Unit 3- Ch. 13-17 Flashcards
Differentiate between the structures of DNA & RNA
- DNA nucleotides contain deoxyribose, RNA nucleotides have ribose sugars
- pyrimidine uracil is present in RNA instead of thymine, one of the two pyrimidines found in DNA
- RNA consists of a single polynucleotide strand whereas DNA consists of two polynucleotide strands joined by hydrogen bonding between complementary bases
what is the location and functions of rRNA
-make up the ribosome, the site of protein assembly
-located in the cytoplasm
-pro and eukaryotic in cytoplasm
what is the location and functions of mRNA
-carries the coding instructions for a polypeptide chain from DNA to a ribosome
-after attaching to the ribosome, mRNA molecule specifies the sequence of AA in a polypeptide chain and provide a template for the joining of those AA
-located in nucleus and cytoplasm
-pro and eukaryotes
what is the location and functions of tRNA
-serves as link between the coding sequence of nucleotides in an mRNA molecule and the AA sequence of a polypeptide chain
-each tRNA attaches to one type of AA and helps incorporate that AA into a polypeptide chain
-located in cytoplasm
-pro and eukaryotes
Define transcription and know the general characteristics of the process
-Transcription: synthesis of an RNA molecule from a DNA template
-RNA synthesis is complementary and antiparallel to template strand
-only certain regions of the DNA are transcribed (mostly gene regions) rather than the entire chromosome
-only 1 of the 2 DNA strands is transcribed for each gene
-nucleotides are added to the 3’ end of the RNA molecule so the direction of synthesis is 5’ to 3’
Draw and label a transcription unit
promoter, RNA-coding region, terminator
Describe the bacterial transcription apparatus
-at the heart of most bacterial RNA polymerases are five subunits that make up the core enzyme
—–>catalyzes the elongation of the RNA molecule by the addition of RNA nucleotides
—–>sigma factor controls the binding of RNA polymerase to the promoter
-after sigma has associated with the core enzymes, RNA polymerase binds stably only to the promoter and initiates transcription at the proper start site
Bacterial RNA polymerase
-bacteria only have 1 RNA polymerase that transcribes all RNA (mRNA, tRNA, rRNA),
Eukaryotic RNA polymerases
-many RNA polymerases (named with Roman numerals) that transcribe different RNAs
-here are different RNA polymerases
that transcribe different types of RNA
-RNA pol II transcribes pre-mRNA*
Define and provide the function of the promoter
DNA sequences that transcription apparatus recognizes and binds
- indicates which of the strand to read and the direction of transcription
Describe what a consensus sequence is and why its presence is important within a promoter
short stretches of common nucleotides
the presence of consensus in a set of nucleotides usually implies that the sequence is associated with an important function
What are the steps of bacterial transcription
Initiation:
- sigma + core RNA polymerase bind to promoter
- unwinds DNA
-nucleotides incorporated—no primer required
- 2 phosphates cleaved for each new nucleotide added
-sigma released
Elongation:
-RNA polymerase continues adding nucleotides
-unwinding at front of bubble & rewinding behind bubble
-proofreading
Termination:—different genes can use different termination mechanisms. You only need to understand the
following mechanism:
1. RNA polymerase transcribes a terminator sequence, which consists of a sequence that forms a
hairpin structure in the RNA followed by several uracils in the RNA
2. the hairpin structure causes the RNA polymerase to pause
3. at this point, only A-U bonds (weak bonds) are holding DNA and RNA together— DNA and RNA
separate
Differentiate between general transcription factors and other transcription factors
General transcription factors:
-Necessary for any transcription to occur
-Combine with RNA pol and other proteins to form the basal transcription apparatus
Other transcription factors:
-Can increase or decrease transcription levels but not required for transcription
Differentiate between a core promoter and a regulatory promoter within eukaryotes
Core:
- immediately upstream of a gene
- basal transcription apparatus binds here, which is required for transcription
- there are several consensus sequences within the core promoter that are recognized by many different
proteins, but the only one you need to remember is the TATA box at -25
Regulatory:
- immediately upstream of core promoter
- Affects rate of transcription
- Not required for transcription
Know the steps of eukaryotic transcription (for RNA polymerase II)
Initiation:
- TFIID contains a TATA-binding protein (TBP) which binds to the TATA box within the core promoter
- General TFs + RNA pol + mediator bind to core promoter via TFIID
— TBP of TFIID positions active site of RNA polymerase over start site
- Other transcription factors:
— may bind to regulatory promoter
— may bind to enhancers
— affect transcription rate by interacting with the basal transcription apparatus via the mediator
Elongation:
- Similar to bacterial elongation
Termination:
- Just know that there are proteins that assist to remove the RNA polymerase and the RNA transcript from the DNA
-TFIID CAN BIND TO THE TATA BOX DUE TO THE TBP
Differentiate between an exon and an intron
many eukaryotic genes contain
- exons: RNA coding regions
- introns: noncoding regions called intervening sequences
Which organisms are introns common within and rare within
Introns are common in eukaryotic genes but are rare in bacterial genes
Introns are present in mitochondrial and chloroplast genes as well as in the nuclear genes of eukaryotes
List the 3 main steps in pre-mRNA processing
- addition of the 5’ cap
- addition of the poly(A) tail
- internal modifications (aka splicing)
Draw and label the structure of a mature mRNA
each AA of a protein is specified by a set of three nucleotides (a codon)
Describe the mRNA molecule
both eukaryotic and prokaryotic mRNA contains three primary regions:
- 5’ untranslated region - nucleotides do not encode any of the AA of a protein
- protein-coding region - comprises the codons that specify the AA sequence of the protein
- 3’ untranslated region - nucleotides not translated into AA
Know the type of cells that pre-mRNA processing occurs in as well as the location within the cell
Only in Eukaryotes within the nucleus
Describe how the 5’ cap is added as well as its function
-it is a guanine nucleotide added backwards (5’ to 5’ bond) to the 5’ end of the mRNA
-then methyl groups are added to the mRNA
Function:
-assists with binding of ribosome to mRNA for translation
-stabilizes mRNA
Describe how the poly(A) tail is added as well as its function
How? 2 steps:
-Cleavage 11-30 nucleotides downstream of AAUAA consensus near 3’ end
-Polyadenylation: many adenines are added
Function:
-Increases stability of mRNA
-Required for ribosome binding to 5’cap
Define splicing
another major type of modification by eukaryotic pre-mRNA
the removal of introns
Where does splicing occur?
takes place in the nucleus before RNA moves to the cytoplasm
Define spliceosome as well as its makeup
-Spliceosome is a large complex where splicing occurs
-Consists of several snRNPs (‘snurps’)
-snRNP = 1 snRNA + proteins
-each snRNP plays a different role in the splicing process but we will not worry about those details
Explain alternative splicing
the same pre-mRNA can be spliced in more than one way to yield different mRNAs that are translated into different amino acid sequences and thus different proteins
Explain multiple 3’ cleavage sites
another type of alternative processing requires multiple 3’ cleavage sites
- 2 or more potential sites for cleavage and polyadenylation are present in the pre-mRNA
2.may or may not produce a different protein, depending on whether the site is located before or after the stop codon
conclusion: both alternative splicing and multiple 3’ cleavage sites produce what?
different mRNAs from a single pre-mRNA
Explain the purpose of RNA interference (RNAi)
-Defense mechanism against viruses
-Regulation of gene expression
-NOTE: RNAi is only found in eukaryotes
Describe how RNAi works
-Double-stranded RNA gets chopped up by the enzyme Dicer
-The resulting RNA pieces are called microRNAs (miRNAs) or small interfering RNAs (siRNAs)—there are slight
difference between these 2 types of RNAs, but we won’t worry about that
-miRNAs/siRNAs form a complex with proteins—this complex is called RISC
-RISC pairs with the target mRNA (because the miRNA/siRNA is complementary to the target mRNA)
-The target mRNA can no longer be translated
-Thus the mRNA has been “interfered” with so that the gene that the mRNA came from will no longer be
expressed (no protein produced)
Explain the purpose of CRISPR RNA (crRNA)
-Think about this as the “immune system” of prokaryotes
-They defend prokaryotic cells against invasion of foreign DNA (DNA from bacteriophages and plasmids)
Describe the action of CRISPR RNAs
ACQUISITION:
-Foreign DNA (ie phage DNA) gets incorporated into the bacterial DNA—specifically it is inserted into a CRISPR array region of the bacterial genome
EXPRESSION:
-The CRISPR array gets expressed (transcribed into crRNA)
-The crRNA and CAS protein form a complex
INTERFERENCE:
-Now when the foreign DNA enters the cell again, the crRNA-CAS complex will bind to it (because the crRNA is complementary to it) and CAS cuts the foreign DNA rendering it nonfunctional
Define codon
one AA is encoded by 3 consecutive nucleotides in mRNA
What is meant by the degeneracy of the genetic code?
How many possible codons are there?
4^3 = 64 possible codons, but only 20 amino acids
3 codons are stop codons (specify end of translation)
61 codons are sense codons (encode AA)
Know what is meant by the degeneracy of the genetic code
What does degeneracy mean?
Degeneracy: 1 amino acid may be specified by more than one codon
there are 61 sense codons and only 20 different AA commonly found in proteins; the code contains more information than is needed to specify the AA and is said to be degenerate
AA may be specified by more than one codon
Know what is meant by the degeneracy of the genetic code
Some amino acids are carried by more than one _____
tRNA
each tRNA attaches to a single AA
there are 30-50 different tRNAs and only 20 different AA in proteins
some AA are carried by more than one tRNA
What is meant by wobble in the code?
Wobble (nonstandard base pairing at 3rd position (3’) of codon) allows 1 anticodon to pair with > 1 codon
wobble allows some tRNA to pair with more than one mRNA codon
Define translation
how AA are assembled into proteins
only mRNAs are translated into proteins
Where does translation occurs within the cell?
takes place on ribosomes
ribosome attaches near 5’ end of mRNA strand and moves to 3’ end, translating the codon as it goes
Explain the process of translation initiation in prokaryotes
-IF-3 binds to small ribosomal subunit
-IF-3/small subunit bind to mRNA
-The rRNA in the small subunit is complementary to Shine-Dalgarno sequence
-tRNA carrying n-formyl methionine + IF-2 + GTP all come in together and anticodon binds to start codon
-IF-1 joins this complex
-GTP is hydrolyzed to GDP and all IF’s leave the complex
-Large subunit joins
Explain the process of translation elongation
-After initiation, the fMet-tRNA is in the P site of the ribosome
-EF-Tu + GTP + charged tRNA enter A site
-GTP hydrolyzed to GDP and EF-Tu/GDP leaves
-Peptide bond forms between amino acids in P and A sites (peptidyl transferase is the enzyme that does this)
-tRNA in P site releases its amino acid so now the peptide chain is entirely on the tRNA at the A site
-ribosome shifts down by one codon (translocation) with the help of EF-G and GTP
-tRNA that was in P site is now in E site and then immediately leaves
-A site is now available to receive the next charged tRNA and the cycle continues
Explain the process of translation termination
-When stop codon is in the A site, a release factor comes to the A site
-Another release factor joins the ribosome and all components are released
Explain simultaneous transcription and translation and state whether this occurs in prokaryotic or eukaryotic cells.
in prokaryotic cells, transcription and translation are simultaneous; multiple ribosomes may be attached to the 5’ end of the mRNA while transcription is still taking place at the 3’ end
in eukaryotes, transcription and translation are separated in time and space
—–>transcription in nucleus
—–>translation in cytoplasm
Explain the relationship between how tightly packaged DNA is and whether transcription occurs or not
in the nucleus, histone proteins form octamers around which DNA tightly coils to create chromatin
chromatin structure represses gene expression
for a gene to be transcribed, proteins called transcription factors must bind to the DNA (and regulator proteins and RNA polymerase must also bind)
Provide 2 ways that histones can be modified and describe the effects of these modifications as well as the enzymes
involved
- Methylation of histones
—–>addition of methyl groups to histone proteins
—–> can either activate/repress transcription depending on which histone is modified and which particular AA in the histone tail is methylated - Acetylation of histones
—->histone methyltransferases add methyl groups
—->histone demethylases remove methyl groups from histones
Describe how DNA methylation is associated with gene regulation. What gets methylated and how does this result
in gene regulation?
another change in chromatin structure associated with transcription is the methylation of cytosine bases
Define CpG islands
DNA methylation is most common on cytosine bases adjacent to guanine nucleotides (CpG where p=phoshpate group in DNA backbone)
Define general transcription factors, other transcription factors, basal transcription apparatus, core promoter, and regulatory promoter.
-transcription in eukaryotes is regulated by proteins called transcription factors that bind to specific DNA sequences
-many transcription factors regulate by recruiting other proteins
-general transcription factors bind to the core promoter and are part of the basal transcription apparatus: the complex of RNA polymerase, transcription factors, and other proteins that assemble to carry out transcription
Differentiate between enhancers and silencers. What type of transcription factors bind to these? Where are enhancers and silencers located with respect to a gene?
-other TFs bind to regulatory promoter and to enhancers (located some distance from the gene)
-enhancers are regulatory elements that affect the transcription of distant genes
-other sequences, called silencers, have an inhibitory effect on the transcription of distant genes
-like enhancers, silencers are position and orientation independent, and they contain binding sites for transcription factors that decrease transcription
Describe how regulatory promoters and mediators are associated with eukaryotic gene regulation. Where are these located with respect to a gene?
-regulatory promoters typically contain several different consensus sequences to which different transcription factors can bind
-one of the components of the basal transcription apparatus is a complex of proteins called the mediator which interacts with RNA polymerase
Explain the purpose of an insulator and how they work
