in chronic anemias associated with bone marrow failures in AML and MDS patients. [Display omitted] DisclosuresPakulska: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Obacz: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Goller: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Combik: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Keska: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Mazan: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Juszczynski: Ryvu Therapeutics: Current equity holder in publicly-traded company. Brzozka: Ardigen: Current Employment, Membership on an entity's Board of Directors or advisory committees; Selvita SA: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees. Dziedzic: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Angelosanto: Ryvu Therapeutics: Current Employment. Shamsili: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Rzymski: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company.

A stem cell marker KLF5 regulates CCAT1 via three-dimensional genome structure in colorectal cancer cells

BackgroundKLF5 plays a crucial role in stem cells of colorectum in cooperation with Lgr5 gene. In this study, we aimed to explicate a regulatory mechanism of the KLF5 gene product from a view of three-dimensional genome structure in colorectal cancer (CRC).MethodsIn vitro engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP)-seq method was used to identify the regions that bind to the KLF5 promoter.ResultsWe revealed that the KLF5 promoter region interacted with the KLF5 enhancer region as well as the transcription start site (TSS) region of the Colon Cancer Associated Transcript 1 (CCAT1) gene. Notably, the heterodeletion mutants of KLF5 enhancer impaired the cancer stem-like properties of CRC cells. The KLF5 protein participated in the core-regulatory circuitry together with co-factors (BRD4, MED1, and RAD21), which constructs the three-dimensional genome structures consisting of KLF5 promoter, enhancer and CCAT1 TSS region. In vitro analysis indicated that KLF5 regulated CCAT1 expression and we found that CCAT1 expression was highly correlated with KLF5 expression in CRC clinical samples.ConclusionsOur data propose the mechanistic insight that the KLF5 protein constructs the core-regulatory circuitry with co-factors in the three-dimensional genome structure and coordinately regulates KLF5 and CCAT1 expression in CRC.

High protein diet-induced metabolic changes are transcriptionally regulated via KLF15-dependent and independent pathways

High protein diet (HPD) is an affordable and positive approach in prevention and treatment of many diseases. It is believed that transcriptional regulation is responsible for adaptation after HPD feeding and Kruppel-like factor 15 (KLF15), a zinc finger transcription factor that has been proved to perform transcriptional regulation over amino acid, lipid and glucose metabolism, is known to be involved at least in part in this HPD response. To gain more insight into molecular mechanisms by which HPD controls expressions of genes involved in amino acid metabolism in the liver, we performed RNA-seq analysis of mice fed HPD for a short period (3 days). Compared to a low protein diet, HPD feeding significantly increased hepatic expressions of enzymes involved in the breakdown of all the 20 amino acids. Moreover, using KLF15 knockout mice and in vivo Ad-luc analytical system, we were able to identify Cth (cystathionine gamma-lyase) as a new target gene of KLF15 transcription as well as Ast (aspartate aminotransferase) as an example of KLF15-independent gene despite its remarkable responsiveness to HPD. These findings provide us with a clue to elucidate the entire transcriptional regulatory mechanisms of amino acid metabolic pathways.

Loss of proximal tubular transcription factor Krüppel-like factor 15 exacerbates kidney injury through loss of fatty acid oxidation

Loss of fatty acid β-oxidation (FAO) in the proximal tubule is a critical mediator of acute kidney injury and eventual fibrosis. However, transcriptional mediators of FAO in proximal tubule injury remain understudied. Krüppel-like factor 15 (KLF15), a highly enriched zinc-finger transcription factor in the proximal tubule, was significantly reduced in proximal tubule cells after aristolochic acid I (AAI) treatment, a proximal tubule-specific injury model. Proximal tubule specific knockout of Klf15 exacerbated proximal tubule injury and kidney function decline compared to control mice during the active phase of AAI treatment, and after ischemia-reperfusion injury. Furthermore, along with worsening proximal tubule injury and kidney function decline, knockout mice exhibited increased kidney fibrosis as compared to control mice during the remodeling phase after AAI treatment. RNA-sequencing of kidney cortex demonstrated increased transcripts involved in immune system and integrin signaling pathways and decreased transcripts encompassing metabolic pathways, specifically FAO, and PPARα signaling, in knockout versus control mice after AAI treatment. In silico and experimental chromatin immunoprecipitation studies collectively demonstrated that KLF15 occupied the promoter region of key FAO genes, CPT1A and ACAA2, in close proximity to transcription factor PPARα binding sites. While the loss of Klf15 reduced the expression of Cpt1a and Acaa2 and led to compromised FAO, induction of KLF15 partially rescued loss of FAO in AAI-treated cells. Klf15, Ppara, Cpt1a, and Acaa2 expression was also decreased in other mouse kidney injury models. Tubulointerstitial KLF15 independently correlated with eGFR, PPARA and CPT1A appearance in expression arrays from human kidney biopsies. Thus, proximal tubule-specific loss of Klf15 exacerbates acute kidney injury and fibrosis, likely due to loss of interaction with PPARα leading to loss of FAO gene transcription.

Isoform-specific effects of transcription factor TCFL5 on the pluripotency-related genes SOX2 and KLF4 in colorectal cancer development.

Colorectal cancer (CRC) is a very common life‐threatening malignancy. Transcription factor‐like 5 (TCFL5) has been suggested to be involved in CRC. Here, we describe the expression of four alternative transcripts of TCFL5 and their relevance in CRC. Complete deletion of all isoforms drastically decreased pro‐tumoural properties such as spheroids formation and in vivo tumour growth, although increased migration in CRC cell lines. Overexpression of the two main isoforms, TCFL5_E8 and CHA, had opposite effects: TCFL5_E8 reduced proliferation and spheroids formation, while CHA increased them. TCFL5_E8 reduced in vivo tumour formation, while CHA had no effect. In addition, TCFL5_E8 and CHA have different roles in the regulation of the pluripotency‐related genes SOX2 and KLF4. Both isoforms bind directly to their promoters; however, TCFL5_E8 induced SOX2 and reduced KLF4 mRNA levels, whereas CHA did the opposite. Together, our results show that TCFL5 plays an important role in the development of CRC, being however isoform‐specific. This work also points to the need to analyse separately TCFL5 isoforms in cancer, due to their different and opposite functions.

Multi-species single-cell transcriptomic analysis of ocular compartment regulons

The retina is a widely profiled tissue in multiple species by single-cell RNA sequencing studies. However, integrative research of the retina across species is lacking. Here, we construct the first single-cell atlas of the human and porcine ocular compartments and study inter-species differences in the retina. In addition to that, we identify putative adult stem cells present in the iris tissue. We also create a disease map of genes involved in eye disorders across compartments of the eye. Furthermore, we probe the regulons of different cell populations, which include transcription factors and receptor-ligand interactions and reveal unique directional signalling between ocular cell types. In addition, we study conservation of regulons across vertebrates and zebrafish to identify common core factors. Here, we show perturbation of KLF7 gene expression during retinal ganglion cells differentiation and conclude that it plays a significant role in the maturation of retinal ganglion cells.

Krüppel-Like Factor 15 Reduces Ischemia-Induced Apoptosis Involving Regulation of p38/MAPK Signaling.

Cardiomyocyte apoptosis is a characteristic of a variety of cardiac diseases, including myocardial infarction (MI). Krüppel-like factor 15 (KLF15) is a transcription factor of Krüppel family that plays an important part in cardiovascular diseases. However, the function and the underlying mechanism of KLF15 in MI remain unknown. The expression of KLF15 was downregulated both in ischemic myocardium of MI mice model and hypoxia-treated neonatal rat ventricular myocytes (NRVCs). KLF15 overexpression mediated by adeno-associated virus significantly abrogated the ischemia-induced cardiac dysfunction, increased the survival rate, and reduced infarct size after MI. Meanwhile, KLF15 overexpression dramatically reduced the myocardial apoptosis, regulated apoptosis-related genes, such as Bcl2 and Bax, diminished the activities of caspase-9/3, and inactivated p38/MAPK signaling in the border zone. Similar results were observed in NRVCs exposed to hypoxia. We demonstrated for the first time that KLF15 overexpression could reduce cardiomyocyte apoptosis and improve cardiac dysfunction in MI mice at least partially by inhibiting p38/MAPK signaling pathway.

Intermittent High Glucose Elevates Nuclear Localization of EZH2 to Cause H3K27me3-Dependent Repression of KLF2 Leading to Endothelial Inflammation

Epigenetic mechanisms have emerged as one of the key pathways promoting diabetes-associated complications. Herein, we explored the role of enhancer of zeste homolog 2 (EZH2) and its product histone 3 lysine 27 trimethylation (H3K27me3) in high glucose-mediated endothelial inflammation. To examine this, we treated cultured primary endothelial cells (EC) with different treatment conditions—namely, constant or intermittent or transient high glucose. Intermittent high glucose maximally induced endothelial inflammation by upregulating transcript and/or protein-level expression of ICAM1 and P-selectin and downregulating eNOS, KLF2, and KLF4 protein levels. We next investigated the underlining epigenetic mechanisms responsible for intermittent hyperglycemia-dependent endothelial inflammation. Compared with other high glucose treatment groups, intermittent high glucose-exposed EC exhibited an increased level of H3K27me3 caused by reduction in EZH2 threonine 367 phosphorylation and nuclear retention of EZH2. Intermittent high glucose also promoted polycomb repressive complex-2 (PRC2) assembly and EZH2′s recruitment to histone H3. Abrupt enrichment of H3K27me3 on KLF2 and KLF4 gene promoters caused repression of these genes, further supporting endothelial inflammation. In contrast, reducing H3K27me3 through small molecule and/or siRNA-mediated inhibition of EZH2 rescued KLF2 level and inhibited endothelial inflammation in intermittent high glucose-challenged cultured EC and isolated rat aorta. These findings indicate that abrupt chromatin modifications cause high glucose-dependent inflammatory switch of EC.

Gene-Specific DNA Methylation Profiles in Pediatric Medulloblastomas.

Medulloblastoma is the most common pediatric central nervous tumor of high malignancy that has been classified into both histological subtypes and molecular subgroups by the 2016 World Health Organization classification. However, there is a still need to understand the genomic characteristics and predict the clinical course. The aim of the study is to investigate the significance of the methylation profiles in molecular subclassification and precision medicine of the disease. The study enrolled 47 pediatric medulloblastoma patients. DNA methylation levels of KLF4, SPINT2, RASSF1A, EZH2, ZIC2, and PTCH1 genes were analyzed using methylation-specific pyrosequencing. The significance of the statistical relationship between methylation profiles and clinicopathological parameters including molecular subgroups and histological subtypes, the status of metastasis, and event-free survival were analyzed. DNA methylation analysis demonstrated that KLF4, PTCH1, and ZIC2 hypermethylation were associated with the SHH-activated subgroup, whereas both SPINT2 and RASSF1A hypermethylation were associated with metastatic disease. EZH2 gene was not methylated in any of the samples. We think that customized DNA methylation profiling may be a useful tool in the molecular subclassification of pediatric medulloblastoma and a potential technical approach in precision medicine.

Kruppel-like factor 15 induces the development of mature hepatocyte-like cells from hepatoblasts

The liver is an important metabolic organ that controls homeostasis in the body. Moreover, it functions as a hematopoietic organ, while its metabolic function is low during development. Hepatocytes, which are parenchymal cells of the liver, acquire various metabolic functions by the maturation of hepatic progenitor cells during the fetal period; however, this molecular mechanism is still unclear. In this study, Kruppel-like factor 15 (KLF15) was identified as a new regulator of hepatic maturation through a comprehensive analysis of the expression of transcriptional regulators in mouse fetal and adult hepatocytes. KLF15 is a transcription factor whose expression in the liver increases from the embryonic stage throughout the developmental process. KLF15 induced the overexpression of liver function genes in mouse embryonic hepatocytes. Furthermore, we found that the expression of KLF15 could also induce the expression of liver function genes in hepatoblasts derived from human induced pluripotent stem cells (iPSCs). Moreover, KLF15 increased the promoter activity of tyrosine aminotransferase, a liver function gene. KLF15 also suppressed the proliferation of hepatoblasts. These results suggest that KLF15 induces hepatic maturation through the transcriptional activation of target genes and cell cycle control.

Krueppel-Like Factor 4 Expression in Phagocytes Regulates Early Inflammatory Response and Disease Severity in Pneumococcal Pneumonia.

The transcription factor Krueppel-like factor (KLF) 4 fosters the pro-inflammatory immune response in macrophages and polymorphonuclear neutrophils (PMNs) when stimulated with Streptococcus pneumoniae, the main causative pathogen of community-acquired pneumonia (CAP). Here, we investigated the impact of KLF4 expression in myeloid cells such as macrophages and PMNs on inflammatory response and disease severity in a pneumococcal pneumonia mouse model and in patients admitted to hospital with CAP. We found that mice with a myeloid-specific knockout of KLF4 mount an insufficient early immune response with reduced levels of pro-inflammatory cytokines and increased levels of the anti-inflammatory cytokine interleukin (IL) 10 in bronchoalveolar lavage fluid and plasma and an impaired bacterial clearance from the lungs 24 hours after infection with S. pneumoniae. This results in higher rates of bacteremia, increased lung tissue damage, more severe symptoms of infection and reduced survival. Higher KLF4 gene expression levels in the peripheral blood of patients with CAP at hospital admission correlate with a favourable clinical presentation (lower sequential organ failure assessment (SOFA) score), lower serum levels of IL-10 at admission, shorter hospital stay and lower mortality or requirement of intensive care unit treatment within 28 days after admission. Thus, KLF4 in myeloid cells such as macrophages and PMNs is an important regulator of the early pro-inflammatory immune response and, therefore, a potentially interesting target for therapeutic interventions in pneumococcal pneumonia.

Krüppel-like factor gene function in the ctenophore Mnemiopsis leidyi assessed by CRISPR/Cas9-mediated genome editing.

The Krüppel-like factor (Klf) gene family encodes transcription factors that play an important role in the regulation of stem cell proliferation, cell differentiation and development in bilaterians. Although Klf genes have been shown to specify functionally various cell types in non-bilaterian animals, their role in early-diverging animal lineages has not been assessed. Thus, the ancestral activity of these transcription factors in animal development is not well understood. The ctenophore Mnemiopsis leidyi has emerged as an important non-bilaterian model system for understanding early animal evolution. Here, we characterize the expression and functional role of Klf genes during M. leidyi embryogenesis. Zygotic Klf gene function was assessed with both CRISPR/Cas9-mediated genome editing and splice-blocking morpholino oligonucleotide knockdown approaches. Abrogation of zygotic Klf expression during M. leidyi embryogenesis resulted in abnormal development of several organs, including the pharynx, tentacle bulbs and apical organ. Our data suggest an ancient role for Klf genes in regulating endodermal patterning, possibly through regulation of cell proliferation.

Inhibition of the MAP2K7-JNK pathway with 5Z-7-oxozeaenol induces apoptosis in T-cell acute lymphoblastic leukemia.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive pediatric leukemia with a worse prognosis than most frequent B-cell ALL due to a high incidence of treatment failures and relapse. Our previous work showed that loss of the pioneer factor KLF4 in a NOTCH1-induced T-ALL mouse model accelerated the development of leukemia through expansion of leukemia-initiating cells and activation of the MAP2K7 pathway. Similarly, epigenetic silencing of the KLF4 gene in children with T-ALL was associated with MAP2K7 activation. Here, we showed the small molecule 5Z-7-oxozeaenol (5Z7O) induces dose-dependent cytotoxicity in a panel of T-ALL cell lines mainly through inhibition of the MAP2K7-JNK pathway, which further validates MAP2K7 as a therapeutic target. Mechanistically, 5Z7O-mediated apoptosis was caused by the downregulation of regulators of the G2/M checkpoint and the inhibition of survival pathways. The anti-leukemic capacity of 5Z7O was evaluated using leukemic cells from two mouse models of T-ALL and patient-derived xenograft cells generated using lymphoblasts from pediatric T-ALL patients. Finally, a combination of 5Z7O with dexamethasone, a drug used in frontline therapy, showed synergistic induction of cytotoxicity. In sum, we report here that MAP2K7 inhibition thwarts survival mechanisms in T-ALL cells and warrants future pre-clinical studies for high-risk and relapsed patients.

KLF4 Inhibits the Differentiation of Goat Intramuscular Preadipocytes Through Targeting C/EBPβ Directly.

Intramuscular fat (IMF) deposition is a complicated process, and most of the underlying regulators of this biological process are unknown. Here, we cloned the intact CDS of KLF4 gene, investigated the role of KLF4 by gaining or losing function in vitro and further explored the pathways of KLF4 regulating differentiation of intramuscular preadipocytes in goat. Our results show that goat KLF4 gene consists of 1,536 bp encoding a protein of 486 amino acids. The expression of KLF4 is higher in the lung while lower in the heart and muscle in goat. Knockdown of KLF4 mediated by siRNA technique significantly promotes intramuscular preadipocyte lipid accumulation and upregulates mRNA expression of adipogenic related genes including C/EBPα, C/EBPβ, and PPARγ in vivo cultured cells. Consistently, overexpression of KLF4 inhibits intramuscular adipocyte lipid accumulation and significantly downregulation gene expression of C/EBPβ, PPARγ, aP2, and Pref-1. Further, we found that other members of KLFs were upregulated or downregulated after interference or overexpression of KLF4, including KLF2 and KLF5–7. We also found that C/EBPβ was a potential target of KLF4, because it had an opposite expression pattern with KLF4 during the differentiation of intramuscular preadipocytes and had putative binding sites of KLF4. The dual-luciferase reporter assay indicated that overexpression of KLF4 inhibited the transcriptional activity of C/EBPβ. These results demonstrate that KLF4 inhibits the differentiation of intramuscular preadipocytes in goat by targeting C/EBPβ.

Evaluation of the Effect of Orlistatorlistat on Expression of OCT4, Nanog, SOX2, and KLF4 Genes in Colorectal Cancer SW40 Cell Line

Background and Objective:Orlistat drug is one of the most criticalanti-obesity drugs that widely used around the world. The aim of this study was evaluation the effect of orlistat on the expression of OCT4, Nanog, SOX2, and KLF4 genes in the colorectal cancer SW40 cell line. Materials and Methods:SW40 cell line was cultured in DMEM medium contained orlistat for 24h, and cell viability was assessed by MTT assay. The fold changes of expression of OCT4, NANOG, KLF4, and SOX2 at mRNA level against β-actin were determined by real-timePCR. Two-sample t-test and one-way ANOVA were used to compare the mean of expression of different genes in different groups and different concentrations; a significant level of 0.05 was considered in all tests. Results:Our results showed a significant difference in cell viability, when different doses of Orlistat were used for 24 hour. concentrations of 25 and 100 μM reduce significantly the expression of OCT4 (P<0.05) and SOX2 (P<0.05) in the treated group in comparison to control (P<0.05). Also, the mRNA expression of KLF4 and Nanog was reduced significantly after treatment of SW40 cell lines was performed with 100 μM doses of Orlistat (P<0.05). Conclusion:It appears that after further studies in animal and human phases, orlistat can be used for the treatment of Colorectal Cancer.

Dietary amino acid supplementation affects temporal expression of amino acid transporters and metabolic genes in selected and commercial strains of rainbow trout (Oncorhynchus mykiss).

Replacement of fishmeal as the major protein source in feeds is critical for continued growth and sustainability of the aquaculture industry. However, numerous studies have shown suboptimal fish growth performance and reduced protein retention efficiency when carnivorous fish species are fed low fishmeal-high plant protein feeds. A study was conducted using a commercial strain and a genetically improved strain of rainbow trout selected for improved performance when fed an all plant protein diet to identify physiological differences associated with growth performance in the selected trout strain. Fifty individuals per strain (average weight~580g) were force-fed a plant-protein blend with and without amino acid supplementation (lysine, methionine and threonine) at 0.5% body weight and sampled at intervals over 24h. Samples from intestine and liver were analyzed for specific gene expression analysis related to amino acid transporters, digestive process control, protein degradation and amino acid metabolism. The results showed that expression levels of various intestinal amino acid transporters (SLC1A1, SLC7A9, SLC15A, SLC1A5 SLC6A19 and SLC36A1) were affected by strain, diet and time. Moreover, significant interactions were found regarding the temporal expression levels of cholecystokinin (CCK-L), Krüppel-like factor 15 (KLF15) and aspartate aminotransferase (GOT) transcripts in the examined tissues. The results provide evidence that improved growth and protein retention of the selected strain fed an all-plant protein diet is a result of nutritional adaptation and an overall change in physiological homeostatic control.

MiR-7 regulates the apoptosis of chicken primary myoblasts through the KLF4 gene

ABSTRACT 1. MicroRNAs (miRNAs) play a vital role in the proliferation, differentiation, and apoptosis of myoblasts. However, the effect of miR-7 on the apoptosis of chicken primary myoblasts (CPMs) and its mechanism is still unclear. 2. In this study, the expression of apoptosis marker genes (RAF1, Caspase3, Caspase9, Cytc, Fas) in CPMs was significantly increased after transfection of miR-7 mimic. The expression of the apoptosis marker genes in CPMs was significantly reduced after transfection with miR-7 inhibitor. Flow cytometry showed that the late apoptosis rate of the mimic group was significantly higher than the negative control (NC). The viable cells of the mimic group were significantly lower than the NC. In contrast, inhibition of miR-7 had the opposite effect. 3. The dual-luciferase assay showed that the KLF4 was a target gene of miR-7. The rescue experiment showed that the KLF4 gene could attenuate the effect of miR-7 on the expression of apoptosis marker genes in CPMs. 4. Determination of the function the KLF4 gene showed that the expression of the apoptosis marker genes in CPMs decreased significantly compared with the NC after its overexpression. Inhibition of KLF4 gene had the opposite effect. Flow cytometry showed that overexpression of the KLF4 gene inhibited early apoptosis of myoblasts (P ≤ 0.01), while interference with the KLF4 gene could promote early apoptosis of myoblasts (P ≤ 0.001). 5. The results demonstrated, for the first time, that miR-7 promotes apoptosis in chicken primary myoblasts by regulating the expression of the KLF4 gene.

Severe neonatal haemolytic anaemia caused by compound heterozygous KLF1 mutations: report of four families and literature review

Mutations in the KLF1 gene, which encodes a transcription factor playing a role in erythropoiesis, have recently been demonstrated to be a rare cause of hereditary haemolytic anaemia. We described the genotypic and phenotypic spectra of four unrelated families with compound heterozygous class 2/class 3 KLF1 mutations. All patients had p.G176RfsX179 on one allele and either p.A298P, p.R301H or p.G335R on the other allele. All presented on the first day of life with severe haemolytic anaemia with abnormal red blood cell morphology, markedly increased nucleated red blood cells and hyperbilirubinaemia. Three patients later became transfusion-dependent. All parents with heterozygous KLF1 mutation without co-inherited thalassaemia had normal to borderline mean corpuscular volume (MCV) and normal to slightly elevated Hb F. Fifteen previously reported cases of biallelic KLF1 mutations were identified from a literature review. All except one presented with severe haemolytic anaemia in the neonatal period. Our finding substantiates that compound heterozygous KLF1 mutations are associated with severe neonatal haemolytic anaemia and expands the haematologic phenotypic spectrum. In carriers, the previously suggested findings of low MCV, high Hb A2 and high Hb F are inconsistent; thus this necessitates molecular studies for the identification of carriers.

MiR-933 inhibits proliferation, migration and invasion of lung cancer cell lines by regulation of KLF6 gene

miR-933 inhibits proliferation, migration and invasion of lung cancer cell lines by regulation of KLF6 gene