The structure of the Klf4 DNA-binding domain links to self-renewal and macrophage differentiation

Schuetz, A.; Nana, Didier; Rose, Charlotte; Zocher, Georg; Milanovic, Maja; Koenigsmann, Jessica; Blasig, Rosel; Heinemann, Udo; Carstanjen, Dirk

doi:10.1007/s00018-010-0618-x

Cited by 76 publications

(97 citation statements)

References 44 publications

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“…Previous studies showed that the second zinc finger recognizes the GTG or GCG triplet in the corebinding site, whereas the third zinc finger plays an important role in overall binding site preferences (56,57). Among the KLFs analyzed in this study, the third zinc finger shows the greatest variation in amino acid sequence.…”

Section: Regulation Of Epha5 Expression By Klfs and Dna Methylation-tmentioning

confidence: 71%

Novel Roles and Mechanism for Krüppel-like Factor 16 (KLF16) Regulation of Neurite Outgrowth and Ephrin Receptor A5 (EphA5) Expression in Retinal Ganglion Cells

Wang¹,

Galvão

Beach³

et al. 2016

Journal of Biological Chemistry

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Regenerative medicine holds great promise for the treatment of degenerative retinal disorders. Krüppel-like factors (KLFs) are transcription factors that have recently emerged as key tools in regenerative medicine because some of them can function as epigenetic reprogrammers in stem cell biology. Here, we show that KLF16, one of the least understood members of this family, is a POU4F2 independent transcription factor in retinal ganglion cells (RGCs) as early as embryonic day 15. When overexpressed, KLF16 inhibits RGC neurite outgrowth and enhances RGC growth cone collapse in response to exogenous ephrinA5 ligands. Ephrin/EPH signaling regulates RGC connectivity. The EphA5 promoter contains multiple GC-and GT-rich KLF-binding sites, which, as shown by ChIP-assays, bind KLF16 in vivo. In electrophoretic mobility shift assays, KLF16 binds specifically to a single KLF site near the EphA5 transcription start site that is required for KLF16 transactivation. Interestingly, methylation of only six of 98 CpG dinucleotides within the EphA5 promoter blocks its transactivation by KLF16 but enables transactivation by KLF2 and KLF15. These data demonstrate a role for KLF16 in regulation of RGC neurite outgrowth and as a methylation-sensitive transcriptional regulator of EphA5 expression. Together, these data identify differential low level methylation as a novel mechanism for regulating KLF16-mediated EphA5 expression across the retina. Because of the critical role of ephrin/EPH signaling in patterning RGC connectivity, understanding the role of KLFs in regulating neurite outgrowth and Eph receptor expression will be vital for successful restoration of functional vision through optic nerve regenerative therapies. Progressive loss of retinal ganglion cells (RGCs)2 leads to blindness in diseases such as glaucoma, the leading cause of irreversible bilateral blindness worldwide (1). Current treatments can slow progression but cannot restore lost vision. Recent advances in generating retinal cells from embryonic and induced pluripotent stem cells have stimulated interest in the potential for neuronal regeneration to treat blindness (2-4). For successful optic nerve regeneration, newly regenerated RGCs will need to extend axons considerable distances to reach the visual centers of the brain, where they must make the correct patterns of termination on post-synaptic neurons to reestablish retinotopic maps. Thus, it is critical to understand the mechanisms that regulate expression of molecules involved in axonal growth and connectivity.Krüppel-like factors (KLFs) are members of the SP/KLF family of transcription factors that are characterized by three zinc finger DNA binding domains near their C terminus (5). KLFs bind to GT-or GC-rich sequences and can function as either transcriptional activators or repressors, depending on the promoter context (6). At least 15 of the 17 members of the KLF transcription factor family are expressed in RGCs, and overexpression of different KLF transcription factors in RGCs, hippocampal neurons, or cor...

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Section: Regulation Of Epha5 Expression By Klfs and Dna Methylation-tmentioning

confidence: 71%

Novel Roles and Mechanism for Krüppel-like Factor 16 (KLF16) Regulation of Neurite Outgrowth and Ephrin Receptor A5 (EphA5) Expression in Retinal Ganglion Cells

Wang¹,

Galvão

Beach³

et al. 2016

Journal of Biological Chemistry

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“…The defining feature of KLFs is the presence of a conserved, tandem repeat of three Cys 2 -His 2 -type zinc fingers at their C termini, through which KLFs make sequence-specific contacts with GC-rich and CACCC-related elements of DNA (3)(4)(5)(6). While as a whole this domain is highly conserved within the family, specific amino acid differences between the various KLFs result in different DNA-binding preferences (7). As such, individual KLFs have been shown to regulate overlapping, but also distinct, sets of target genes (8,9).…”

mentioning

confidence: 99%

Generation of Mice Deficient in both KLF3/BKLF and KLF8 Reveals a Genetic Interaction and a Role for These Factors in Embryonic Globin Gene Silencing

Funnell

Mak

Twine

et al. 2013

Molecular and Cellular Biology

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f Krüppel-like factors 3 and 8 (KLF3 and KLF8) are highly related transcriptional regulators that bind to similar sequences of DNA. We have previously shown that in erythroid cells there is a regulatory hierarchy within the KLF family, whereby KLF1 drives the expression of both the Klf3 and Klf8 genes and KLF3 in turn represses Klf8 expression. While the erythroid roles of KLF1 and KLF3 have been explored, the contribution of KLF8 to this regulatory network has been unknown. To investigate this, we have generated a mouse model with disrupted KLF8 expression. While these mice are viable, albeit with a reduced life span, mice lacking both KLF3 and KLF8 die at around embryonic day 14.5 (E14.5), indicative of a genetic interaction between these two factors. In the fetal liver, Klf3 Klf8 double mutant embryos exhibit greater dysregulation of gene expression than either of the two single mutants. In particular, we observe derepression of embryonic, but not adult, globin expression. Taken together, these results suggest that KLF3 and KLF8 have overlapping roles in vivo and participate in the silencing of embryonic globin expression during development.

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“…The zinc finger binds to a DNA triplet (blue) through -1, +3 and +6 amino acid residues (yellow) of the finger (relative to the start of the α-helix). Figure was generated with Cn3D (NCBI) from the structure of KLF4 (zf2) bound to DNA; PBD ID 2WBU (Schuetz et al, 2011).…”

Section: Common Structures Of Klfsmentioning

confidence: 99%

“…2WBU) (Schuetz et al, 2011) was used to interpret KLF1-DNA interaction, as DNA-contacting residues from ZF2 of KLF4 and KLF1 are identical. The Nan-KLF1 and CDA-KLF1 mutant conformations was modelled by substituting E339 for D and K, respectively, with the sidechain conformation chosen from a backbone-dependent rotamer library (Dunbrack & Karplus, 1993).…”

Section: Structural Predictionsmentioning

confidence: 99%

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Investigating the Molecular Pathogenesis of Inherited Childhood Anaemia

Ilsley¹

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Krüppel-like factors (KLFs) are a group of 17 transcription factors with diverse functions such as early embryo patterning, organogenesis and tissue homoeostasis. All 17 members contain highlyconserved DNA-binding domains consisting of three C-terminal C2H2-type zinc fingers. This high level of conservation allows all members of the KLF family to bind to a 9 base pair CACCC-box DNA sequence (CCM CRC CCN) which is commonly found in many tissue-specific gene promoters and enhancers. The activity of each member of the KLF family is largely determined by a more variable N-terminal domain through which they interact with co-factors such as p300 and C-terminal binding protein 2 (CtBP2).Many KLFs are often expressed in the same cells. In these situations, they can form transcriptional networks where they coordinate to control developmental programs. For example, KLF2, KLF4, and KLF5 work redundantly in embryonic stem cells where they regulate key pluripotency genes such as Nanog to maintain the 'stemness' of the cell. Multiple KLFs are expressed during erythropoiesis, the ongoing production of red blood cells, where it is likely that they form transcriptional networks to control differentiation.Krüppel-like factor 1 (KLF1) is an erythroid-specific transcription factor that is responsible for coordinating the expression of a large cohort of genes required for nearly all aspects of erythropoiesis. As the founding member of the KLF group, KLF1 contains three C-terminal C2H2-type zinc fingers that bind to DNA and an N-terminal proline-rich activation domain that interacts with co-factors to initiate transcription of target genes.This thesis investigates the molecular mechanisms of Klf1 binding specificity to DNA and how this is affected by mutations, and by competition between family members.A disease that is known as Congenital Dyserythropoietic anaemia type IV (CDAIV) is a rare autosomal-dominant inherited erythrocyte disorder characterised by ineffective erythropoiesis and haemolysis. CDAIV is caused by a point mutation in the second zinc finger of KLF1 which results in substitution of an amino acid predicted to bind to DNA.A mouse carrying a point mutation in the corresponding amino acid residue in murine Klf1 to that mutated in CDAIV patients has been previously generated in an ENU screen. This mouse displays a semi-dominant haemolytic anaemia and is named 'Nan' for 'neonatal anaemia'. The Nan mouse shares a similar phenotype to that of patients with CDAIV and hereditary spherocytosis.Incredibly, both the 'Nan' and CDAIV mutations cause disease which is far more severe than that of commonly occurring Klf1 haploinsufficiency.ii ChIP-seq analysis in cell lines developed to model Klf1 mutations identified aberrant DNAbinding events genome-wide for both the mouse 'Nan' mutation and the human CDAIV mutation.Next-generation sequencing of newly-synthesised RNA (4sU-RNA-seq) reveals ectopic transcriptional consequences of this aberrant binding. Novel sequence specificity of the mutant protein was confirmed by performi...

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The structure of the Klf4 DNA-binding domain links to self-renewal and macrophage differentiation

Cited by 76 publications

References 44 publications

Novel Roles and Mechanism for Krüppel-like Factor 16 (KLF16) Regulation of Neurite Outgrowth and Ephrin Receptor A5 (EphA5) Expression in Retinal Ganglion Cells

Novel Roles and Mechanism for Krüppel-like Factor 16 (KLF16) Regulation of Neurite Outgrowth and Ephrin Receptor A5 (EphA5) Expression in Retinal Ganglion Cells

Generation of Mice Deficient in both KLF3/BKLF and KLF8 Reveals a Genetic Interaction and a Role for These Factors in Embryonic Globin Gene Silencing

Investigating the Molecular Pathogenesis of Inherited Childhood Anaemia

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