L13 - From Genotype to Phenotype Flashcards
How do we get from genotype to phenotype?
Genes must be expressed to have any effect on phenotype. The central dogma of molecular biology.. flow of information from DNA to RNA to protein. A small number of exceptions exist e.g. RNA genome of some viruses.
What happens in transcription pt1?
DNA is used as a template to make complimentary RNA transcript. Only the template strand is transcribed. Its partner, the nontemplate strand is not transcribed.
What happens in transcription pt2?
Ribonucleotides are added to the 3’ end of the RNA transcript via the polymerisation reaction. Bases complimentary to those in the DNA are added. Remember in RNA, U (not T) pairs with A.
What happens in transcription pt3?
The enzyme RNA polymerase carries out the polymerisation. Within the polymerase, the two DNA strands separate to form a transcription bubble.
What happens in transcription pt4?
Transcription doesn’t happen at random. A DNA molecule usually contains many genes that are transcribed individually at different times, often from opposite strands. Transcription starts with a promoter (region of a few hundred bases where the RNA polymerase and associated proteins divide), which usually contains a TATA (TATAAA-3’) box, stops at terminator region.
What happens in transcription pt5?
In bacteria, promoter recognition is facilitated by sigma factor protein, which facilitates RNA polymerase binding to specific promoters. In eukaryotes, promoter recognition is considerably more complex (at least general transcription factor proteins) - provides additional regulation of gene expression. General transcription factors bind to the promoter, and transcriptional activator proteins bind to enhancers.
What happens in transcription pt6?
General transcription factors recruit RNA polymerase 2 (Pol 2) to the site for transcription. In addition, proteins bound to an enhancer sequence need to recruit a mediator complex, which in turn interacts with the Pol 2 complex. Through looping of DNA, transcriptional activator proteins, mediator complex, RNA Pol 2, and general transcription factors are brought into close proximity, allowing transcription to proceed
What happens in transcription pt7?
Transcription bubble is ~14 base pairs in length, the RNA-DNA duplex is ~8 base pairs in length. Transcription takes place in what looks like a bubble that is about 14 base pairs in length
Overall what happens in transcription?
Goes from genes to proteins.
What is the RNA transcript that comes off the DNA template?
The primary transcript. In prokaryotes, the primary transcript is the messenger RNA (mRNA). Carries the genetic message to the ribosome, translation occurs right now. Some prokaryotic mRNAs code for multiple proteins, usually successive steps in the same biochemical pathways.
Overview of RNA processing?
In eukaryotes, translation can’t happen right away because the primary transcript is in the nucleus and ribosomes are in the cytoplasm. Primary transcripts go through several types of chemical modification, known as RNA processing.
What happens in step 1 of RNA processing?
5’ caps, made of a modified nucleotide, enable the ribosome to recognise mRNA.
What happens in step 2 of RNA processing?
The poly(A) tail (about 250 consecutive As to the 3’ end of the mRNA) is added to the 3’ end. This plays a important role in transciption termination and the export of the mRNA to the cytoplasm. The 5’ cap and the poly(A) tail help to stabilise the RNA transcript.
What happens in step 3 of RNA processing?
RNA splicing. Exons - protein coding, Introns - not protein coding. During RNA splicing, introns are removed. This is catalysed by a complex of RNA and proteins called a spliceosome
What happens in step 3 of RNA processing - why bother with introns?
In alternative splicing, primary transcripts from the same gene can be spliced in different ways to yield different mRNAs and ultimately different protein products.
What are the non-protein coding RNAs?
- Ribosomal RNA
- Transfer RNA
- Small nuclear RNA
What is Ribosomal RNA?
Makes up the bulk of ribosomes (used for translation)
What is Transfer RNA?
Carries individual amino acids for use in translation
What is Small Nuclear RNA?
Essential component of the spliceosome
What are the most abundant transcripts in mammal cells?
rRNA (80%), and tRNA (10%), they are needed in large amounts to synthesis proteins.
Translation - How do we get proteins from mRNA?
Proteins are made from amino acids - there are 20 commonly found in proteins. The mRNA bases code for different amino acids. A codon is each group of three adjacent nucleotides coding for a single amino acid. mRNA therefore has three different reading frames - need to start translating in the right place.
Translation - codons?
There are 64 codons (each nucleotide can occupy 3 different positions of the codon -> 4cubed. 61 codons represent amino acids, which encode 20 amino acids. Almost all amino acids are represented by more than one codon. Except for methionine (AUG) and tryptophan (UGG). Often the third base of the codon is not important - third base degeneracy. 3 codons terminate translation.
Where does translation occur?
Occurs within ribosomes
What happens in stage 1 of translation?
Initiation - Initiation factors recruit the small ribosomal subunit and tRNA and scan the mRNA for an AUG codon. When the complex reaches an AUG, the large ribosomal subunit joins, the initiation factors are released, and a tRNA complementary to the next codon binds to the A site. A reaction transfers the met to the amino acid on the tRNA in the A site, forming a peptide bond.
What happens in stage 2 of translation?
Elongation - The ribosome moves down one codon, which puts the tRNA carrying the polypeptide into the P site and the now uncharged tRNA into the E site, where it is ejected. A new tRNA complementary to the next codon binds to the A site. The polypeptide transfers to the amino acid on the tRNA in the A site. The polypeptide is elongated
What happens in stage 3 of translation?
The process continues until one of the stop is encountered (UAA, UAG, UGA). When the stop codon is encountered, a protein release factor binds to the A site of the ribosome, causing the bond connected to the polypeptide of the tRNA to break. The breaking of the bond creates the carboxyl terminus of the polypeptide and completes the chain.
What do proteins provide the basis for?
Many chemical pathways e.g. MC1R gene -> melanocortin 1 receptor protein. Different MC1R alleles lead to differences in hair and skin colour. Receptor plays a important role in normal pigmentation.
What is the point in controlling gene expression?
We don’t want to express all genes at all times. E.g. we don’t want to produce melanin in our pancreas and insulin in our skin. So genes can be turned on and off. Regulation can occur at any stage
What are the methods of controlling gene expression?
- Epigenetics
- Chromosome-level gene regulation
- Transcriptional regulation
- Post-translational regulation
What are epigenetics?
A - Chromatin remodelling exposes different stretches of DNA to the nuclear environment. Can occur via histone modification - these can prevent or promote transcription. B - Chemical modification of some DNA base e.g. DNA methylation. Adds a CH3 group to the nucleotide (usually cytosine) that results in restricted access of transcription factors to promoters (prevents transcription)
What is chromosome-level gene regulation?
E.g. one copy of the X chromosome is inactivated in human females shortly after the embryo implants. The copy that is inactivated is random, leading to different chromosomes being inactivated in different clusters of cells
What is transcriptional regulation?
Regulatory transcription factors can promote transcription by binding with enhancer sequences or recruiting general transcription factors. Other regulatory transcription factors bind with DNA sequences known as silencers and repress transcription.
What is post-transcriptional regulation?
Regulation at this level is essential because some proteins are downright dangerous - e.g. proteases such as the digestive enzyme trypsin must be kept inactive until secreted out of the cell. Post-translational modification also helps regulate protein activity. E.g. by the addition of one or more sugar molecules or a phosphate group to the side chains of some amino acids. Marking proteins for enzymatic destruction by the addition of chemical groups after translation is also important in controlling their activity.
What do several covid mRNA vaccines make use of?
The body’s natural transcription process.
