Gene Regulation in Eukaryotes

In both eukaryotes and prokaryotes, transcriptional regulation occurs through the attachment of _____ ______ ______ to specific DNA sequences that are in the vicinity of the transcription unit itself. However, additional levels of complexity are both possible and necessary for controlling expression in eukaryotes for several reasons. Name 4 such reasons

As in proks. gene expression in euks can be regulated at the time of transcription _________, when RNA polymerase starts to make a primary transcript. Important decisions concerning the amount of ____ ______ in the cell are indeed often made at this point

Name 5 activities in the process of gene expression that can be regulated and lead to an active gene product (excluding transcription initiation).

Three types of RNA polymerases transcribing genes in euks and what they regulate

Although each of the regulatory regions of the thousands of pol II transcribed genes in a euk genome are unique, genes that are transcribed in a cell-type specific manner all contain two kinds of essential DNA sequences namely:

The promoter, which is always very close to the gene's ______ ______ region, usually contains a ______ _____ (or _____ _____), consisting of roughly seven nucleotides of the sequence T-A-T-A-(A or T)-A-(A or T), located just _______ of the transcription initiation site. Binding of RNA pol to the _____ _____ allows a low, so called ____ level of transcription

The second type of DNA sequence element important for transcription in eukaryotes is called an _______: a regulatory site that can be quite distant (10,000s of bases) from the promoter. In specific cell types, binding of proteins to enhancers can ______ or _______ basal levels of transcription

Enhancers may be located either ___ or ____ to the transcription start site (some are even found in ______) and a single gene can have ____ or _____ enhancers. A single enhancer may have multiple ______ _____ for different transcription factors

Scientists often use a gene from jellyfish that encodes green flourescent protein (GFP) as a reporter to identify enhancers in euks. How does it work? (5-story)

Researchers can generate mutations in the control region of the GFP reporter construct and then breed transgenic animals with the ______ DNA. If GFP fluorescence is no longer seen in the same tissue, then the mutation must have affected the ______-specific enhancer. In this way, scientists can define the actual DNA sequences comprising particular ______

The binding of proteins to a gene's promoter and enhancer (or enhancers) controls the rate of ________ ________. Different types of proteins bind to each of the cis-acting regulatory regions: _____ factors bind to the promoter, while ______ and _______ bind to the enhancers

Define transcription factors

Once the transcription factors bind to the DNA, they recruit additional ______ to the gene that can also influence _________

Basal factors assist the binding of RNA polymerase II to the ______. The key component of the basal factor complex that forms on most ______ is the ____ _____ _____ protein. This protein recruits other proteins called _____-_______ factors to the promoter in an ordered pathway of assembly

Once the basal factor complex has formed, RNA polymerase can initiate a _____ level of transcription. The primary sequences and three-dimensional structures of the basal factors are highly ______ in all euks, from yeasts to humans

Transcription of many euk genes requires a multi subunit complex called _____ that contains more than 20 proteins. _______ does not bind DNA directly but instead serves as a bridge between the ___ ____ complex at the _______, and ______ or _______ proteins bound at the enhancer

The enormous range of transcriptional regulation occurs through the binding of different ______ _____ to distinct _______ elements associated with different ______. When bound to an _____ element, transcriptional activators can interact _______ or ________ with basal factors at the promoter in a three-dimensional protein/DNA complex to cause a/an _______ in transcriptional activity

Why must DNA between the enhancer and the promoter loop for interactions to occur between basal factors, mediator and activators?
How can enhancer sequences still function if moved to different positions relative to the promoter

At the mechanistic level, transcriptional activator proteins bind their target sites on DNA and increase RNA synthesis by doing one or both of the following:

Transcriptional activator proteins must bind to enhancer DNA in a _______-specific way, and after binding, they must be able to interact with other ______ (a _____ factor or a _______) to activate transcription. Name the two structural domains within the activator protein that mediate these two biochemical functions

Transcription factors belong to several protein families that share similar DNA-binding domains. Two well characterized DNA-binding structures are _____ _____ _____ domains and _____ _______. Where are each found?

DNA-binding domains fit within the _____ groove of DNA. Subtle differences in the _____ ____ sequences among activators of the same family can allow these proteins to recognize specific DNA sequence at different ______ ______

The activation domains of transcription factors are less well __________ than the DNA-binding domains, and may be less ______. The amino acid sequences of activation domains depends on whether the activator interacts with the ______ complex or with one or more _________

Many activator polypeptides also have a third domain (what is it called/what does it do?)

One structural motif in many dimerization domains is a ______ _____, a helix with leucines at regular intervals. The ability of two such _____ _____ proteins to interlock depends on the specific amino acids that lie between the ______. Most such proteins also have a characteristic DNA-binding domain, called a ______ domain

Among the best-characterized transcripton factors with a leucine zipper is Jun, a protein important for cellular proliferation and other processes such as the ____ and _______ of the uterine endometrium during the mammalian menstrual cycle. Jun can form dimers either with itself, making Jun-Jun ________, or with other proteins called Fos, making Jun-Fos ________

Repressors

In prokaryotes, negative regulators generally work by ______ ______ RNA polymerase from binding the promoter. In euks, the primary function of repressors in different (explain)

What are the two possible functions of corepressors?

How are repressor structures similar to activators?

The drosophila transcription factor called Dorsal is an activator when bound to the enhancers of some target genes and a repressor when bound to the enhancers of other genes. How can one protein have two opposing functions in transcription?

Many regulatory proteins, called ______ _______, prevent transcription initiation indirectly, not by recruiting corepressors, but instead by interfering with the function of activators. Name the four ways this is accomplished

Scientists often use GFP reporter genes to verify that a transcription factor suspected of regulating the expression of a target gene indeed plays such a role. What are the two requirements of this experimental approach?

By contrast, loss of function mutations in a gene encoding a repressor may yield _____ levels of GFP flourescence than normal, and/or the expression of GFP in _________ cells

In complex multicellular organisms, a large percentage of genes are devoted to transcriptional regulation. Of the 25,000 genes in the human genome, scientists estimate that more than _____ of them encode transcription factors. Many proteins can regulate the ______ of any one gene, and each regulatory protein may act on ______ different genes

Operational definition of an enhancer

Why is it that a gene's regulatory region may contain several enhancer elements, such as an eye enhancer that activates transcription in eye cells and a skin enhancer that activates transcription in skin cells

Each enhancer in turn has one ore more binding sites with varying _______ for each of several different activators and repressors. At any moment, there may be dozens of _____ and ________ in the cell that compete for these binding sites, as well as _______ and _______ that compete with each other for binding to different ________ or ________. All this info biochemically integrated from an enhancer guides the cell to do what two things?

Within a multicellular organism, all cells have the same ______ _____, so a gene's enhancer elements are present in ____ types of cells. The cell-type specificity of transcription therefore depends on changes that occur during which process?

Euk transcription factor activity can be regulated through allosteric interactions with small molecules. Steroid hormone receptor transcription factors constitute an important exmaple of allostery in euks. (explain the example)

Transcription factor proteins can be modified after they are synthesized by the ______ addition of any of several different chemical groups. One of the most important of these modifications is phosphorylation (define)

Phosphorylations can either ______ or ______ a transcription factor in any of a number of ways. List 4

Cells often rely on phosphorylations to control events that must occur _______ (example)

Not all transcription factors are made in ____ cells at ____ times. Clearly, if a factor is not present in a given cell, it will be ______ to influence the initiation of the ________ of any target genes. (explain)

An enhancer may be located ______ or ______ of the promoter that it regulates and in either orientation with respect to the promoter. These facts pose a conceptual problem. Pg 555 top right

Insulators (define)
How do we test if something is an insulator in a reporter gene?

How insulators work (2-story)

Cis acting elements
Trans acting elements

chromatin immunoprecipitation sequencing