Krüppel-like factor 6 (KLF6) requires its amino terminal domain to promote villous trophoblast cell fusion.

INTRODUCTION: Villous cytotrophoblast (vCTB) cells fuse to generate and maintain the syncytiotrophoblast layer required for placental development and function. Krüppel-like factor 6 (KLF6) is a ubiquitous transcription factor with an N-terminal acidic transactivation domain and a C-terminal zinc finger DNA-binding domain. KLF6 is highly expressed in placenta, and it is required for proper placental development. We have demonstrated that KLF6 is necessary for cell fusion in human primary vCTBs, and in the BeWo cell line. MATERIALS AND METHODS: Full length KLF6 or a mutant lacking its N-terminal domain were expressed in BeWo cells or in primary vCTB cells isolated from human term placentas. Cell fusion, gene and protein expression, and cell proliferation were analyzed. Moreover, Raman spectroscopy and atomic force microscopy (AFM) were used to identify biochemical, topography, and elasticity cellular modifications. RESULTS: The increase in KLF6, but not the expression of its deleted mutant, is sufficient to trigger cell fusion and to raise the expression of ß-hCG, syncytin-1, the chaperone protein 78 regulated by glucose (GRP78), the ATP Binding Cassette Subfamily G Member 2 (ABCG2), and Galectin-1 (Gal-1), all molecules involved in vCTB differentiation. Raman and AFM analysis revealed that KLF6 reduces NADH level and increases cell Young's modulus. KLF6-induced differentiation correlates with p21 upregulation and decreased cell proliferation. Remarkable, p21 silencing reduces cell fusion triggered by KLF6 and the KLF6 mutant impairs syncytialization and decreases syncytin-1 and ß-hCG expression. DISCUSSION: KLF6 induces syncytialization through a mechanism that involves its regulatory transcriptional domain in a p21-dependent manner.

Expression of the bovine KLF6 gene polymorphisms and their association with carcass and body measures in Qinchuan cattle (Bos Taurus).

Kruppel-like factor 6 (KLF6) genes plays a significant role in the regulation of cell differentiation, proliferation and muscle development. The aim of this study is to investigate the genetic variation and the haplotype combination of the KLF6 gene in Qinchuan cattle and verify its contribution to bovine carcass traits and body measurements. The data were analyzed by real-time quantitative PCR (qPCR) to detect the expression profile of the KLF6 gene in the various tissues of Qinchuan cattle. PCR amplicons sequencing explored three novel SNPs at loci 3332C > G; 3413C > T and 3521G > A in the 2nd exon region of the KLF6 gene. The expression of KLF6 in the liver, kidney and lung was greater than that of other tissues. Allelic and genotypic frequencies of these SNPs were found to be in Hardy Weinberg equilibrium (P < 0.05). In SNP1, genotype CC, in SNP2, genotype CT and in SNP3 genotype GG were associated (P < 0.05) with larger body and carcass measurements. Association analysis results indicated that individuals with the Hap1/4 diplotype had a longer body and rump, were taller at the withers, and were wider at the hip than the other combinations. In terms of ultrasound carcass measures, Hap1/4 was associated with a larger muscle area and more intramuscular fat than other combinations. The bioinformatics study of the KLF6 protein showed a high degree of conservation in different mammalian species. The above results suggest that the KLF6 gene can used as potential candidate markers gene for the beef breed improvement through marker assisted selection of Qinchuan cattle.

MicroRNA-191 promotes hepatocellular carcinoma cell proliferation by has_circ_0000204/miR-191/KLF6 axis.

OBJECTIVES: MicroRNAs are powerful regulators in hepatocellular carcinoma (HCC) tumorigenesis. MicoRNA-191 (miR-191) has been reported to play an important role in HCC, However, the regulatory mechanism is still unclear. In this study, we investigated the role of miR-191 in HCC and studied its underlying mechanisms of action. MATERIALS AND METHODS: The expression of miR-191 in HCC tissues was determined by quantitative real-time PCR (qRT-PCR). The role of miR-191 in HCC cells was examined by using both in vitro and in vivo assays. Downstream targets of miR-191 were determined by qRT-PCR and Western blot analysis. Dual-luciferase assays were performed to validate the interaction between miR-191 and its targets. RESULTS: The expression of miR-191 was significantly higher in HCC patients and a higher miR-191 expression predicted poorer prognosis. Analysis of The Cancer Genome Atlas data sets suggested that miR-191 positively correlated with cell cycle progression. Gain and loss of function assays showed that miR-191 promoted cell cycle progression and proliferation. Luciferase reporter assay showed that miR-191 directly targeted the 3'-untranslated region of KLF6 mRNA. Furthermore, circular RNA has_circ_0000204 could sponge with miR-191, resulting in inactivation of miR-191. CONCLUSIONS: Our study sheds light on the novel underlying mechanism of miR-191 in HCC, which may accelerate the development of cancer therapy.

MiR-630 promotes epithelial ovarian cancer proliferation and invasion via targeting KLF6.

OBJECTIVE: MicroRNAs play critical roles in post-translational gene expression. The current study was to investigate the effects of miR-630 in epithelial ovarian cancer. PATIENTS AND METHODS: Thirty epithelial ovarian cancer tissue and thirty normal ovarian tissue samples were collected and were detected miR-630 expression level with qRT-PCR. MiR-630 mimics, inhibitors and negative controls were transfected into SKOV3 and Cell Counting Kit-8 (CCK-8) assay, and transwell experiment were performed to detect the proliferation rate and migration, respectively. The luciferase reporter assay was utilized to identify miR-630's target gene. Balb/c nude mice were utilized to verify the effect of miR-630 in vivo. RESULTS: QRT-PCR showed a significantly high miR-630 expression in epithelial ovarian cancer relative to normal ovarian tissue. The miR-630 overexpression promoted epithelial ovarian cancer cell SKOV3 proliferation and migration. Krüppel-like factor 6 (KLF6) was predicted as the target of miR-630. In vivo study also verified that miR-630 overexpression stimulated ovarian cancer growth. CONCLUSIONS: We propose that targeting miR-630 might be a promising therapeutic approach for ovarian cancer.

Functional and Structural Insights of the Zinc-Finger HIT protein family members Involved in Box C/D snoRNP Biogenesis.

Zf­HIT family members share the zf­HIT domain (ZHD), which is characterized by a fold in "treble-clef" through interleaved CCCC and CCHC ZnF motifs that both bind a zinc atom. Six proteins containing ZHD are present in human and three in yeast proteome, all belonging to multimodular RNA/protein complexes involved in gene regulation, chromatin remodeling, and snoRNP assembly. An interesting characteristic of the cellular complexes that ensure these functions is the presence of the RuvBL1/2/Rvb1/2 ATPases closely linked with zf­HIT proteins. Human ZNHIT6/BCD1 and its counterpart in yeast Bcd1p were previously characterized as assembly factors of the box C/D snoRNPs. Our data reveal that the ZHD of Bcd1p is necessary but not sufficient for yeast growth and that the motif has no direct RNA-binding capacity but helps Bcd1p maintain the box C/D snoRNAs level in steady state. However, we demonstrated that Bcd1p interacts nonspecifically with RNAs depending on their length. Interestingly, the ZHD of Bcd1p is functionally interchangeable with that of Hit1p, another box C/D snoRNP assembly factor belonging to the zf­HIT family. This prompted us to use NMR to solve the 3D structures of ZHD from yeast Bcd1p and Hit1p to highlight the structural similarity in the zf­HIT family. We identified structural features associated with the requirement of Hit1p and Bcd1p ZHD for cell growth and box C/D snoRNA stability under heat stress. Altogether, our data suggest an important role of ZHD could be to maintain functional folding to the rest of the protein, especially under heat stress conditions.