USF1
Upstream stimulatory factor 1 is a protein that in humans is encoded by the USF1 gene.
Gene
The upstream stimulatory factor gene encodes a transcription factor USF that belongs to the proto-oncogene MYC family and features a basic helix-loop-helix leucine zipper motif in the protein structure. USF was originally identified as regulating the major late promoters of adenovirus, and recent research has further revealed its role in tissue protection. The bHLH-LZ motif enables the transactivation capacity of the USF protein through interacting with the initiator element and E-box motif on the bound DNA. In the context of insulin and glucose-induced USF activities, those E-box motifs can act as a glucose-responsive element and a part of the carbohydrate response element to interact with transcription factors.Isoforms
USF comprises two major isoforms: USF1 and USF2. The USF1 gene is located on the chromosome region 1q22-q23 in both humans and mice; the USF2 gene is located on the chromosome 19q13 in humans and chromosome 19q7 in mice, respectively. Both USF1 and USF2 transcripts comprise 10 exons and can undergo exon 4-excision during alternative splicing. From an auto-regulation perspective, these exon 4-excision products act as dominant negative regulators and are found to suppress USF-dependent gene expression.Protein
Although USF1 and USF2 share 70% of the amino acid sequence in their bHLH-LZ region, only 40% similarity is found in their full-length proteins. In addition, USF1 and USF2 exhibit different protein abundances in a cell type-specific manner. It has been found that USF1 and USF2 expression increases during the differentiation of erythroid cells. Despite the ubiquitous expression of both isoforms, USF1 and USF2 mediate different biological processes and functions in cells. While USF1 modulates metabolism, immune response, and tissue protection, USF2 primarily controls embryonic development, brain function, iron metabolism, and fertility. Structurally, the highly conserved bHLH-LZ structure on the C-terminus of USF yields high binding specificity and promotes the formation of USF1 homodimers or USF1-USF2 heterodimers for DNA binding. The USF-specific region on the N-terminal region, on the other hand, facilitates the nuclear translocation and activation of USF1.Function
The USF1 gene encodes a member of the basic helix-loop-helix leucine zipper family and can function as a cellular transcription factor. The encoded protein can activate transcription through pyrimidine-rich initiator elements and E-box motifs. This gene has been linked to familial combined hyperlipidemia. Two transcript variants encoding distinct isoforms have been identified for this gene.A study of mice suggested that reduced USF1 levels increase metabolism in brown fat.
Regulation
Modulation of DNA binding affinity
The symmetrical E-box motif is the main target of bHLH-LZ transcription factors, and USF1 has a high binding affinity for the core sequence CACGTG in the motif. USF1-DNA binding activity can be modulated by cell type-specific DNA methylation and acetylation on the E-box motif or by post-transcriptional modifications of the USF1 protein. For example, CpG methylation on the central E-box motif inhibits the complex formation of USF1 with its co-transcription factors and therefore decreases the corresponding gene expression in mouse lymphosarcoma cells. In contrast, phosphorylation of USF1 by p38 mitogen-activated protein kinases, protein kinase A, or protein kinase C increases its binding to the E-box motif and activates gene transcription.Phosphorylation
phosphorylates serine and threonine residues of substrate proteins and converts extracellular signals induced by growth factors, mitogens, or cytokines into intracellular phosphorylation cascades, which regulate cell proliferation, differentiation, stress responses, and apoptosis.Phosphorylation by MAPKs induce a conformational change of the USF protein and exposes its DNA-binding domain for interaction. This increased structural exposure enhances DNA binding and therefore the transcriptional activity of USF.
- ERK1 and ERK2 phosphorylate USF1 in response to TFG-β signaling in vascular smooth muscle cells. SMAD2 and SMAD3 signaling, following the TFG-β receptor activation, can also cooperate with EGFR / ERK pathways to activate USF1, which in turn regulates the gene expression of plasminogen activator inhibitor-1, a significant biomarker and predictor of cardiovascular disease-related death and a marker of poor prognosis in breast cancer.
- Casein kinase 2 or CK-II is a tetrameric enzyme composed of two catalytic and two regulatory subunits. In pancreatic cells, CK2 phosphorylates USF1, PDX1, and MST1 to suppress insulin expression.
| Proteins mediating USF1 modification | |
| Phosphorylation | p38, pKA and pKC, ERK1/2, DNA-PK |
| Acetylation | PCAF |
| Methylation' | SET7/92 |
| USF1-interacting proteins | |
| Transcription co-factors | USF2, SP1, PEA3, MTF1, SREBP1-c, MED17, BAF60 |
Gene transcription
- Transforming growth factor β 1 is encoded by the TFGB1 gene, which contains an E-box within the promoter region and has been implicated in excessive extracellular matrix accumulation under a high-glucose condition. Overexpression of either USF1 or USF2 is found to elevate the TFGB1 promoter activity in human embryonic kidney cells. However, only USF1 overexpression leads to increased TGF-β1 secretion.
- Thrombospondin 1 is involved in the development of diabetic nephropathy. USF1/2 binds to the E-box motif on the human THBS1 promoter and regulates high-glucose-induced TSP1 expression in mesangial cells. USF2 overexpression has been found to augment THBS1 promoter activity and TSP1 expression. The resulting increase in TSP1 expression further promotes the formation of active TGF-β.
- AP-1 transcription factor refers to a complex of dimeric transcription factors composed of c-Jun, c-Fos, or activating transcription
Interaction between USF1 and other transcription factors, including SP1, PEA3 and MTF1, also leads to cooperative transcriptional regulation. For instance, the leucine zipper motif of USF1 recruits PEA3 to form a ternary complex and co-regulates the transcription of BAX, an apoptosis regulator. Another USF1-regulated target is topoisomerase III, which catalyzes the topological changes of DNA, modifies DNA supercoil structures, and increases the chromatin accessibility for gene expression. Similar interactions exist between USF1 and JMJD1C or H3K9 demethylase, in which the molecular interactions change chromatin accessibility and elevate the transcription of a series of lipogenic genes, including those for fatty acid synthase, acetyl-CoA carboxylase, ATP citrate synthase, and sterol regulatory element-binding protein 1.
Chromosome boundary by USF
Chromosomes are generally classified as euchromatin or heterochromatin with distinct histone modifications, compaction levels, and the resulting gene expression patterns. Heterochromatin is a tightly condensed and transcriptionally repressed chromatin domain that is characterized by distinct combinations of histone post-translational modifications. Heterochromatin is required for genome stability and gene expression regulation. However, it can spread into neighboring DNA regions and inactivate gene expression. Chromosome boundary elements are thus necessary to block such stochastic spreads of heterochromatin and maintain stable gene expression. USF1 and USF2 have been found to recruit various histone-modifying complexes, including the histone H3 methyltransferase Set1 complex and the H4 arginine 3 methyltransferase PRMT1, with the latter known to establish active chromatin domains. USF1/USF2 binding deposits a high level of activating histone modifications on adjacent nucleosomes and thus prevents the propagation of chromatin silencing modifications from the heterochromatin, such as H3K9 and K27 methylation.Other USF1/USF2-related chromatin modifications include the recruitment of the E3 ubiquitin ligase, RNF20, to monoubiquitinate histone H2B. The loss of RNF20 is found to cause an extension of the silencing modifications from the 16 kb heterochromatic domain into the β-globin locus. Moreover, USF1 and USF2 can bind to the 5' DNase I hypersensitive site HS4 and recruit an H3 acetyltransferase, PCAF, which blocks the heterochromatin spread into the β-globin locus.
FASN transactivates for lipogenesis
USF is known to bind the L-type pyruvate kinase promoter on DNA at high glucose and insulin levels. Excessive insulin activates kinases and phosphatases that post-translationally modify USF, sterol regulatory element-binding protein 1C, carbohydrate-responsive element-binding protein, and Liver X receptor. With insulin stimulation, USF1 and USF2 bind to the E-boxes at -332 and -65 in the promoter region of FASN that encodes fatty acid synthase for lipogenesis.Various post-translational modifications of USF1 determine its activity and signaling pathways, and can affect the lipogenesis process. An abnormal increase in the USF-mediated de novo fatty acid synthesis is found to cause intracellular fatty acid accumulation and to deregulate gene expression and cellular processes such as tumor cell survival.