Transcription and RNA
Explain some mutations.
a mutation in a consensus sequence of a promoter will prevent the binding of transcription factors and or of RNA polymerase. A mutation in a enhancer could prevent binding of an activator.
Types of RNA?
rRNA: about 120-4718 nucleotides long. Functions include translation (ribosome structure and catalytic activity). tRNA: 54-100 nucleotides long. functions include delivery of amino acids to ribosome during translation. only need to know top two. recognize the rest and know some of there possible roles. Small interfering RNA siRNA: 20-25 nucleotides long. functions include sequence-specific inactivation of mRNA. Micro RNA (miRNA): 20-25 nucleotides long and functions to sequence specific inactivation of mRNA. Large intervening noncoding RNA (lincRNA): up to 17,200 nucleotides long and functions: transcriptional control. Small nuclear RNA (snRNA) 60-300 nucleotides long and functions: RNA splicing. Small nucleolar RNA (snoRNA): 70-100 nucleotides long and functions: sequence specific methylation of rRNA.
What percent of the human genome undergoes transcription to produce noncoding RNAs?
roughly 80%
What do the + and - #s mean on a promoter?
the - means that it is upstream and + indicates that it is downstream from the start of transcription. The number signifies the number of nucleotides away from the start site.
Consensus sequence:
A consensus sequence is a common nucleotide sequence across all promoters. One of them is called the TATAAT box (-10 in pro and -30 in euk). Only have to remember the TATAAT box.
Gene: Most RNA transcripts correspond to a single functional unit.
A gene is a segment of DNA that is transcribed. It will not always code for a protein. Only about 1.5 or 2% of genes actually code for proteins. For protein encoding genes, the mRNA sequence will include all the information specifying the amino acid sequence of a polypeptide.
Operon: in Eukaryotes, every single gene will have its own promoter. In prokaryotes, functionally related genes are grouped together and share a single promoter.
A group of genes that fall under control of one promoter. Functionally related genes. Only found in prokaryotic cells. in the lac operon, transcription of several genes is regulated by one repressor.
Enhancer:
A section of DNA that will have consensus sequences that allow binding of proteins to enhance transcription. This allows an activator to bind to it.
Spliceosome:
A spliceosome is a large and complex molecular machine found primarily within the splicing speckles of the cell nucleus of eukaryotic cells. The spliceosome is assembled from snRNAs and protein complexes. The spliceosome removes introns from a transcribed pre-mRNA, a kind of primary transcri
How does RNA polymerase transcribe a gene through a nucleosome?
Acetylation happens and loosens up the bond between the histones and the DNA so it can transcribe the gene. Methylation of the histones will increase the association between the histone and the DNA and this will make it so that the DNA gene cannot be transcribed (it is silenced).
Alternative splicing:
Alternative splicing is one of the explanations as to why we have over 100,000 proteins created from only about 20,000 genes. Essentially this is a controlled way that removes particular exons. It will remove some exons that allow for different combinations of exons to be expressed.
Activator: conformational changes in one of these proteins will change the conformation in other associated proteins.
An activator is the protein that binds to the DNA sequence (enhancer). Acts similar to transcription factor. Will bind to a mediator and this will allow it to fold over the DNA and will help with the construction of RNA polymerase. The main point of the activator, mediator, and transcription factors is that they create a good environment where the RNA polymerase can bind to the promoter.
Describe two ways that transcription in prokaryotes is terminated
Destabilizing of RNA polymerase DNA complex by: Hairpin loop: This is when the RNA sequence loops back on itself and actually becomes double stranded. This requires the sequence of mRNA to be complementary to itself. The end of the gene has to code for RNA that is complementary to itself to form the hairpin loop. Rho: When Rho binds to the consensus sequence of the transcript, it causes conformational changes that cause transcription to stop.
Outline transcription initiation
Histone modification must be required first so that the DNA can be access. This is called chromatin remodeling. After this, transcription begins when RNA polymerase binds to the promoter with the help of facilitation factors (these allow RNA polymerase to bind to promoter). The transcription factors will also unwind the DNA to form the transcription bubble.
What mechanism is employed in the sequences to allow some sequences to be alternatively spliced out?
Possibly there might be some methylation or other modification to the DNA that will allow the 3' splice site to be skipped over. Some proteins could bind to the 3' site and allow them to be skipped. RNA molecules could overlap in a complementary way and cover over the 3' site and make them inaccessible (by hiding it).
List which RNA polymerases transcribe the major types of RNA
RNA polymerase I: transcribes rRNAs RNA polymerase II: transcribes mRNAs RNA polymerase III: transcribes tRNAs
Difference between RNA polymerase and DNA polymerase?
RNA polymerase does not require a primer to start and DNA polymerase does. Also it is important to notice that transcription and replication occur in different parts of the cell. Stains of RNA and DNA polymerase reveal that they occur in different parts.
Transcription factor: Eukaryotic transcription factors interact with DNA, with RNA polymerase, and with each other.
RNA polymerase requires transcription factors to bind to promoter areas. Transcription factors are proteins that also recognize the promoter region and allow transcription to happen. They facilitate the binding of RNA polymerase to the consensus sequence of the promoter region. Transcription factors also unwind DNA to form a transcription bubble. Transcription factors are proteins that facilitate binding of RNA polymerase by altering the structure/form of DNA at the promoter. They bind to the consensus sequences and also to the RNA polymerase
What strand does RNA polymerase transcribe?
RNA polymerase transcribes the template strand, not the coding strand (the coding strand contains the information is useful to us and that we can decipher. Read from 5' to 3' end).
mRNA turnover
Some times the end of the poly A tail or 5' cap is removed so that the mRNA is degraded and removed. This could happen if the protein the mRNA codes for does not need to be made in the body at that time.
Discuss the steps and purpose of RNA processing Endonucleases are not as specific so they do not attack the middle often. As soon as you have available splice sites in the transcript, the spliceosome will begin splicing even if transcription is not finished.
The addition of a 5' cap: A 7-methyl G nucleotide is added to the 5' end to protect the mRNA from degradation by the exonucleases. The exonucleases would chew on the ends of the mRNA. The 5' cap is added to the end so that the mRNA does not get damaged and missing part of the gene. Added as soon as 5' end is made. Splicing: Splicing is the removal of introns and joining of exons in an RNA molecule. It is controlled by the consensus sequences. A big complex of proteins called the spliceosome will recognize the consensus sequence and it will bind to the 5' splice site right before the intron and the 3' splice site at the end of the intron to cut it out with endonuclease activity. Must have a ligase ability so it can combine the two exons together after it cuts out the introns. Addition of a 3' poly(A) tail: a sequence of As (AAAAA...) that is added to the 3' end. A consensus sequence on the nucleotide strand will trigger the enzyme poly(A)polymerase to create the poly(A) tail. The tail is then bound by poly(A)-BP. The mRNA has to be transported out of the nucleus to protect the transcript.
What would happen if you have a mutation in the splice site?
The spliceosome reads along the mRNA and looks for the splice sites to start and stop cutting. If there is a mutation in a 5' splice site, the spliceosome will not start the cut and it will include the entire intron in the transcript. This will include a lot of new nucleotides in the transcript and will probably change the reading frame. A mutation in the 3' splice site will mean that the spliceosome will not know where to cut again at the end of the intron. This will cause it to splice again at the next 3' splice site and this will cause it to cut out the subsequent exon!
Intron:
These are noncoding sections of the gene that will be cut from the transcript. Introns usually span large areas of the gene.
Exon:
These are the sections of a gene that will be sent to the ribosome where proteins will be made
Promoter:
Transcription is initiated at the promoter (a section of the DNA where RNA polymerase can bind) which is upstream (towards the 5' end) of the actual start codon. It starts before the start codon because you want a little bit of extra RNA sequence in front of the start codon so the transcript will not be degraded. It acts as sort of a buffer zone. A promoter is not a molecule that binds to the gene, it is a section of the gene itself where RNA polymerase binds and transcription will begin.
Transcription:
Transcription is the process of using a gene as a template to create an mRNA strand that will code for a gene. mRNA includes all of the information specifying the amino acid sequence of a polypeptide. These RNA transcripts correspond to a single functional unit.
Explain why transcription happens:
Transcription occurs so that the genetic information in DNA can be copied into mRNA sequences.
Why does transcription start before the start codon and why does it end after the stop codon?
Transcription starts before the start codon and after the stop codon so that it will have a little extra bit of transcribed RNA to act as a buffer region on either end. This will protect the entirety of the transcript from degradation.
Explain the purpose of chromatin remodeling Transcription in eukaryotes may requre alteration of histones and chromatin structure.
chromatin remodeling is moving around the protein complexes so that we can access the DNA and perform transcription. Change the covalent bonds on a histone between DNA so that it can change the availability of the DNA. Acetylation will repel the DNA from the histone and this will for it to be accessed. Methylation will tighten the association and will not let the DNA gene to be expressed.
Outline the process of transcription During transcription, the DNA strands separate, and RNA polymerase constructs an RNA molecule that forms a short hybrid helix with the template strand. RNA polymerase is a processive enzyme that proofreads its work. The shift from initiation to elongation involves structural changes in RNA polymerase (it does not bring along the transcription factors with it), including phosphorylation of its C-terminal domain. The mechanisms for transcriptional termination differ in prokaryotes and eukaryotes.
mRNA strand is being created 5' to 3'. Same way as DNA replication. Nucleotides are added on to the 3 prime end. So activators bind to the enhancer region of the DNA template strand that is upstream of the promoter region. This helps construct the set up for the addition of the RNA polymerase II molecule. Transcription factors also recognize the promoter region of the DNA template strand and they bind to the consensus sequence and facilitate the binding of RNA polymerase II to the consensus sequence in the promoter region. The transcription factors also open up the DNA strand and unwind it and create the transcription bubble. At this point the RNA polymerase II molecule starts to create a nucleotide sequence by reading off the template strand of DNA. It reads off the promoter sequence (5' tail) and then over the start codon. The sequence is read off until it reads the stop codon and then it creates a 3' cap by transcribing a little bit more after the stop codon.