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Krüppel‐like Factor 7 (KLF7) is a zinc finger transcription factor that has a critical role in cellular differentiation, tumorigenesis, and regeneration. Mutations in Klf7 are associated with autism spectrum disorder, which is characterized by neurodevelopmental delay and intellectual disability. Here we show that KLF7 regulates neurogenesis and neuronal migration during mouse cortical development. Conditional depletion of KLF7 in neural progenitor cells resulted in agenesis of the corpus callosum, defects in neurogenesis, and impaired neuronal migration in the neocortex. Transcriptomic profiling analysis indicated that KLF7 regulates a cohort of genes involved in neuronal differentiation and migration, including p21 and Rac3. These findings provide insights into our understanding of the potential mechanisms underlying neurological defects associated with Klf7 mutations. [ABSTRACT FROM AUTHOR]

Abstract Background Ribosomal biogenesis and ribosomal proteins have attracted attention in the context of tumor biology in recent years. Instead of being mere translational machineries, ribosomes might play an active role in tumor initiation and progression. Despite its importance, regulation of ribosomal biogenesis is still not completely understood. Methods Using Gene Set Enrichment Analysis of RNA sequencing and proteomical mass spectrometry data in breast cancer cells expressing Krüppel-like factor 7 (KLF7), we identified processes altered by this transcription factor. In silico analyses of a cohort of breast cancer patients in The Cancer Genome Atlas confirmed our finding. We further verified the role of KLF7 the identified ribosomal processes in in vitro assays of mammary carcinoma cell lines and analyses of breast cancer patients’ tissue slices. Results We identified the transcription factor Krüppel-like factor 7 (KLF7) as a regulator of ribosomal biogenesis and translation in breast cancer cells and tissue. Highly significant overlapping processes related to ribosomal biogenesis were identified in proteomics and transcriptomics data and confirmed in patients’ breast cancer RNA Seq data. Further, nucleoli, the sites of ribosomal biogenesis, were morphologically altered and quantitatively increased in KLF7-expressing cells. Pre-rRNA processing was identified as one potential process affected by KLF7. In addition, an increase in global translation independent from proliferation and transcription was observed upon exogenous KLF7 expression in vitro. Importantly, in a cohort of breast cancer patients, KLF7-expression levels correlated with aggressiveness of the intrinsic breast cancer subtype and tumor grading. Moreover, KLF7 correlated with nucleolar characteristics in human breast tumor tissue, indicating a role for KLF7 in ribosomal biogenesis. Conclusion In mammary carcinoma, KLF7 is involved in ribosomal biogenesis. Alterations of ribosomal biogenesis has far reaching quantitative and qualitative implications for the proteome of the cancer cells. This might influence the aggressiveness of cancer cells.

Objectives: Atherosclerosis (AS) remains a major contributor to death worldwide. This study sought to explore the role of Krüppel-like factor 7 (KLF7) in AS lesions via regulating glucose metabolic reprogramming (GMR) in macrophages. Methods: AS mouse and cell models were established via high-fat-diet feeding and oxidized low-density lipoprotein (ox-LDL) induction. KLF7, histone deacetylase 4 (HDAC4), miR-148b-3p, and nuclear receptor corepressor 1 (NCOR1) expressions in aortic tissue and cells were detected via reverse transcription quantitative polymerase chain reaction or Western blotting. Parameters of AS lesions and mouse metabolism were detected via hematoxylin-eosin, oil red O, and Masson staining, assay kits, glucose tolerance test, and enzymatic analysis. Peritoneal macrophages of mice were isolated and cellular metabolism was detected via Seahorse metabolic flux analysis, assay kits, ELISA, and Western blotting. Bindings among KLF7, HDAC4, microRNA (miR)-148b-3p, and NCOR1 were testified via the dual-luciferase assay and chromatin immunoprecipitation assay. Results: KLF7 was poorly expressed in AS mice and ox-LDL-induced RAW264.7 cells. KLF7 overexpression attenuated AS lesions and rescued metabolic abnormities in AS mice, and reduced glucose intake and GMR in ox-LDL-induced RAW264.7 cells. Mechanically, KLF7 bound to the HDAC4 promoter to activate HDAC4. HDAC4 reduced H3 and H4 acetylation levels in the miR-148b promoter to inhibit miR-148b-3p and promote NCOR1 transcription. HDAC4 downregulation abolished the protective role of KLF7 overexpression in AS mice and ox-LDL-induced RAW264.7 cells via the miR-148b-3p/NCOR1 axis. Conclusion: KLF7 bound to the HDAC4 promoter to activate HDAC4, inhibit miR-148b-3p via reducing acetylation level, and promote NCOR1 transcription, thereby limiting GMR in macrophages and alleviating AS lesions. [ABSTRACT FROM AUTHOR]

Background: Ribosomal biogenesis and ribosomal proteins have attracted attention in the context of tumor biology in recent years. Instead of being mere translational machineries, ribosomes might play an active role in tumor initiation and progression. Despite its importance, regulation of ribosomal biogenesis is still not completely understood.Methods: Using Gene Set Enrichment Analysis of RNA sequencing and proteomical mass spectrometry data in breast cancer cells expressing Krüppel-like factor 7 (KLF7), we identified processes altered by this transcription factor. In silico analyses of a cohort of breast cancer patients in The Cancer Genome Atlas confirmed our finding. We further verified the role of KLF7 the identified ribosomal processes in in vitro assays of mammary carcinoma cell lines and analyses of breast cancer patients' tissue slices.Results: We identified the transcription factor Krüppel-like factor 7 (KLF7) as a regulator of ribosomal biogenesis and translation in breast cancer cells and tissue. Highly significant overlapping processes related to ribosomal biogenesis were identified in proteomics and transcriptomics data and confirmed in patients' breast cancer RNA Seq data. Further, nucleoli, the sites of ribosomal biogenesis, were morphologically altered and quantitatively increased in KLF7-expressing cells. Pre-rRNA processing was identified as one potential process affected by KLF7. In addition, an increase in global translation independent from proliferation and transcription was observed upon exogenous KLF7 expression in vitro. Importantly, in a cohort of breast cancer patients, KLF7-expression levels correlated with aggressiveness of the intrinsic breast cancer subtype and tumor grading. Moreover, KLF7 correlated with nucleolar characteristics in human breast tumor tissue, indicating a role for KLF7 in ribosomal biogenesis.Conclusion: In mammary carcinoma, KLF7 is involved in ribosomal biogenesis. Alterations of ribosomal biogenesis has far reaching quantitative and qualitative implications for the proteome of the cancer cells. This might influence the aggressiveness of cancer cells. [ABSTRACT FROM AUTHOR]

Our previous study has shown that the transcription factor Krüppel-like factor 7 (KLF7) promotes peripheral nerve regeneration and motor function recovery after spinal cord injury. KLF7 also participates in traumatic brain injury, but its regulatory mechanisms remain poorly understood. In the present study, an HT22 cell model of traumatic brain injury was established by stretch injury and oxygen-glucose deprivation. These cells were then transfected with an adeno-associated virus carrying KLF7 (AAV-KLF7). The results revealed that, after stretch injury and oxygen-glucose deprivation, KLF7 greatly reduced apoptosis, activated caspase-3 and lactate dehydrogenase, downregulated the expression of the apoptotic markers B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax) and cleaved caspase-3, and increased the expression of βIII-tubulin and the antiapoptotic marker Bcl-2. Furthermore, KLF7 overexpression upregulated Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) phosphorylation in HT22 cells treated by stretch injury and oxygen-glucose deprivation. Immunoprecipitation assays revealed that KLF7 directly participated in the phosphorylation of STAT3. In addition, treatment with AG490, a selective inhibitor of JAK2/STAT3, weakened the protective effects of KLF7. A mouse controlled cortical impact model of traumatic brain injury was then established. At 30 minutes before modeling, AAV-KLF7 was injected into the ipsilateral lateral ventricle. The protein and mRNA levels of KLF7 in the hippocampus were increased at 1 day after injury and recovered to normal levels at 3 days after injury. KLF7 reduced ipsilateral hippocampal atrophy, decreased the injured cortex volume, downregulated Bax and cleaved caspase-3 expression, and increased the number of 5-bromo-2′-deoxyuridine-positive neurons and Bcl-2 protein expression. Moreover, KLF7 transfection greatly enhanced the phosphorylation of JAK2 and STAT3 in the ipsilateral hippocampus. These results suggest that KLF7 may protect hippocampal neurons after traumatic brain injury through activation of the JAK2/STAT3 signaling pathway. The study was approved by the Institutional Review Board of Mudanjiang Medical University, China (approval No. mdjyxy-2018-0012) on March 6, 2018.

Objective: The present study was performed to determine the clinical relevance of KLF7 in tongue squamous cell carcinoma (TSCC) and to characterize its potential function and mechanism of action. Materials and methods: KLF7 expression was measured by RT‐qPCR in 21 tongue cancer samples. The clinical relevance of KLF7 was analyzed in another cohort of 127 TSCC samples from a public database. Then, we performed RNA sequencing analysis in KLF7‐overexpressing TSCC (SCC9 and CAL27) cells to define significantly altered pathways. The possible changes in migration and adhesion were then analyzed in KLF7‐overexpressing and knockdown TSCC cells. Results: Our results showed that KLF7 mRNA expression was upregulated in TSCC and was significantly associated with the T and N stages. Patients with high‐KLF7 expression had worse overall survival. RNA sequencing and KEGG enriched pathway analysis showed that altered genes were enriched in extracellular matrix‐receptor interactions and focal adhesions in both cell lines. KLF7‐overexpressing TSCC cell lines showed enhanced migration capacity and cell adhesion ability, and knockdown of KLF7 expression decreased TSCC migration and adhesion ability. Conclusions: We concluded that KLF7 was overexpressed in TSCC and has prognostic value. KLF7 promoted TSCC migration and increased cell adhesion. [ABSTRACT FROM AUTHOR]

Abstract Background Our previous study found that chicken KLF7 was an important regulator in formation of adipose tissue. In the present study, we analyzed the association for DNA methylation in chicken KLF7 with its transcripts of abdominal adipose tissue and blood metabolic indicators. Results The KLF7 transcripts of the adipose tissue of Chinese yellow broilers were associated with age (F = 6.67, P = 0.0035). In addition, the KLF7 transcripts were negatively correlated with blood glucose levels (r = − 0.61841, P = 0.0140). The DNA methylation levels of 26 CpG loci in the chicken KLF7 promoter and Exon 2 were studied by Sequenom MassArray. A total of 22 valid datasets were obtained. None of them was significantly different in relation to age (P > 0.05). However, the DNA methylation levels in the promoter were lower than those in Exon 2 (T = 40.74, P

Non-small-cell lung cancer (NSCLC) remains the leading cause of cancer-related death worldwide. Accumulating researches have highlighted the ability of exosome-encapsulated microRNAs (miRNAs or miRs) as potential circulating biomarkers for lung cancer. The current study aimed to evaluate the significance of mesenchymal stem cells (MSCs)-derived exosomal miR-204 in the invasion, migration, and epithelial-mesenchymal transition (EMT) of NSCLC cells. Initially, the expression of miR-204 in human NSCLC tissues and cells was determined by RT-qPCR, which demonstrated that miR-204 was downregulated in NSCLC tissues and cells. Next, Krüppel-like factor 7 (KLF7) was predicted and validated to be a target of miR-204 using dual-luciferase reporter gene assay. NSCLC A549 cells were treated with MSCs-derived exosomes, after which the migration and invasion of A549 cells were detected and expression of EMT-related proteins (E-cadherin, N-cadherin, and Vimentin), KLF7, p-AKT/AKT, and HIF-1α were measured. The results of gain- and loss-of-function assays revealed that miR-204 overexpression in MSCs-derived exosomes inhibited KLF7 expression and the AKT/HIF-1α pathway activity, resulting in impaired cell migration, invasion, as well as EMT. In conclusion, the key findings of the current study demonstrate that exosomal miR-204 from MSCs possesses anticarcinogenic properties against NSCLC via the KLF7/AKT/HIF-1α axis. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of miR-132 on the proliferation and migration of nasopharyngeal carcinoma (NPC) cells by negatively targeting the expression of Kruppel-like factor 7 (KLF7). Methods NPC CNE-2Z cells were divided into blank group, NC group, miR-132 mimics group, KLF7-OE group and miR-132 mimics + KLF7-OE group. The expressions of miR-132 in NPC tissues and cells and normal nasopharyngeal tissues and cells were detected by fluorescence quantitative PCR (qPCR). The expression level of KLF7 protein was detected by Western blot assay. The targeting relationship between KLF7 and miR-132 was predicted and verified by TargetScan website and dual luciferase detection. CCK-8 test and Transwell test were used to detect the proliferation, migration and invasion of CNE-2Z cells. The tumor formation test in nude mice and immunohistochemistry were used to detect the growth of CNE-2Z cells in vivo and the blood vessels in tumor. Results The expression level of miR-132 was decreased in nasopharyngeal carcinoma tissue than that in normal nasopharyngeal tissue, and the expression of KLF7 protein was increased. The expression level of miR-132 was decreased in nasopharyngeal carcinoma CNE-2Z cells than that in normal nasopharyngeal epithelial cells NP69, and the expression of KLF7 protein was increased (P＜0.05). TargetScan prediction and dual luciferase detection showed that KLF7 was the target gene of miR-132. Compared with those in the blank group and NC group, the expression of KLF7 protein, cell proliferation activity at 36, 48 and 60 h, numbers of migration and invasion, tumor volume and microvessel density (MVD) in CNE-2Z cells were significantly decreased in miR-132 mimics group (P＜0.05), while the results were oppositive in KLF7- OE group (P＜0.05). Compared with those in miR-132 mimics group, the expression level of KLF7 protein, cell proliferation activity at 36, 48 and 60 h, numbers of migration and invasion, tumor volume and MVD were significantly increased in miR132 mimics + KLF7-OE group (P＜0.05). Conclusion miR-132 can affect the proliferation, migration and invasion of NPC cells by negatively regulating the expression of KLF7. [ABSTRACT FROM AUTHOR]

Background: Our previous study found that chicken KLF7 was an important regulator in formation of adipose tissue. In the present study, we analyzed the association for DNA methylation in chicken KLF7 with its transcripts of abdominal adipose tissue and blood metabolic indicators. Results: The KLF7 transcripts of the adipose tissue of Chinese yellow broilers were associated with age (F = 6.67, P = 0.0035). In addition, the KLF7 transcripts were negatively correlated with blood glucose levels (r = − 0.61841, P = 0.0140). The DNA methylation levels of 26 CpG loci in the chicken KLF7 promoter and Exon 2 were studied by Sequenom MassArray. A total of 22 valid datasets were obtained. None of them was significantly different in relation to age (P > 0.05). However, the DNA methylation levels in the promoter were lower than those in Exon 2 (T = 40.74, P < 0.01). Correlation analysis showed that the DNA methylation levels of PCpG6 and E2CpG9 were significantly correlated with KLF7 transcripts and blood high-density lipoprotein levels, respectively, and many CpG loci were correlated with each other (P < 0.05). The methylation data were subjected to principal component analysis and factor analysis. The six principal components (z1–z6) were extracted and named Factors 1–6, respectively. Factor analysis showed that Factor 1 had a higher load on the loci in the promoter, and Factors 2–6 loaded highly on quite different loci in Exon 2. Correlation analysis showed that only z1 was significantly correlated to KLF7 transcripts (P < 0.05). In addition, an established regression equation between z1 and KLF7 transcripts was built, and the contribution of z1 to the variation on KLF7 transcripts was 34.29%. Conclusions: In conclusion, the KLF7 transcripts of chicken abdominal adipose tissue might be inhibited by DNA methylation in the promoter, and it might be related to the DNA methylation level of PCpG6. [ABSTRACT FROM AUTHOR]

Background/Aims: Krüppel-like factor (KLF) 7 protein is a member of the KLF transcription factor family, which plays important roles in regulating the expression of genes involved in cell growth, proliferation, differentiation and metabolism. However, the role of KLF7 in gastric cancer (GC) is unknown. The aim of this study is to explore the role of KLF7 in GC and its correlation with clinicopathological characteristics and prognosis of GC patients. Methods: We first systematically evaluated dysregulation of the KLF family in The Cancer Genome Atlas (TCGA) GC database. Then, 252 patients who underwent surgery for GC were enrolled to validate the results from the TCGA. Functional studies were also used to explore the role of KLF7 in GC. Results: In the TCGA database, we found that KLF7 was an independent predictor for survival by both univariate and multivariate analysis (P

Congenital deafness is prevalent among modern dog breeds, including Australian Stumpy Tail Cattle Dogs (ASCD). However, in ASCD, no causative gene has been identified so far. Therefore, we performed a genome-wide association study (GWAS) and whole genome sequencing (WGS) of affected and normal individuals. For GWAS, 3 bilateral deaf ASCDs, 43 herding dogs, and one unaffected ASCD were used, resulting in 13 significantly associated loci on 6 chromosomes, i.e., CFA3, 8, 17, 23, 28, and 37. CFA37 harbored a region with the most significant association (−log10(9.54 × 10−21) = 20.02) as well as 7 of the 13 associated loci. For whole genome sequencing, the same three affected ASCDs and one unaffected ASCD were used. The WGS data were compared with 722 canine controls and filtered for protein coding and non-synonymous variants, resulting in four missense variants present only in the affected dogs. Using effect prediction tools, two variants remained with predicted deleterious effects within the Heart development protein with EGF like domains 1 (HEG1) gene (NC_006615.3: g.28028412G>C; XP_022269716.1: p.His531Asp) and Kruppel-like factor 7 (KLF7) gene (NC_006619.3: g.15562684G>A; XP_022270984.1: p.Leu173Phe). Due to its function as a regulator in heart and vessel formation and cardiovascular development, HEG1 was excluded as a candidate gene. On the other hand, KLF7 plays a crucial role in the nervous system, is expressed in the otic placode, and is reported to be involved in inner ear development. 55 additional ASCD samples (28 deaf and 27 normal hearing dogs) were genotyped for the KLF7 variant, and the variant remained significantly associated with deafness in ASCD (p = 0.014). Furthermore, 24 dogs with heterozygous or homozygous mutations were detected, including 18 deaf dogs. The penetrance was calculated to be 0.75, which is in agreement with previous reports. In conclusion, KLF7 is a promising candidate gene causative for ASCD deafness.

The biological relation between ubiquitin ligases and their substrates has been largely unclear. We previously developed a method—differential proteomics‐based identification of ubiquitylation substrates (DiPIUS)—for the comprehensive identification of substrates for a given ubiquitin ligase. We have now applied DiPIUS to the F‐box protein Fbxw7 in three cell lines (mHepa, Neuro2A and C2C12) and thereby identified Krüppel‐like factor 7 (KLF7) as a candidate substrate of the SCFFbxw7 ubiquitin ligase complex. KLF7 was shown to interact with Fbxw7 and to undergo Fbxw7‐mediated polyubiquitylation. The stability of KLF7 was increased by depletion of Fbxw7, mutation of a putative Cdc4 phosphodegron (CPD) of KLF7 or exposure to inhibitors of glycogen synthase kinase‐3 (GSK‐3). Over‐expression of Fbxw7 in Neuro2A cells down‐regulated expression of the p21Cip1 gene, which is a transcriptional target of KLF7 in neuronal differentiation and maintenance. Despite the presence of an almost identical CPD sequence in KLF6, the closest paralog of KLF7, mutation of this sequence affected neither the interaction of KLF6 with Fbxw7 nor its half‐life. Our results suggest that KLF7, but not KLF6, is a bona fide substrate of SCFFbxw7, and that control of KLF7 abundance by SCFFbxw7 might contribute to the regulation of neuronal differentiation and maintenance. [ABSTRACT FROM AUTHOR]

Background: Kruppel-like factor 7 (KLF7) is associated with type 2 diabetes and obesity; however, at present the KLF7 level in polycystic ovary syndrome (PCOS) remains unreported. Methods: In this study, serum KLF7, glucose, insulin, C-reactive protein (CRP), follicle-stimulating hormone (FSH), luteinizing hormone (LH), and total testosterone concentrations were measured in 65 women with PCOS and 61 healthy controls. Results: Body mass index (BMI) in PCOS and the control group were all < 25 kg/m2. The median concentration of KLF7 was 3.630 ng/mL [interquartile range (IQR): 1.547 - 7.172] in women with PCOS, which was significantly lower than that of controls (5.282 ng/mL, IQR: 3.128 - 11.263, p = 0.003). The KLF7 level was positively correlated with low-density lipoprotein cholesterol (LDL-C) (r = 0.261, p = 0.018). No significant association was observed between KLF7 and the BMI, total testosterone, and insulin resistance (IR). Conclusions: The KLF7 level decreased in women with PCOS. KLF7 may represent a new therapeutic target in the treatment of PCOS. [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the underlying mechanisms of hypoxia-induced microRNA (miR)-210 effects on mouse GC-2spd (GC-2) cells. GC-2 cells were subjected to hypoxia or normoxia for 12, 24, 48 and 72 h. Apoptosis of GC-2 cells was detected using terminal deoxynucleotidyl-transferase-meditated dUTP nick end labeling and flow cytometry. Reverse transcription-quantitative polymerase chain reaction was performed to analyze the expression of miR-210. Hypoxia-inducible factor-1α (HIF-1α), caspase-3, B-cell lymphoma 2, apoptosis regulator BAX and Kruppel-like factor 7 (KLF7) protein expression levels were detected by western blotting. Luciferase reporter gene assays were used to assess the targeting effects of miR-210 on KLF7. Hypoxia induced GC-2 cell apoptosis and increased the expression of HIF-1α and pro-apoptotic proteins; however, decreased anti-apoptotic protein expression levels. Furthermore, hypoxia resulted in the upregulation of miR-210 in GC-2 cells. HIF-1α and miR-210 were involved in the apoptosis of GC-2 cells by mediating the expression of apoptosis-associated proteins. Furthermore, KLF7 was directly targeted by miR-210 to influence the apoptosis of GC-2 cells subjected to hypoxia. The results suggested that hypoxia-induced miR-210 stimulated the activation of the apoptosis signaling pathway and contributed to the apoptosis of GC-2 cells by targeting KLF7. [ABSTRACT FROM AUTHOR]

Background: Metastasis is a major cause of HCC-related deaths with no effective pharmacotherapies. Chronic inflammation promotes HCC dissemination, however, its underlying mechanisms are not fully understood. Here, we investigated the role of Krüppel-like factor 7 (KLF7) in inflammation-provoked HCC metastasis and proposed therapeutic strategies for KLF7-positive patients. Methods: The expression of KLF7 in human HCC specimens were examined by immunohistochemistry and quantitative real-time PCR. The luciferase reporter assays and chromatin immunoprecipitation assays were conducted to explore the transcriptional regulation related to KLF7. Orthotopic xenograft models and DEN/CCl 4 -induced HCC models were established to evaluate HCC progression and metastasis. Results: KLF7 overexpression promotes HCC metastasis through transactivating toll-like receptor 4 (TLR4) and protein tyrosine kinase 2 (PTK2) expression. High mobility group box 1 (HMGB1) upregulates KLF7 expression through the TLR4/advanced glycosylation end-product specific receptor (RAGE)-PI3K-AKT-NF-κB pathway, forming an HMGB1-KLF7-TLR4 positive feedback loop. The HMGB1-KLF7-TLR4/PTK2 axis is gradually activated during the progression of inflammation-HCC transition. Genetic depletion of KLF7 impedes HMGB1-mediated HCC progression and metastasis. The combined application of TLR4 inhibitor TAK-242 and PTK2 inhibitor defactinib alleviates HCC progression and metastasis induced by the HMGB1-KLF7 axis. In human HCCs, KLF7 expression is positively correlated with cytoplasmic HMGB1, p-p65, TLR4, and PTK2 levels, and patients positively co-expressing HMGB1/KLF7, p-p65/KLF7, KLF7/TLR4 or KLF7/PTK2 exhibit the worst prognosis. Conclusions: HMGB1-induced KLF7 overexpression facilitates HCC progression and metastasis by upregulating TLR4 and PTK2. Genetic ablation of KLF7 via AAV gene therapy and combined blockade of TLR4 and PTK2 represents promising therapy strategies for KLF7-positive HCC patients., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background/Aims: Krüppel-like factor (KLF) 7 protein is a member of the KLF transcription factor family, which plays important roles in regulating the expression of genes involved in cell growth, proliferation, differentiation and metabolism. However, the role of KLF7 in gastric cancer (GC) is unknown. The aim of this study is to explore the role of KLF7 in GC and its correlation with clinicopathological characteristics and prognosis of GC patients. Methods: We first systematically evaluated dysregulation of the KLF family in The Cancer Genome Atlas (TCGA) GC database. Then, 252 patients who underwent surgery for GC were enrolled to validate the results from the TCGA. Functional studies were also used to explore the role of KLF7 in GC. Results: In the TCGA database, we found that KLF7 was an independent predictor for survival by both univariate and multivariate analysis (P<0.05). In a validation cohort, KLF7 expression was significantly increased in GC tissues compared with adjacent normal controls (P=0.013). High KLF7 expression correlated with inferior prognostic factors, such as T stage (P=0.022), N stage (P =0.005) and lymphovascular invasion (P=0.009). Furthermore, we observed a strong negative correlation between KLF7 expression and 5-year overall survival and disease-free survival in GC patients (P<0.05). Moreover, our in vitro studies showed a notable decrease in migration in KLF7 knockdown cells. Conclusion: KLF7 has an important role in GC progression, as it inhibits GC cell migration and may serve as a prognostic marker. [ABSTRACT FROM AUTHOR]

Krüppel-like factor 7 (KLF7) is a member of the KLF family of zinc finger transcription factors, and was the first KLF cloned using complementary DNA and polymerase chain reaction (PCR) techniques with human vascular endothelial cells as a template. In addition, KLF7 is known as the ubiquitous Krüppel-like factor, as it is widely expressed in numerous human tissues at low levels. In the present study, the function of KLF7 in migration and epithelial-mesenchymal transition (EMT), which are associated with tumor progression, was investigated in human oral squamous cell carcinoma (OSCC) cells. Genes that were differentially expressed in normal vs. OSCC tissue were identified in the Gene Expression Omnibus database, which identified upregulation of KLF7 in OSCC. The expression and subcellular location of KLF7 was then analyzed using immunohistochemistry. KLF7 expression was measured in three OSCC cell lines, and the two cell lines with the highest (HN13) and lowest (CAL27) KLF7 expression were selected for further analysis. Subsequently, HN13 cells with reduced KLF7 expression (sh-HN13) and CAL27 cells overexpressing KLF7 (OE-CAL27) were constructed. Transwell migration and wound healing assays were then used to analyze the migration of the cells. In addition, mRNA and protein expression levels of the EMT markers E-cadherin, N-cadherin, vimentin and snail were detected using reverse transcription-quantitative PCR and western blotting. KLF7 overexpression in OSCC was validated using tissue immunohistochemistry, which identified moderate to high cytoplasmic staining of KLF7 in OSCC cells. KLF7 knockdown and overexpression altered the migration ability of sh-HN13 and OE-CAL27 cells, which decreased and increased significantly respectively. Expression of E-cadherin, N-cadherin, vimentin and snail was markedly altered in sh-HN13 and OE-CAL27 cells, indicating changes in EMT status. The results of the present study suggest that KLF7 overexpression changes the migratory behavior of OSCC cells, and induces EMT and lymph node metastasis through the expression of snail. [ABSTRACT FROM AUTHOR]

Abstract: Gene targeting in mice has recently demonstrated that transcription factor KLF7 plays a critical role in neurite outgrowth and neuronal survival. Here we extended this genetic evidence by establishing the transcriptional profile of differentiating olfactory sensory neurons (OSNs) in Klf7 −/− mice, and by identifying relevant genes that are directly regulated by KLF7. Functional clustering of DNA microarray data revealed that loss of KLF7 affects primarily the activity of genes involved in OSN differentiation, axonal growth, cytoskeletal dynamics, cell adhesion and synaptogenesis. Cell transfection experiments, on the other hand, demonstrated that the promoters of the genes encoding the OSN-specific OMP and the adhesion molecule L1 are both activated by KLF7 binding to CACCC motifs. Collectively, these results advance knowledge of transcriptional regulation of olfactory neurogenesis and KLF7 action. [Copyright &y& Elsevier]

Aims/hypothesis: Although genetic susceptibility plays an important role in the pathogenesis of type 2 diabetes, most of the genes that influence susceptibility to type 2 diabetes have yet to be identified. Krüppel-like transcription factors are known to play important roles in development and cell differentiation, and have recently been implicated in the pathogenesis of type 2 diabetes. The present study aimed to examine the associations of single nucleotide polymorphisms (SNPs) in genes encoding members of the Krüppel-like-factor (KLF) family with type 2 diabetes in a large cohort of Japanese subjects. Methods: We genotyped 33 SNP loci found in 12 KLF genes in subjects with type 2 diabetes and in subjects from the general population using the PCR-Invader assay. We also examined the effects of the overexpression of KLF7 on adipogenesis in 3T3-L1 cells. Results: We identified a significant association between an SNP in KLF7 and type 2 diabetes (A vs C: p=0.004 after Bonferroni's correction, odds ratio=1.59, 95% CI 1.27-2.00). The expression of Klf7 decreased in response to the differentiation of 3T3-L1 adipocytes, and the overexpression of KLF7 resulted in significant inhibition of adipogenesis in 3T3-L1 cells. Conclusions/interpretation: These results indicate that the gene encoding KLF7 is a novel candidate for conferring genetic susceptibility to type 2 diabetes. [ABSTRACT FROM AUTHOR]

Morphine was previously found to compromise the chemotactic mobility of leukocytic cells toward inflammatory sites as well as induce the expression of chemokine receptor CCR5, a coreceptor for HIV entry, on lymphocytes. Both effects increase the host's susceptibility for HIV infection. In this report, we used a reverse transcription-polymerase chain reaction-based differential display (RT-PCR DD), an RNA finger printing procedure, coupled with Northern slot blot hybridization, in identifying gene or genes regulated by morphine in human lymphocytic cells. Ten positive gene segments, including two morphine downregulated genes and eight morphine upregulated genes, were identified. Analysis of the DNA sequences of these gene segments showed high homologies to known DNA and protein sequences on databases, of which two apoptosis/cell growth-related genes (PNAS-133 and Kru¨ppel-like factor 7) were revealed. PNAS-133 gene was downregulated by morphine whereas the Kru¨ppel-like factor 7 (KLF7), a zinc finger transcription factor, was upregulated by morphine. The upregulation of KLF7 by morphine was further studied and found to be at both the transcriptional and the translational levels in a naloxone-reversible manner. Cell growth was promoted by morphine up to day 4 (72 h) post treatment. These findings suggest a unique role for morphine in regulating cell proliferation and differentiation of immune cells. [Copyright &y& Elsevier]

Our previous study has shown that the transcription factor Krüppel-like factor 7 (KLF7) promotes peripheral nerve regeneration and motor function recovery after spinal cord injury. KLF7 also participates in traumatic brain injury, but its regulatory mechanisms remain poorly understood. In the present study, an HT22 cell model of traumatic brain injury was established by stretch injury and oxygen-glucose deprivation. These cells were then transfected with an adeno-associated virus carrying KLF7 (AAV-KLF7). The results revealed that, after stretch injury and oxygen-glucose deprivation, KLF7 greatly reduced apoptosis, activated caspase-3 and lactate dehydrogenase, downregulated the expression of the apoptotic markers B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax) and cleaved caspase-3, and increased the expression of βIII-tubulin and the antiapoptotic marker Bcl-2. Furthermore, KLF7 overexpression upregulated Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) phosphorylation in HT22 cells treated by stretch injury and oxygen-glucose deprivation. Immunoprecipitation assays revealed that KLF7 directly participated in the phosphorylation of STAT3. In addition, treatment with AG490, a selective inhibitor of JAK2/STAT3, weakened the protective effects of KLF7. A mouse controlled cortical impact model of traumatic brain injury was then established. At 30 minutes before modeling, AAV-KLF7 was injected into the ipsilateral lateral ventricle. The protein and mRNA levels of KLF7 in the hippocampus were increased at 1 day after injury and recovered to normal levels at 3 days after injury. KLF7 reduced ipsilateral hippocampal atrophy, decreased the injured cortex volume, downregulated Bax and cleaved caspase-3 expression, and increased the number of 5-bromo-2'-deoxyuridine-positive neurons and Bcl-2 protein expression. Moreover, KLF7 transfection greatly enhanced the phosphorylation of JAK2 and STAT3 in the ipsilateral hippocampus. These results suggest that KLF7 may protect hippocampal neurons after traumatic brain injury through activation of the JAK2/STAT3 signaling pathway. The study was approved by the Institutional Review Board of Mudanjiang Medical University, China (approval No. mdjyxy-2018-0012) on March 6, 2018., Competing Interests: None

The purpose of this study is to investigate the effects of Krüppel-like factor 7 (KLF7) on retinal ganglion cells (RGCs) and retinal function after retinal ischemia-reperfusion (RIR) injury in mice. Male C57BL/6J mice were intravitreally injected with recombinant adeno-associated vectors (rAAV-KLF7-EGFP or rAAV-EGFP), and subsequently used to induce RIR injury. Retinal cryosections were used to access the efficacy of virus transfection, 1, 2, 3, and 4 weeks after rAAV-KLF7-EGFP transfer. RGCs survival rate was observed and quantified by immunofluorescent staining, 7 days after RIR injury. Meanwhile, electroretinogram (ERG) and optomotor response were used to evaluate the electrophysiological functions and visual acuity. Apoptosis was evaluated by TUNEL staining 1 day after RIR injury. Expression of KLF7, Akt, phospho-Akt, Bcl-2, and Bax were further detected by western blot to excavate the underlying mechanism. The transfection efficiency of rAAV-KLF7-EGFP was increased in a time-dependent manner, and the number of EGFP-positive cells was increased significantly 3 weeks after rAAV-KLF7-EGFP transfer. RGCs survival rates, amplitudes of ERG a-, b-wave, Ops, PhNR, and visual acuity of mice were decreased after RIR injury. With the increase of light intensity, the amplitudes of scotopic ERG a- and b-wave were gradually increased while the incubation period was gradually shortened. RGCs survival rates, amplitudes of ERG a-, b-wave, Ops, PhNR, and visual acuity of mice were increased after rAAV-KLF7-EGFP transfer. The protein level of KLF7 was up-regulated after rAAV-KLF7-EGFP transfer. Up-regulation of KLF7 significantly inhibited cells apoptosis, increased phospho-Akt and Bcl-2 expression, and decreased Bax expression. There were no significant changes in Akt expression. Overexpression of KLF7 can not only prevent the loss of RGCs, but also preserve the electrophysiological function. In addition, overexpression of KLF7 can ameliorate the retinal dysfunction after RIR injury, and ultimately improve the visual acuity of mice. The activation of Akt pathway and the suppression of the mitochondrial apoptotic pathway contribute to the neuroprotection of KLF7. • KLF7 overexpression prevented the loss of RGCs after retinal ischemia-reperfusion injury. • KLF7 overexpression preserved the electrophysiological function of RGCs after retinal ischemia-reperfusion injury. • KLF7 overexpression ameliorated the retinal dysfunction after retinal ischemia-reperfusion injury. • The neuroprotection of KLF7 achieved by activating Akt pathway and inhibiting the mitochondrial apoptosis pathway. [ABSTRACT FROM AUTHOR]

Congenital deafness is prevalent among modern dog breeds, including Australian Stumpy Tail Cattle Dogs (ASCD). However, in ASCD, no causative gene has been identified so far. Therefore, we performed a genome-wide association study (GWAS) and whole genome sequencing (WGS) of affected and normal individuals. For GWAS, 3 bilateral deaf ASCDs, 43 herding dogs, and one unaffected ASCD were used, resulting in 13 significantly associated loci on 6 chromosomes, i.e., CFA3, 8, 17, 23, 28, and 37. CFA37 harbored a region with the most significant association (−log10(9.54 × 10−21) = 20.02) as well as 7 of the 13 associated loci. For whole genome sequencing, the same three affected ASCDs and one unaffected ASCD were used. The WGS data were compared with 722 canine controls and filtered for protein coding and non-synonymous variants, resulting in four missense variants present only in the affected dogs. Using effect prediction tools, two variants remained with predicted deleterious effects within the Heart development protein with EGF like domains 1 (HEG1) gene (NC_006615.3: g.28028412G>C; XP_022269716.1: p.His531Asp) and Kruppel-like factor 7 (KLF7) gene (NC_006619.3: g.15562684G>A; XP_022270984.1: p.Leu173Phe). Due to its function as a regulator in heart and vessel formation and cardiovascular development, HEG1 was excluded as a candidate gene. On the other hand, KLF7 plays a crucial role in the nervous system, is expressed in the otic placode, and is reported to be involved in inner ear development. 55 additional ASCD samples (28 deaf and 27 normal hearing dogs) were genotyped for the KLF7 variant, and the variant remained significantly associated with deafness in ASCD (p = 0.014). Furthermore, 24 dogs with heterozygous or homozygous mutations were detected, including 18 deaf dogs. The penetrance was calculated to be 0.75, which is in agreement with previous reports. In conclusion, KLF7 is a promising candidate gene causative for ASCD deafness. [ABSTRACT FROM AUTHOR]

Osteosarcoma (OS) is a type of primary malignant cancer occurring in the bone and poses a threat to the lives of children and young adults. Long non‑coding RNAs (lncRNAs) have been certified to play important roles in various human malignant tumors, including OS. lncRNA KCNQ1OT1 has been investigated in certain types of cancer; however, its role and molecular mechanisms in OS remain to be determined. In the present study, a high KCNQ1OT1 expression was detected in human OS tissues and cell lines. Moreover, patients with OS with a high expression of KCNQ1OT1 presented a worse prognosis. Loss‑of‑function assays demonstrated that KCNQ1OT1 silencing suppressed cell proliferative, migratory and invasive abilities in OS. Importantly, the knockdown of KCNQ1OT1 suppressed the Wnt/β‑catenin signaling pathway in OS. In vivo assays displayed the inhibitory role of the silencing of KCNQ1OT1 in OS tumor growth. As regards the underlying mechanisms, KCNQ1OT1 could sponge miR‑3666, and its expression was negatively associated with that of miR‑3666 in OS tissues. Thereafter, Kruppel‑like factor 7 (KLF7), upregulated in OS tissues and cells, was discerned as a target gene of miR‑3666. Furthermore, KLF7 expression negatively correlated with miR‑3666 expression, whereas it positively correlated with KCNQ1OT1 expression. A rescue assay delineated that the overexpression of KLF7 counteracted the KCNQ1OT1 knockdown‑induced suppression of OS cell proliferation, migration, invasion and Wnt/β‑catenin signaling. Collectively, the present study demonstrates that KCNQ1OT1 facilitates OS progression and activates Wnt/β‑catenin signaling by targeting the miR‑3666/KLF7 axis.

Introduction: MicroRNA (miRNA) is known to be involved in nerve injury. Our study aimed to identify the role and mechanism of miR-1b in rat neuron proliferation, regeneration and apoptosis., Material and Methods: Neurons were successfully separated and identified using a microscope and immunofluorescence staining of microtubule-associated protein 2 (MAP-2). The expressions of miR-1b and Krüppel-like factor 7 (KLF7) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Neuron viability and apoptosis were detected by MTT assay and flow cytometry, respectively. Neuron regeneration states were observed using a microscope and analysed by the ImageJ software. Expressions of C-caspase-3 and cell regeneration-related proteins (nerve growth factor [NGF], ciliary neurotrophic factor [CNTF] and brain-derived neurotrophic factor [BDNF]) were measured by Western blot. Target genes and potential binding sites of KLF7 and miR-1b were predicted by TargetScan 7.2 and confirmed by dual luciferase reporter assay., Results: Neurons were identified as MAP-2-positive. Up-regulation of miR-1b reduced neuron viability and regenerative ability, promoted neuron apoptosis and C-caspase-3 expression, and down-regulated the expressions of cell regeneration-related proteins. KLF7 was the target gene of miR-1b. Overexpressed KLF7 rescued the effects of up-regulation of miR-1b on neuron viability, regeneration and apoptosis. Expressions of NGF, CNTF and BDNF were suppressed yet C-caspase-3 expression was up-regulated by miR-1b mimic, which was partially rescued by overexpressed KLF7., Conclusions: Up-regulation of miR-1b promoted rat neuron proliferation and regeneration yet inhibited apoptosis via targeting KLF7.

The biological relation between ubiquitin ligases and their substrates has been largely unclear. We previously developed a method-differential proteomics-based identification of ubiquitylation substrates (DiPIUS)-for the comprehensive identification of substrates for a given ubiquitin ligase. We have now applied DiPIUS to the F-box protein Fbxw7 in three cell lines (mHepa, Neuro2A and C2C12) and thereby identified Krüppel-like factor 7 (KLF7) as a candidate substrate of the SCF Fbxw7 ubiquitin ligase complex. KLF7 was shown to interact with Fbxw7 and to undergo Fbxw7-mediated polyubiquitylation. The stability of KLF7 was increased by depletion of Fbxw7, mutation of a putative Cdc4 phosphodegron (CPD) of KLF7 or exposure to inhibitors of glycogen synthase kinase-3 (GSK-3). Over-expression of Fbxw7 in Neuro2A cells down-regulated expression of the p21 Cip1 gene, which is a transcriptional target of KLF7 in neuronal differentiation and maintenance. Despite the presence of an almost identical CPD sequence in KLF6, the closest paralog of KLF7, mutation of this sequence affected neither the interaction of KLF6 with Fbxw7 nor its half-life. Our results suggest that KLF7, but not KLF6, is a bona fide substrate of SCF Fbxw7 , and that control of KLF7 abundance by SCF Fbxw7 might contribute to the regulation of neuronal differentiation and maintenance., (© 2019 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Krüppel-like factor 7 (KLF7) is a member of the KLF family of zinc finger transcription factors, and was the first KLF cloned using complementary DNA and polymerase chain reaction (PCR) techniques with human vascular endothelial cells as a template. In addition, KLF7 is known as the ubiquitous Krüppel-like factor, as it is widely expressed in numerous human tissues at low levels. In the present study, the function of KLF7 in migration and epithelial-mesenchymal transition (EMT), which are associated with tumor progression, was investigated in human oral squamous cell carcinoma (OSCC) cells. Genes that were differentially expressed in normal vs. OSCC tissue were identified in the Gene Expression Omnibus database, which identified upregulation of KLF7 in OSCC. The expression and subcellular location of KLF7 was then analyzed using immunohistochemistry. KLF7 expression was measured in three OSCC cell lines, and the two cell lines with the highest (HN13) and lowest (CAL27) KLF7 expression were selected for further analysis. Subsequently, HN13 cells with reduced KLF7 expression (sh-HN13) and CAL27 cells overexpressing KLF7 (OE-CAL27) were constructed. Transwell migration and wound healing assays were then used to analyze the migration of the cells. In addition, mRNA and protein expression levels of the EMT markers E-cadherin, N-cadherin, vimentin and snail were detected using reverse transcription-quantitative PCR and western blotting. KLF7 overexpression in OSCC was validated using tissue immunohistochemistry, which identified moderate to high cytoplasmic staining of KLF7 in OSCC cells. KLF7 knockdown and overexpression altered the migration ability of sh-HN13 and OE-CAL27 cells, which decreased and increased significantly respectively. Expression of E-cadherin, N-cadherin, vimentin and snail was markedly altered in sh-HN13 and OE-CAL27 cells, indicating changes in EMT status. The results of the present study suggest that KLF7 overexpression changes the migratory behavior of OSCC cells, and induces EMT and lymph node metastasis through the expression of snail.