Lect. 9/23 - Transcription + Ch. 7 RQ

What are general transcription factors?

(Euk only) -assemble at the core promoter -recruit RNA Polymerase II to enable transcription. -bind to specific sites on DNA to activate transcription

What is TBP? What does it do?

(GTF) TATA binding protein -Binds to the TATA Box -Signals binding site for RNA pol I, II, and III. -Subunit of TFIID

What is 5' capping and what is it's purpose?

-Addn' of backwards guanine to 5' end of an mRNA mol. connected by a triphosphate bridge -Addn' of methyl to backwards guanine -It is a form of mRNA modification in eukaryotic cells

What is TFIIE?

-GTF -Attracts and regulates TFIIH

What is TFIIB and what does it do?

-GTF -Recognizes the B recognition element in promoters -Pos. RNA Pol's at the start site of transcription -Connection of this and RNA pol stabilized by TFIIF

What is TFIIF?

-GTF -Stabilizes RNA pol. interaction w/ TFIIB and TBP -Attracts TFIIE and TFIIH

What is TFIIH?

-GTF -Unwinds DNA @transcr. start point -Phosphorylates C-terminal domain on RNA pol. -Releases RNA pol from promoter

Why is RNA pol. 2 the one we focus on most

-Its the most similar to bacterial RNA pol. - It transcribes all protein encoding genes

What is splicing and what is its purpose?

-Removal of introns from a DNA seq. -Form of mRNA mod.

How is RNA different than DNA?

-Single stranded -Uricil instead of thymine -Ribose instead of deoxyribose -usually a lot shorter

What is 3' tailing and what is its purpose?

-Stop codon reached -Ribonuclease cleaves off 3' of RNA -Poly A polymerase adds 100's of adenine residues to the cleaved 3' of the mRNA -Form of mRNA mod.

What does RNA polymerase do?

-binds to DNA and separates DNA strands -uses one strand of DNA as a template from -assembles nucleotides into a complementary strand of RNA -rewinds DNA behind it

What type of cell uses a sigma factor for binding of the RNA polymerase to the DNA promoter?

Bacteria

How are transcription and translation modulated with respect to their final products?

More RNA can be made from a given gene during transcription which can then produce more protein

Where does transcription and translation occur in a eukaryote? Do they happen at they same time? Why or why not?

Transcr=nucleus Transl= cytoplasm They do not happen at the same time bc pre mRNA made that needs to be processed during transcription

The process of gene expression always involves which process(es) described in the central dogma? -transcription -translation -replication -transcription and translation

Transcription Some genes that are expressed are functional RNA genes. These RNAs don't undergo translation to protein because they do their job in the cell as an RNA.

The catalytic sites for peptide bond formation during translation is found in which part of the ribosome? -large subunit RNAs -large subunit proteins -small subunit RNAs -small subunit proteins

large subunit RNAs The catalysis of the peptide bonds in the growing polypeptide chain during translation is performed by the rRNA of the large subunit. The ribosome is a ribozyme; proteins play a largely structural role.

What is miRNA and what does it do?

micro RNA; able to silence mRNA by binding to it preventing the its coding of proteins

What is pre-mRNA? How do we get from that to mature mRNA

(in prok. no pre-mRNA) Pre-mRNA is what is produced from transcription. It is then processed through the techniques of intron splicing, 5' capping, and addn' of a poly-A tail on the 3' end.

At which step of gene expression can cells amplify the number of copies of a protein made from a single gene? -transcription -translation -neither transcription nor translation -both transcription and translation

-both transcription and translation Proteins can be made in large quantities by transcribing many mRNAs from the gene, and then each mRNA can be translated into many copies of the protein. In contrast, if just a few mRNAs are made, only a few copies of the protein are made.

What is TFIID?

-composed of TBP and TAFs -Regulates binding by TBP

What is tRNA? How does it help facilitate elongation?

-transfer RNA -have anticodons that bind to specific codons and at the other end of the molecule are already bound to an a.a. -tRNA enters (w/ a.a.) on A-site of ribosome by binding of its anitcodon to the given codon -peptide bond forms between that a.a. and the one on the tRNA in the P-site of the ribosome -Ribosome shifts to right to translate next codon on DNA

What is a promoter? What does it consist of?

A sequence of nucleotides on the DNA that designates a start point for trx. Made of: -recognition site for the RNA polymerase to recog. and bind to DNA -TATA box where RNA pol seperates the two strands of DNA

Why and when is the C-terminal tail of RNA pol II phosphorylated?

Allows for RNA processing Done as soon at the RNA comes free

How does RNA polymerase know where to start transcribing a gene into mRNA?

Attaches to the promoter sequence on the DNA which consists of the B recognition element and the TATA box

Why does transcription happen in the cytoplasm of bacteria?

Because they don't have a nucleus

What does the sigma factor do?

Bind to RNA polymerase to recognize and binds it to the promoter sequence in bacteria Released shortly after transcription begins

Which of the following is a difference between the mechanisms of DNA polymerase and RNA polymerase? -The direction of polymerization for DNA polymerase is 5′ to 3′, but RNA polymerase is 3′ to 5′. -DNA polymerase uses DNA as a template for making DNA copies; RNA polymerase uses RNA as a template for making RNAs. -DNA polymerase uses the energy from the hydrolysis of the nucleotide triphosphates to drive the reaction; RNA polymerase uses reduced electron carriers. -DNA polymerase needs a base-paired 3′ −OH for a polymerization reaction to occur; RNA polymerase can polymerize two nucleotides without a base-paired 3′ −OH.

DNA polymerase needs a base-paired 3′ −OH for a polymerization reaction to occur; RNA polymerase can polymerize two nucleotides without a base-paired 3′ −OH. DNA polymerase needs a primer to provide a base-paired 3′ −OH to catalyze the polymerization reaction. RNA polymerase does not need a base-paired 3′ −OH, it can join two nucleotides together without a primer.

Protein concentration can be regulated by all of the steps listed EXCEPT -nuclear export. -RNA processing. -mRNA stability. -DNA replication.

DNA replication Protein levels can be tuned by changing the amount of processed mRNA available to ribosomes. Protein levels are not typically regulated by making more copies of the DNA instructions.

If RNA polymerase cannot intiate transcription on its own and eukaryotic cells do not have a sigma factor, how does transcription get initiated in eukaryotic cells?

General transcription factors

True or False: Once initiation of RNA transcription begins, the whole RNA strand muct be synthesized and released before that RNA pol. can go back to synthesize another RNA mol.

False A soon as a region of the gene is free, a different RNA pol will attach to synthesize another RNA mol. even while the first one is attached

True or False: Transcription is always followed by translation giving the central dogma of molecular biology: "DNA-->RNA-->protein"

False For most genes, a protein is the final product, but for some genes, a functional RNA like tRNA or rRNA is the final product.

True or False: All genes that are transcribed come from the same strand

False Genes can be transcribed from either strand of DNA. Where transcription of the gene occurs is based on the location of the promoter relative to the gene

True or False: All newly transcribed RNA stays attached to the DNA until a stop terminator seq. is reached.

False Only the RNA actively being synthesized is attached to the DNA. Otherwise, the DNA is rewound after transcription by the RNA polymerase and the synthesized RNA floats free in the nucleus

What is mRNA and what does it do?

Messenger RNA; mRNA carries instructions on how to make specific proteins. (coding RNA)

Where does energy required for transcription come from and go to?

Hydrolysis of the second and third PO₄ bond on the incoming nucleotide in formation of the phosphodiester backbone bond between nucleotides

How does being single stranded alter the function of RNA so that it's different from a DNA double helix?

Noncoding RNA can acquire a 3D shape by H-bonding with complementary sequences on itself. EX: tRNA that facilitates a.a. addn' to a growing peptide strand

The splicing of introns out of an mRNA molecule is catalyzed by: -RNA molecules that base pair with the splice sites to promote intron removal. -proteins that contain metal ions to pull electrons from the phosphate bonds. -RNA molecules that act as a template for new RNA synthesis using exons only. -proteins that act as nucleases to chew away and remove the introns.

RNA molecules that base pair with the splice sites to promote intron removal. Splicing is actually an RNA-catalyzed reaction. The RNA molecules in the snRNPs base pair with regions of the RNA that are to be spliced. This base pairing helps promote the rearrangement of bonds to remove the intron.

What rewinds DNA after transcription?

RNA polymerase

What unwinds DNA for transcription?

RNA polymerase

What is the complementary RNA transcript synthesized by? How about the protein?

RNA polymerase synthesizes RNA from a DNA template Ribosomes synthesize the protein using mRNA as a template

What recognizes the stop codons in an mRNA? How does it do it? -release factor -a specialized tRNA linked to methionine -nothing -the 3′ poly-A tail

Release factor When the ribosome encounters a stop codon, instead of a tRNA binding, a protein called release factor binds and catalyzes the addition of a water molecule to the carboxyl end of the polypeptide and releases it.

*FLIP FOR QUESTION* *ANSWER ON THIS SIDE* It would use the bottom strand because the promoter sets the direction and the polymerase moves from 3′ to 5′ along the template strand. RNA polymerase binds to promoter sequences in a specific orientation. The RNA polymerase will then move away from the promoter using the promoter-dictated directionality and uses the 3′ to 5′ strand as a template to make a new RNA in the 5′ to 3′ direction.

Shown below is a gene with the direction of transcription noted. How does the RNA polymerase know which strand to use as a template for the RNA, and which strand would it choose in this case? -It would use the top strand as a template because the polymerase always moves from left to right. -It would use the top strand as a template because the promoter marks beginning of the gene. - It would use the bottom strand because the promoter sets the direction and the polymerase moves from 3′ to 5′ along the template strand. -It would use the bottom strand because the bottom strand is always the template strand.

What is the equivalent of general transcription factors in bacteria?

Sigma factor

What happens to the sigma factor and RNA polymerase once transcription is complete?

Sigma factor is released as soon as RNA pol. is bound to DNA. RNA pol released after transcription is complete. After transcription, they rebind together to prepare for another round of transcription

What are the ways that pre mRNA is processed?

Splicing capping poly-A tail addn'

Which characteristic of a replicating RNA polymerase allows multiple transcripts to be made simultaneously from the same region of DNA? -The RNA transcript dissociates from the DNA template immediately once complete. -RNA polymerase is very small. -RNA remains bound to the gene and helps template more transcripts. -RNA polymerases can jump over slower RNA polymerases to make more transcript.

The RNA transcript dissociates from the DNA template immediately once complete. RNA transcripts dissociate immediately from the DNA template once they are made. This allows many RNA polymerases to form a "caravan" on the gene, producing many transcripts.

How do you know whether the top or bottom strand will serve as the template strand for transcription?

The location of the promoter relative to the gene

Where does transcription and translation occur in a prokaryote? Do they happen at the same time?

They happen at the same time (bc mature mRNA made; no processing required) in the cytoplasm (bc no nucleus in prok.

How do tRNAs become attached to the correct amino acid? -aminoacyl-tRNA synthetases -RNA polymerases -rRNA ribozymes -translation initiation factors

aminoacyl-tRNA synthetases Enzymes called aminoacyl-tRNA synthetases recognize tRNAs with a specific anticodon as well as the amino acid for that tRNA and catalyze a reaction to join them together. This is called "charging" a tRNA.

What performs the function of bacterial sigma factor in eukaryotes? -general transcription factors -initiation proteins -nucleosomes -promoters

general transcription factors Sigma factor is an accessory protein to RNA polymerase in bacteria that helps the polymerase bind to the promoter region. In eukaryotes, this function is performed by a set of proteins called the general transcription factors.

Which type of RNA is converted into protein for performing its cellular function? -tRNA -rRNA -mRNA -miRNA

mRNA There are many different types of RNA that perform functions as RNA in the cell and are never translated into protein. These include tRNA, rRNA, and miRNA. The only RNAs that code for proteins are mRNAs.

What is the ribosome made of?

proteins and rRNA

What is rRNA and what does it do?

ribosomal RNA; it actually makes up the ribosome in addn' to protein. Preforms catalysis of the peptide bonds in the growing polypeptide chain during translation.