Question

Discuss how promoters, enhancers and the TF proteins that interact with them are used in eukaryotic cells to control transcription of housekeeping genes versus cell specific/regulated genes? Be specific about everything’s role – what binds where and does what!

A) 4pts - Regulated cell specific genes -

B) 4pts - Housekeeping genes –

Answer 1

Much of the complexity in differentiation in animal and plant cells can be attributed to the evolution of elaborate systems made up of short (6 to 8 base pair) cis-regulatory DNA sequences or motifs, as well as the TFs that bind to the motifs, interact with each other to form complexes, and recruit RNA polymerase II. Most eukaryotic genes have promoters that consist of the TATA box close to the 5' end of the gene and, farther upstream, several motifs recognized by specific transcription factors. In addition, many genes have one or more other nearby sequences called enhancers. Enhancers affect transcription; these sequences occur upstream, downstream, or within introns, and they continue to work whether in the normal orientation or turned backward in the genome. In yeast, no enhancers are known; instead, there are only upstream activator sequences (UASs). Enhancers can be found thousands of base pairs from a promoter, whereas UASs are generally within a few hundred base pairs upstream. Typical RNA polymerase II promoters can be influenced by many enhancers and by multiple factors bound to the promoter and enhancer sequences.

The mode of action of TFs is to recognize and bind to a segment of DNA in the promoter and/or enhancer region. Often, a change in the conformation, or three-dimensional structure of a TF, will accompany DNA binding. For example, the two loops in NFATC1 that interact with DNA are found in different conformations, depending on whether NFATC1 is complexed with DNA or not. Moreover, the structure of different TF families, described later in this article, results in specific areas in these protein complexes that interact with the DNA recognition motif. The recognition motif is usually only about 6 to 10 base pairs long.

Experiments have shown that TFs can bind tightly, both within cells and in vitro. After TFs bind to promoter or enhancer regions of the DNA, they interact with other bound TFs and recruit RNA polymerase II. Their influence, however, can be either positive or negative, depending on the presence of other functional domains on the protein and the overall impact of the entire TF complex. A typical TF has multiple functional domains, not only for recognizing and binding to the appropriate DNA strand, but also for interactions with other TFs, with proteins called coactivators, with RNA polymerase II, with chromatin remodeling complexes, and with small noncoding RNAs.

TFs control many important parts of development; therefore, organisms with a deletion of a TF gene exhibit profound irregularities in organization and development. For example, in Drosophila, deletion of the TF antennapedia gene results in the development of the antennal imaginal disc into legs rather than antennae.

A) Regulated cell specific genes

Cell-specific gene regulation is often controlled by specific combinations of DNA binding sites in target enhancers or promoters. Thus, SPS+A is an architectural DNA transcription code that programs a cell-specific pattern of gene expression

Notch signaling in Drosophila proneural clusters, cell-specific activation of certain Notch target genes is known to require transcriptional synergy between the Notch intracellular domain (NICD) complexed with CSL proteins bound to "S" DNA sites and proneural bHLH activator proteins bound to nearby "A" DNA sites. Previous studies have implied that arbitrary combinations of S and A DNA binding sites (an "S+A" transcription code) can mediate the Notch-proneural transcriptional synergy.

Gene regulation is how a cell controls which genes, out of the many genes in its genome, are "turned on" (expressed). Thanks to gene regulation, each cell type in your body has a different set of active genes – despite the fact that almost all the cells of your body contain the exact same DNA. These different patterns of gene expression cause your various cell types to have different sets of proteins, making each cell type uniquely specialized to do its job.

For example, one of the jobs of the liver is to remove toxic substances like alcohol from the bloodstream. To do this, liver cells express genes encoding subunits (pieces) of an enzyme called alcohol dehydrogenase. This enzyme breaks alcohol down into a non-toxic molecule. The neurons in a person's brain don’t remove toxins from the body, so they keep these genes unexpressed, or “turned off.” Similarly, the cells of the liver don’t send signals using neurotransmitters, so they keep neurotransmitter genes turned off.

B) Housekeeping genes

Transcription start sites of housekeeping genes can span over a region of around 100 bp where as transcription start sites of developmentally regulated genes are usually focused in a narrow region. Little is known about how the dispersed transcription initiation of housekeeping gene is established.

Most housekeeping genes utilize a promoter lacking the common TATA and CAAT boxes, and having instead a series of GC boxes (consensus sequence GGGCGG). GC boxes provide binding sites for the transcription factor Sp1 and, like the TATA box, direct the start of transcription. Since there are several GC boxes in the promoters of many housekeeping genes, the transcription start site is ambiguous. Indeed, many housekeeping gene transcripts have heterogeneous 5′ start sites. The coding function of these genes is not impaired, however, because all of the alternative start sites are within the 5′ untranslated region of the mRNA. As an example, the human c-Ha-ras oncogene promoter has about 80% G+C content, 10 GC boxes, and at least four transcription start sites.