central dogma: transcription- lecture 2 Flashcards
c value paradox
the “constant” value of DNA content in a cell
the total haploid content of a genome doesnt correlate strongly with level of complexity (onion has more dna but isnt smarter than human, despite having 5* as much dna)
what is a gene
a gene is the dna sequence required for the production of an rna that either has its own particular biological activity or that can in turn be translated into a functional protein
main function of dna
to encode the instructions needed to produced rna, which in turn are read by the cell to produce proteins
how many genes in human
20000
what does natural selection do with genes
eliminates mutations that disrupt function (mutations that land in regions of genomes that are functionally important)
The main enzyme involved in transcription
RNA polymerase
RNA polymerase builds an RNA strand in what direction
5’ to 3’ direction, adding each new nucleotide to the 3’ end of the strand
stages of transcription: Initiation
RNA polymerase binds to a sequence of DNA called the promoter, found near the beginning of a gene. Each gene (or group of co-transcribed genes, in bacteria) has its own promoter. Once bound, RNA polymerase separates the DNA strands, providing the single-stranded template needed for transcription
stages of transcription: Elongation
One strand of DNA, the template strand, acts as a template for RNA polymerase. As it “reads” this template one base at a time, the polymerase builds an RNA molecule out of complementary nucleotides, making a chain that grows from 5’ to 3’. The RNA transcript carries the same information as the non-template (coding) strand of DNA, but it contains the base uracil (U) instead of thymine (T)
stages of transcription: Termination
Sequences called terminators signal that the RNA transcript is complete. Once they are transcribed, they cause the transcript to be released from the RNA polymerase. An example of a termination mechanism involving formation of a hairpin in the RNA is shown below
In bacteria, RNA transcripts can act as messenger RNAs (mRNAs) right away. In eukaryotes, the transcript of a protein-coding gene is called a
pre-mRNA and must go through extra processing before it can direct translation
how are pre-mrna’s modified
Eukaryotic pre-mRNAs must have their ends modified, by addition of a 5’ cap (at the beginning) and 3’ poly-A tail (at the end).
Many eukaryotic pre-mRNAs undergo splicing. In this process, parts of the pre-mRNA (called introns) are chopped out, and the remaining pieces (called exons) are stuck back together
End modifications increase the stability of the mRNA, while splicing gives the mRNA its correct sequence. (If the introns are not removed, they’ll be translated along with the exons, producing a “gibberish” polypeptide.)
are all genes transcribed at the same time
Not all genes are transcribed all the time. Instead, transcription is controlled individually for each gene (or, in bacteria, for small groups of genes that are transcribed together). Cells carefully regulate transcription, transcribing just the genes whose products are needed at a particular moment
the stretch of dna transcribed into an rna molecule and the bits of nearby dna that control it is called
a transcriptional unit
promoter
sequence at which proteins that use the dna as a template to make rna bind and initiate synthesis (like rna polymerase)
rna coding region
dna sequence that codes for the rna that is transcribed
transcription start site
location in the dna corresponding to the first rna nucleotide incorporated into the transcribed rna
terminator
a dna sequence that disrupts the activity of the rna polymerase and induces it to stop transcription
transcriptional termination site
dna site of the last nucleotide incorporated into the transcribed rna
upstream
sequences before the transcription start site such as promoter and other regulatory elements
downstream
sequences after the transcription start site including the coding sequence of the gene and transcriptional termination site
non template strand
after it is transcribed the rna will have a sequence similar to this strand of dna
template strand
the rna polymerases uses base-pairing with this strand to create a complementary rna transcript
why can you have transcription in either direction
because dna is double stranded
explain transcription
which mushroom is a potent inhibitor of rna polymerase
amanita
lac operon
typically needs glucose, but in absence of glucose begins to break down lactose for energy
transcriptional activator
According to the conventional wisdom, transcription factors are typically classified as “activators” or “repressors”. Activators recruit coactivators, resulting in gene activation, while repressors recruit corepressors, leading to transcriptional repression.
A transcriptional activator is a protein (transcription factor) that increases transcription of a gene or set of genes. Activators are considered to have positive control over gene expression, as they function to promote gene transcription and, in some cases, are required for the transcription of genes to occur
An activator facilitates the up regulation of the transcription process by binding to enhancers, while promoter is the site at which RNA polymerase binds, and transcription initiation takes place, and repressor down regulates transcription by binding to silencers
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what is the difference between exons and introns
exons are amino acid coding
dna is transcribed into rna, and introns are removed by rna splicing
post-transcriptional processing
capping and poly-a tail addition make the message stable enough to leave nucleus to go to ribosome
alternative splicing
different exon are used in a different situation, meaning that a single gene can produce a large number of different proteins
how to shut down genes using rna (rna interference or RNAi)
inject complementary strand of the messenger rna, so double stranded rna which cant be translated so gene essentially shuts down
Dicer protein
The Dicer protein aids in the production of a molecule called microRNA (miRNA). MicroRNAs are short lengths of RNA, a chemical cousin of DNA. Dicer cuts (cleaves) precursor RNA molecules to produce miRNA. MicroRNAs control gene expression by blocking the process of protein production
Small interfering RNA (siRNA)
siRNA is a synthetic RNA duplex designed to specifically target a particular mRNA for degradation
Small interfering RNA (siRNA) are double-stranded RNA molecules that offer new opportunities for therapeutic intervention because they act inside the cell to reduce protein expression. They do this by targeting RNA to prevent the production of disease-associated proteins
micro rna
MicroRNA are small, single-stranded, non-coding RNA molecules containing 21 to 23 nucleotides. Found in plants, animals and some viruses, miRNAs are involved in RNA silencing and post-transcriptional regulation of gene expression
dont need the exact complement, more tolerating than sirna
siRna vs miRna
sirna: specifically target a particular mRNA for degradation
miRNAs (microRNAs) are short non-coding RNAs that regulate gene expression post-transcriptionally. They generally bind to the 3’-UTR (untranslated region) of their target mRNAs and repress protein production by destabilizing the mRNA and translational silencing.
what is upstream of the rna coding region
promoter
what is a promoter
dna sequence at which proteins such as rna polymerase bind to initiate transcription
what do activator and repressor transcription factors do
enhance or inhibit rna polymerase binding to the promoter
what does the core/minimal promoter contain
minimal sequences required for transcription
tata box
dna sequence bound by the tata binding protein and other core transcriptional machinery (deletion/disruption of tata box prevent transcriptions entirely, because without it, the general transcription
factors and RNA polymerase will be unable to bind and initiate RNA synthesis)
which forms of small rna molecules involved in gene regulation start out as double stranded rna molecules
small interfering rnas and micro rnas
what enzyme complex picks up double stranded rna molecules and dices them into short sequences
dicer
these 21 base pair RNAs then form a complex with more proteins to produce the
RNA-induced Silencing Complex (RISC). The “silencing” component of the name comes from its activity: it acts to silence the target gene by interfering with
its translation into protein. One of the two strands of the RNA (either siRNA or miRNA) is eliminated.
The active RISC has a single stranded
21 nucleotide RNA that is called the
guide RNA. This is complementary to
a target sequence on the mRNA of the gene that is being regulated. For siRNAs, that complementarity applies to the entire 21 nucleotide sequence. Therefore, siRNA binding is highly specific (it will only bind to a single specific sequence). For miRNAs, that complementarity only applies to a short segment of the guide RNA, meaning that an miRNA will bind to
many more mRNA sites than an siRNA (i.e. an miRNA’s mRNA binding is much less specific than an siRNA’s).
Both siRNAs and miRNAs mess up translation of the protein, effectively silencing the gene. how?
siRNAs typically result in the mRNA being cleaved at the site of siRNA attachment; a snapped mRNA cannot be used for translation and is degraded by the cell. miRNAs, in contrast, do not typically break the mRNA, but, rather, inhibit translation by simply getting in the way of the translation machinery
core promoter
The core promoter region is located most proximal to the start codon and contains the RNA polymerase binding site, TATA box, and transcription start site
regulatory
The regulatory sequences include the promoter region together with enhancer elements. Every gene has a promoter, which is the binding site for the basal transcriptional apparatus - RNA polymerase and its co-factors. This provides the minimum machinery necessary to allow transcription of the gene
