Regulation of gene expression in eukaryotes
Gene expression in a multi-cellular organism is very different than in a prokaryote. One such difference is cellular differentiation.
-Cellular differentiation: start out with general undefined cells then turn on different genes to make a different type of cell or cell types. This process is CRITICAL. You need to turn on the correct genes at the correct times otherwise you will get death.
- ex. of cellular differentiation --> you need different things in a muscle cell then in a nerve cell.
(3:35) How the regulation of gene expression differs in Eukaryotes
- Eukaryotic cells are larger and more complex than prokaryotes. Within this, DNA is packed into chromatin with histone proteins. Chromatin remodeling is a key step in the regulation of gene expression in a eukaryote. So if you don't have your DNA in a form that is accessible to RNA POL to copy you can essentially turn off transcription by shuttin down chromatin remodiling.
- Eukaryotes typically have their DNA in multiple chromosomes not one chromosome (typically seen in prokaryotic organism)
- Because DNA is in the nucleus and the ribosomes are at the endoplasmic reticulum (ER) in the cytosol, transcription and translation are seperated spatially and temporally (occur at different times). In a prokaryote: almost as quickly transcription starts to produce RNA, ribosomes come in, bind that RNA and start to translate that RNA. In eukaryotes these processes are seperated.
- mRNA molecules are modified prior to exiting the nucleus in a eukaryote. That modification includes splicing.
- (8:50) Eukaryotic mRNA molecules are more stable than prokaryotic, they have a longer half-life (amount of time they exist). Partly because Prokaryotes need to rapidly respone to changing conditions. Eukarytoes don't experience this as much.
- In eukaryotes regulation can also occur at the level of translation. You have a more stable mRNA molecule but you may alter or regulate the translation of an individual mRNA molecule because it's not being produced and degrated as quickly.
- While all eukaryotic cells contain complete copies of their genome (all chromosomes + DNA in all cells) different cells express different subsets of genes.
- Broadly: the process of regulation in a eukaryote is a more complex process then what is seen in a prokaryote.
| Differences in regulation of gene expression | Prokayote | Eukaryote |
| 1st difference | larger and more complex | |
| 2nd difference | DNA typically in one chromosome | DNA in multiple chromosomes |
| 3rd difference | transcription and translation happen almost simultaneously | transcription and translation are seperated spatially and temporally |
| 4th difference | mRNA molecules are modified prior to exiting the nucleus | |
| 5th difference | Prokaryotes need to rapidly respond to changing conditions - less stable. | more stable, longer half-life |
| 6th difference | regulation can also occur at the level of translation | |
| 7th difference | contain complete copies of their genome in each cell |
(14:45) Chromosome organization in the nucleus influences gene expression
- During interphase, the DNA found in the nucleus is in a relaxed state. However, there is stil orginization to how DNA is organized within the nucleus. This organization plays a key role in the regulation of gene expression.
- Within the nucleus, each unique chromosome exists in a chromosome territory.
- The regions between the chromosome territory's are called interchromosomal domains.
- (19:45) Within the nucleus the arrangement is as follows:
- Chromosomes with small numbers's of genes have thier chromosome territory on the outside of the nuclues.
- Chromosomes with larger numbers of genes exist in chromsome territiores towared the inside of the nucleus.
- It has been proposed that the genes being actively transcribed on a chromosome will be found toward the edge of the chromosome territory. This suggests that RNA POL lives in the interchromsomal domains. The genes need to be brought close to those areas so they can be transcribed.
- Once we get a chromosome in position the intition of transcription begins --> two steps.
- Chromatin Remodeling
- We need to recruit a number of factors (typically proteins) that help initiate transcription.
(27:10) Transcription Initiation
- There are three common cis acting elements in Eukaryotic transcription initiation: promoters, enhancers and silencers.
- The process requires chromatin remodeling, a number of DNA sequences and over 100 proteins. Just to initiate transcription.
- Promoters: The site where the transcription machinary binds to start transcription.
- The promoter typically facilitates a basal level of transcription
- Typically adjacent to the gene (upstream)
- contains a few 100 nucleotides.
- (31:30) Within those nucleotides there are number of key sequences of the promoter
- TATA Box a.k.a. Core promoter - 25-30 BP region of DNA that is bound by RNA POL. This contians a 7-8 BP consensus sequence. The consensus sequence contains a number of nucleotide sequences and the TATA sequence. This is the RNA POL "docking site".
- mutations in this sequence decrease the level of transcription. Deletion of the sequence results in loss of transcription.
- CAAT Box - these element contain the sequence CAAT or CCAAT located 70 BP upstream of the start of the gene.
- Mutations in this sequence=decreased transcription.
- (36:15) GC Box - GGGCGG and is found 110 BP upstream of the transcription start site.
- both the GC box and the CAAT box can serve as enhancers as well as part of the promoter.
- (40:00) Enhancers: Can be found on either side of a gene and can some times be great distances away.
- enhancers are typically bound by multiple proteins with a net effect of stimulation of transcription.
- (42:50) How do we differentiate an enhancer from a promoter?
- Promoter regions are found at fixed locations. Enhancers are not found in fixed locations (they can move around.
- You can invert an enhancer without affecting its activity. (not the case for a promoter)
- If you move an enhancer to a different gene, that gene will now be regulated by the enhancer.
- Promoters are responsible for the basal level of gene expression. Enhancers are necessary for the full expression of a gene.
- Enhancers can be cell type specific and promoters are not.
(47:45) How do enhancers stimulate the level of transcription?
- Factors can bind enhancers which help with chromatin remodeling.
- When a factor binds the enhancer it bends the DNA bringing the enhancer and promoter closer together. This can help stimulate RNA POL binding to the promoter
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