Regulation of gene expression is central to a wide variety of cellular functions. The fate of a cell--whether it will be a muscle or fat cell, bone, or neuron--is determined by expression of genes in a temporal and tissue-specific manner. The cell's response to external stimuli, such as growth factors, hormones, infections, injury, and even stress, depends on the regulation of gene expression. The differential expression of genes in response to signals originating from the cell membrane determines whether a cell divides, differentiates, or dies.
Studies on the mechanism of gene expression have identified a group of proteins called transcription factors (TFs) as the central regulators of gene expression. These factors bind to specific regions of the DNA called promoters (since they promote transcription) and enhancers (which enhance transcription from a certain promoter).
Transcription factors can either stimulate or inhibit transcription of target genes by regulating the formation of messenger RNA (mRNA) and ultimately altering the production of specific proteins. Transcription in eukaryotic cells occurs in the nucleus and is catalyzed by RNA polymerase II and several associated TF proteins binding to this enzyme (also known as the core initiation complex). Basal or general TFs (TATA-binding protein [TBP] and TBP-associated factors, TFIIA, TFIIB, TFIIE, and TFIIF) bind to the promoter region (usually a sequence called a TATA box, although some genes have been found to be TATA-less) in an ordered manner to initiate transcription (Figure 1).
There are also ubiquitous TFs that recognize and bind to specific DNA sequences in the promoter to ensure efficient transcription. An example of this is the SpI factor. Inducible TFs are another group of factors that also bind to specific motifs found in target DNA sequences. These TFs are activated at specific times in specific cells by signals from the cell membrane, and thus, have a key regulatory function in gene expression. Examples are the early response genes (c-fos, c-jun, c-myc), the expression of which is induced by mitogenic signals.
Clinicians see the result of both normal and abnormal gene regulation in their daily routine: in clinical settings, such as wound healing; in the development of the embryo during pregnancy; and in human diseases, such as malignancy. Abnormal gene expression gives rise to cellular abnormalities in diseases, such as metabolic and developmental disorders. This abnormal gene expression can result from abnormal TF function. For instance, a mutation in the gene coding for a TF that plays an essential role in regulating gene expression in the pituitary (called Pit-1) has been identified in patients with combined pituitary hormone deficiency; this mutant gene results in growth deficiency and mental retardation.
In addition, aberrant expression of genes governing cell growth and differentiation (called proto-oncogenes) plays a critical role in the development of cancer. An example of this is Burkitt's lymphoma, in which the oncogene c-myc is deregulated; this results in rapid cell growth, leading to lymphoma. Mutant forms of steroid/thyroid hormone receptors, another class of TFs, have also been identified as the cause of resistance to hormones, such as the glucocorticoids.
The basic functional domains of TFs consist of a transcriptional action domain (activating or repressing), a DNA-binding domain, a dimerization domain, and a nuclear translocation signal (Figure 2A). There are several major classes of TFs based on these structural motifs (see Table 1): (1) helix-loop-helix (Myo-D); (2) leucine zipper (c-Jun); (3) zinc finger (glucocorticoid receptor); and (4) helix-turn-helix (homeobox). A recent review on the classification of TFs can be found in reference 5.
Helix-Loop-Helix--Members of the helix-loop-helix family of TFs oligomerize through a helix-loop-helix motif (two alpha-helices) and interact with DNA via an adjacent basic amino acid domain. Helix-loop-helix TFs include muscle-specific transactivators (Myo-D, myogenin, MRF-4, and myf-5). They form heterodimers that may result in the activation of muscle-specific genes. The proto-oncogenec-Myc is another example of a helix-loop-helix TF.
Leucine Zipper--The leucine zipper consists of an extended alpha-helix, which contains heptad repeats of leucines that interact with each other in a zipper-like conformation. Two proteins containing the leucine zipper motif form dimers consisting of identical proteins (homodimers) or different proteins (heterodimers). Members of the c-Jun and c-Fos family belong to this class and will be discussed in more detail below.
Zinc Finger--The zinc finger motif is formed by a small group of conserved amino acids (cysteines and histidines) binding a zinc ion in a finger-like structure. There are two types of DNA-binding proteins with this structure: the classic zinc finger proteins (TFIIIA required for transcription by RNA polymerase) and the steroid receptors (glucocorticoid receptor, estrogen receptor, retinoic acid receptors).
Helix-Turn-Helix--The helix-turn-helix motif consists of one alpha-helix making contact in the wide groove of the DNA while the other helix lies at an angle across the DNA. The helix-turn helix motif was originally identified in DNA-binding domains of phage repressors, but related forms of this motif have also been found in homeobox (genes that determine the identity of body structure) DNA-binding domains (ie, Pit-1, Oct-1).
Growth factors (eg, epidermal growth factor, platelet-derived growth factor) and cytokines (eg, interleukin-1 [IL-1], IL-6) are ligands that bind to specific receptors and trigger signaling cascades that ultimately lead to cellular proliferation, differentiation, or transformation (Figure 3). Several secondary messengers may be activated by such ligand-receptor combinations, such as phospholipase C-gamma, phosphoinositol kinase, guanosine triphosphate-binding protein ras, protein kinase C, protein kinase A, and other protein tyrosine kinases, such as mitogen-activated protein kinase (MAPK), extra- cellular-signal-regulated kinase (ERK), and c-Jun NH2 terminal kinase (JNK). These signaling cascades eventually lead to either activation or inactivation of nuclear TFs. Several pathways may activate the same TF, identifying TFs as key proteins that translate these varied signals into biologic action(s). Thus, they are important targets for the development of inhibitory agents for a wide variety of cellular functions.
To give a specific example of the broad subject of TF functions and activities, we will focus on studies done on the mammalian TF activating protein-1 (AP-1), which is actually a complex of the Jun and Fos family of proteins.[10,11] Activating protein-1 complexes are formed by dimers of Jun family members (c-Jun, Jun-B, Jun-D) or heterodimers of Jun family members with Fos family members (c-Fos, Fos B, Fra-1, and Fra-2). As shown in Figure 4, activating protein-1 complexes bind to a specific target DNA sequence TGAC/GTCA (also known as the tetradecanoyl phorbolacetate (TPA) response element, or TRE) found in the promoters of several cellular genes, such as human collagenase, stromelysin, and IL-2, and activate the transcription of these genes.
The AP-1 complex either positively or negatively regulates transcription of the target gene, depending on the composition of the heterodimers.[12-14] In response to different stimuli, such as growth factor stimulation, cellular stress, or even ultraviolet light stimulation, the expression and activity of Jun and Fos proteins are rapidly and transiently induced.[15-17] In addition, tumor-promoting agents, such as TPA, which is a potent activator of protein kinase C, have also been shown to act via the AP-1 pathway.[18,19]