Lecture 5: transcription regulation Flashcards
True/false: there are more protein coding genes (functional genes) than that there are non-coding genes.
False, there are approximately 20.000 functional genes and 25.000 non-coding genes
What is an important first checkpoint when a gene (DNA) is translated to a protein?
Transcriptional control
80% of total RNA synthesis is based on….
RNA polymerase I and III
What makes RNAp II different from the other RNAp I and III?
Each RNAp requires distinct DNA sequences/promotor elements. RNAp II only transcribes protein-coding genes and gets directed by specific transcription factors meant for RNAp II.
What function does RNAp I have?
RNAp I synthesizes ribosomal RNA and is located in nucleoli, where ribosome assembly factories are also located.
Describe how a cluster of rRNA genes looks like (what sequences can be found in these operons)?
rRNA genes are tandemly arranged with intergenic spacer sequences between rRNA genes. The rRNA gene itself contains subunits of the small and large ribosomal subunits. The intergenic spacer contain the following:
- terminator sequence
- cis acting elements functioning as enhancers
- Up stream promotor elements (UPE)
- Core promotor sequence
There are approximately 200 rDNA genes (in haploid cells and 400 copies rDNA in diploid genome). Does every cell need these 400 rDNA genes?
No, it depends on how active the cell is in ribosome and protein synthesis. But in general about 1/2 is transcriptionally active (and unmethylated) 1/2 is transcriptionally active (low methylated) and can occasionally lead to transcription and a small part of these rDNA genes are hypermethylated and cannot be transcribed.
What function does RNAp III have?
RNAp III synthesizes a few small, stable and non-translated RNAs (like tRNA, 5s RNA, 7sl RNA, u6 snRNA etc.)
What is interesting about the genes that get transcribed by RNAp III?
RNAp III recognizes its target genes through intragenic promotors; specific sequences that are located within the genes that gets transcribed (so no recognition of upstream promotor elements or core promotor sequences.
Some RNAp III still recognize their target genes through upstream promotors and not via intragenic elements. What’s characteristic about these target genes?
The sequence around this upstream promotor is uracil (U) rich.
Briefly explain the functions of the following sequences in genes for RNAp II:
- core promoter
- distal promoter
- enhancer
- boundary element or matrix attachment region
- Core promoter contains the transcription start site
- Distal promoter binds transcription factors and help with the assembly of the pre-initiating complex
- Enhancer DNA region that stimulates gene expression by helping with the recruitment of RNAp.
- Boundary element or matrix attachment region, DNA elements that bind specifically to the nuclear matrix.
What is the TATA box and what function does it have?
TATA box is a sequence that is located upstreams of the transcription start site. It binds to transcription factor TFIID, that recruits other transcription factors (e.g. TFIIB) that result in the recruitment of RNAp II.
What if the TATA box is not present in a core promoter?
Then there are other elements (like BRE, INR or DPE) that can recruit transcription factors which results in recruitment of RNAp II.
Just know that only 15% of all RNAp II promoters in mammals contain a TATA box. So no TATA box means that one or more of the other elements have to be here. Beside BRE, INR or DPE, what do the majority of promoters appear to consist of?
CpG islands
What are CpG islands?
These are CpG rich DNA sequences with a lenght of 200 bp to several kb
Fill in: unmethylated or methylated.
CpGs within CpG islands are normally … while most CpGs outside CpG islands are …
CpGs within CpG islands are normally unmethylated while most CpGs outside CpG islands are methylated.
What is characteristic for CpG islands (name two)?
- CpG islands nearly always encompass promoters and/or exons
- CpG islands typically lack TATA or DPE elements, but contain myltiple GC box motifs bound by the transcription factor Sp1.
What are DNA cis-acting elements?
Cis acting elements are a region of DNA that regulates the expression of genes located on the same molecule.
What is the function of histone acetyl transferases (HAT)?
These enzymes are involved in acetylation of lysine residues on histone tails. This makes the chromatin open for transcription (euchromatin).
What is the difference in spatial assembly between transcription of genes in a small genome and transcription of genes in a complex genome?
In a small genome there’s a transcriptional unit, where all the things that are needed for gene transcription lay in close proximity of each other. In a complex genome, these elements can be dispersed through the DNA and don’t necessarily need to be in close proximity with each other.
So in complex genomes, if transcriptional elements don’t lay in close proximity with each other: how can transcription still occur?
The DNA will form DNA loops to bring certain elements together.
What’s meant by coordinated expression of genes?
That in order for one gene to be on and actively transcribed, another gene that lays in close proximity needs to be inactive.
What are ‘transcription factories’?
Some genes have an overlapping function or use the same transcription factor to be transcribed. Then it’s useful to have these genes in a cluster so that they can be transcribed more easily.
How is chromosome conformation capture performed?
- Treating cells with formaldehyde, this allows proteins to be covalently bound together and to the DNA.
- Isolation of nuclei and/or chromatin
- Digestion of nuclei and/or chromatin by restriction enzymes.
- Ligation of the restriction ends (5’ ligated to 3’) with ligase.
- Cross-link and get rid of the bound proteins, so you end up with two pieces of DNA attached to each other.
- Sequence the DNA pieces and then match the sequence to the DNA strand, so you know the location of the proteins.
What are Topologically Associated Domains (TADs)?
TADs are linear chromosomes divided into specific domains. Here, each red dot displays a region that is covalently linked to another region. So also in linear chromosomes there are certain domains that interact with each other (A and B), the so called TADs.
What’s important to remember based on this picture?
That by changing the DNA sequence, that the order of interacting groups that are able to communicate with specific amino acids also changes. So you get a different binding site.
What kind of DNA-interacting domain is this?
Helix-turn-helix
What are leucine zippers?
DNA-interacting domains that regulate the DNA through homo- or heterodimers that bind to two major grooves.
Why is it important that leucine zippers can either be homo- or heterodimers?
It enhances the specificy, see this picture. The use of a heterodimer is called combinatorial control.
What are zinc-finger proteins?
You need a Zinc atom that is able to bind to histidine and cysteine residues. These are connected to helices, which bind to the bases.
