Klf4 Directly Interacts with Oct4 and Sox2 to Promote Reprogramming

Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, Keck School of Medicine, University of Southern California, Los Angeles, California 90033, USA.
Stem Cells (Impact Factor: 6.52). 12/2009; 27(12):2969-78. DOI: 10.1002/stem.231
Source: PubMed

ABSTRACT Somatic cells can be reprogrammed to induced pluripotent stem (iPS) cells by ectopic expression of specific sets of transcription factors. Oct4, Sox2, and Klf4, factors that share many target genes in embryonic stem (ES) cells, are critical components in various reprogramming protocols. Nevertheless, it remains unclear whether these factors function together or separately in reprogramming. Here we show that Klf4 interacts directly with Oct4 and Sox2 when expressed at levels sufficient to induce iPS cells. Endogenous Klf4 also interacts with Oct4 and Sox2 in iPS cells and in mouse ES cells. The Klf4 C terminus, which contains three tandem zinc fingers, is critical for this interaction and is required for activation of the target gene Nanog. In addition, Klf4 and Oct4 co-occupy the Nanog promoter. A dominant negative mutant of Klf4 can compete with wild-type Klf4 to form defective Oct4/Sox2/Klf4 complexes and strongly inhibit reprogramming. In the absence of Klf4 overexpression, interaction of endogenous Klf4 with Oct4/Sox2 is also required for reprogramming. This study supports the idea that direct interactions between Klf4, Oct4, and Sox2 are critical for somatic cell reprogramming.

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Available from: Gang Bai, Sep 29, 2015
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    • "Constitutive expression of SOX2 maintains cellular self-renewal, and coordinated with OCT4 and NANOG supports ESC-pluripotency (Boyer et al 2005). Furthermore, ectopic KLF4 expression reprogrammes pluripotency in somatic cells (Wei et al 2009). The characterization of sphere-forming cells showed a regular and marked expression of two self-renewal transcription factors (SOX2 and KLF4) in relation to their matched counterparts, at both gene and protein levels. "
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    ABSTRACT: Osteosarcoma is a bone tumor displaying significant cellular and histological heterogeneity and a complex genetic phenotype. Although multiple studies strongly suggest the presence of cancer stem cells in osteosarcoma a consensus on their characterization is still missing. We used a combination of functional assays (sphere-forming, Aldefluor and side-population) for identification of cancer stem cell populations in osteosarcoma cell lines. Expression of stemness-related transcription factors, quiescent nature, in vivo tumorigenicity and Wnt/β-catenin activation were evaluated. We show that different cancer stem cell populations may co-exist in osteosarcoma cell lines exhibiting distinct functional properties. Osteosarcoma spheres are slowly-proliferating populations, overexpress SOX2 and KLF4 stemness-related genes and have enhanced tumorigenic potential. Additionally, spheres show specific activation of Wnt/β-catenin signaling as evidenced by increased nuclear β-catenin, TCF/LEF activity and AXIN2 expression, in a subset of the cell lines. Aldefluor-positive populations were detected in all osteosarcoma cell lines and overexpress SOX2, but not KLF4. The side-population phenotype is correlated with ABCG2 drug-efflux transporter expression. Distinct functional methods seem to identify cancer stem cells with dissimilar characteristics. Intrinsic heterogeneity may exist within osteosarcoma cancer stem cells and can have implications on the design of targeted therapies aiming to eradicate these cells within tumors.
    Journal of Cellular Physiology 09/2015; DOI:10.1002/jcp.25179 · 3.84 Impact Factor
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    • "There is ample evidence for possible involvement of several other proteins in organization of long-range interactions, including the transcription factor SP1 that contains C2H2-type zinc finger DNA-binding domain and glutamine-rich dimerization domain (Courey et al., 1989; Mastrangelo et al., 1991; Su et al., 1991), the transcription factor Klf4 (Wei et al., 2013) that interact with many transcription regulators, including Oct4 and Sox2 (Wei et al., 2009), general activator p300/CBP, and repressors such as HDAC and CtBP (Swamynathan, 2010), MAR-binding protein SATB1 (Cai et al., 2006; Gong et al., 2011), TFIIIC (Kirkland et al., 2013), and condensins (D’Ambrosio et al., 2008). In any case however, it would be premature to arrive at any definitive conclusions about the role of these proteins in the chromosome architecture. "
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    ABSTRACT: Due to advances in genome-wide technologies, consistent distant interactions within chromosomes of higher eukaryotes have been revealed. In particular, it has been shown that enhancers can specifically and directly interact with promoters by looping out intervening sequences, which can be up to several hundred kilobases long. This review is focused on transcription factors that are supposed to be involved in long-range interactions. Available data are in agreement with the model that several known transcription factors and insulator proteins belong to an abundant but poorly studied class of proteins that are responsible for chromosomal architecture.
    Frontiers in Genetics 02/2014; 5:28. DOI:10.3389/fgene.2014.00028
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    • "RNA interference experiments confirm that Klf4 is redundant with two other family members, Klf2 and Klf5, in regulating expression of pluripotency related genes. In ES cells, Klf4 has been shown to be important to activate Lefty1 together with Oct4 and Sox2 (Chan et al., 2009; Nakatake et al., 2006; Wei et al., 2009). Genome-wide chromatin immunoprecipitation with microarray analysis (ChIP-Chip) demonstrates that the DNA binding profile of Klf4 overlaps with that of Oct4 and Sox2 on promoters of genes specifically underlying establishment of iPSCs (Nakatake et al., 2006), suggesting transcriptional synergy among these factors. "
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    ABSTRACT: Embryonic stem (ES) cells are pluripotent cells with the capacity for unlimited self-renewal or differentiation. Inhibition of MAPK pathways enhances mouse ES cell pluripotency characteristics. Compared to wildtype ES cells, jnk2(-/-) ES cells displayed a much higher growth rate. To determine whether JNKs are required for stem cell self-renewal or differentiation, we performed a phosphorylation kinase array assay to compare mouse ES cells under LIF+ or LIF- culture conditions. The data showed that activation of JNKs was induced by LIF withdrawal. We also found that JNK1 or 2 phosphorylated Klf4 at threonines 224 and 225. Activation of JNK signaling and phosphorylation of Klf4 inhibited Klf4 transcription and transactivation activity. Importantly, jnk1(-/-) and jnk2(-/-) murine embryonic fibroblasts (MEFs) exhibited a significantly greater potency in the ability to increase the number of iPS colonies compared with jnk wildtype MEFs. Overall, our results demonstrated that JNK1 and 2 play a negative role in reprogramming to pluripotent stem cells by suppressing Klf4 activity.
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