Molecular Biology Exam Ii

Front 1

What are Basal factors?

Back 1

required for initiation at nearly all promoters; determine site of initiation; interact with TATA box

Front 2

What are upstream factors?

Back 2

DNA binding proteins that recognize consensus elements upstream of TATA box. Ubiquitous. Increase efficiency of initiation. Interact with proximal promoter elements (such as CCAAT box)

Front 3

What are Inducible (regulated) factors?

Back 3

Work like upstream factors but are regulatory. Made or active only at specific times or in specific tissues. Interact with enhancers or silencers.

Front 4

What are the factors for class II promoters?

Back 4

TFII A, B, D, E, F, H
Many are multi-subunit

Front 5

What are the functions of the TAFIIs (of TFIID)?

Back 5

1. strongly promote txn from promoters with I and D elements
2. X-linking (to DNA) and footprinting with different complexes showed that TAFII250 and TAFII150 bind Inr and D regions in cooperation wtih TBP
3. Promote txn from some class II promoters that lack a TATA box
4. Interact with some upstream activators (e.g. Sp1) and hence act as coactivators

Front 6

What are the characteristics of TFIIF?

Back 6

-2 subunits (RAP 70 and RAP 30)
-binds to RNAP and RAP 30 delivers it to DAB complex.
-Reduces RNAPs nonspecific binding to DNA (function analogous to sigma in e. coli)

Front 7

What are the characteristics of TFIIE?

Back 7

-Binds after Pol/TFIIF binds to the pre-initiation complex.
-2 different subunits, each required to stimulate txn

Front 8

What are the characteristics of TFIIH?

Back 8

1. required fro promoter clearance
2. complex protein with 9 subunits
3. has DNA helicase/ATPase activity for melting DNA at transcription bubble
4. Kinase activity, phosphorylating CTD of RNAP large subunit.

Front 9

What are the three complexes formed with Pol II?

Back 9

1. Minimal initiation complex
2. Active transcription complex
3. Elongation complex

Front 10

What are the factors required for initiation at Class I promoters?

Back 10

nRNAP I, SL1, and UBF

Front 11

Why is the core more important for initation at class I promoters?

Back 11

Because that is what RNAP binds to

Front 12

What is the function of UCE in class I promoters?

Back 12

Stimulate TXN

Front 13

What are the factors for nRNAP III?

Back 13

TFIIIA, C, and B.

Front 14

What is the model for assembly of pre-initiation complex on class III tRNA promoter?

Back 14

1. TFIIIC binds
2. TFIIIB binds (TBP is a subunit)
3. PolIII binds TFIIIB
4. When TXN starts, Pol III displaces TFIIIC

Front 15

What are three distinctive characteristics of ribosomal genes?

Back 15

1. different base composition
2. highly repetitive
3. localized to nucleolus

Front 16

How did Roeder and Rutter find 3 RNA polymerases?

Back 16

Using DEAE-Sephadex (ion-exchange) chromatography, they saw three peaks when assessed by both spectrometry (amount of protein) and activity (amount of radioactive UTP incorporated).

Front 17

What genes does each RNA Polymerase transcribe?

Back 17

I: rRNA precursor
II: hnRNA(become mRNA) and snRNA
III: tRNA precursor, 5s rRNA, other small RNAs

Front 18

What does alpha-amanitin inhibit?

Back 18

Pol II (and III at hight concentrations)

Front 19

What are the steps in epitope tagging?

Back 19

1. genetically attach small epitope to subunit
2. put gene in cell w/o functional gene
3. label proteins w/ 35S
4. Precipitate complex from solution using antibody
5. separate labelled polypeptides w/ SDS-PAGE
6. detect with autoradiography

Front 20

What do class II promoters consist of?

Back 20

Core Promoter and Upstream Promoter element. The core promoter can contain TATA box, TFIIB recognition element (BRE), Inr, or DPE.

Front 21

What genes do not usually have TATA boxes?

Back 21

1. Housekeeping genes
2. Developmentally regulated genes

Front 22

Is the TATA box always necessary for TXN?

Back 22

No, in some cases it only positions TXN start site

Front 23

What are initiators?

Back 23

Conserved sequences around txn start sites that are required for optimal txn. Efficiency can be enhanced by other promoter elements.

Front 24

What are two UPEs found in Class II promoters?

Back 24

GC boxes, CCAAT box

Front 25

What are the elements of class I promoters?

Back 25

1. Core element
2. UPE

Front 26

What are the different types of class III promoters?

Back 26

- promoter located entirely w/in gene itself
- promoter not w/in gene (similar to class II promoters)

Front 27

What are the major classes for DNA-binding motifs?

Back 27

1. Zinc-containing modules (such as zinc fingers and those in glucocorticoid receptors)
2. Homeodomains
3. bZIP and bHLH motifs

Front 28

What are the main classes of transcription-activating domains?

Back 28

1. acidic domains
2. glutamine-rich domains
3. proline-rich domains

Front 29

What are the main subunits of TFIID?

Back 29

TBP and 8-10 TAFIIs

Front 30

Which subunit of TF11D is highly evolutionarily conserved?

Back 30

TBP

Front 31

What is the function of TBP?

Back 31

Bind the TATA box in Class II promoters. Also operates at TATA-less promoters and with RNA Pol I and III

Front 32

How does TBP in TFIID bind to the TATA box?

Back 32

through the minor groove of the TATA box

Front 33

How does TFIID associate with the TATA box minor groove?

Back 33

TBP sits on the DNA and forces it to bend 80 degrees, opening the minor groove. Since the T-A step in a DNA helix is relatively easy to distort, this may explain why the TATA box is so conserved.

Front 34

What are the functions of TAFIIs?

Back 34

- Interact with promoter
- Interact with gene-specific transcription factors.

Front 35

Do zinc fingers bind in the minor or major groove of DNA?

Back 35

Major groove

Front 36

What are homeotic genes?

Back 36

transcription factors with a conserved DNA-binding domain (homeodomain) that resembles a HTH. bind as monomers

Front 37

What are the characteristics of bZIP/bHLH proteins?

Back 37

1. have DNA binding and dimerization domains
2. DNA binding region is very basic
3. Dimerization involves Leucine zipper or Helix-loop-helix domain
4. Can form heterodimers

Front 38

What is the structure of a leucine zipper?

Back 38

alpha helices form a coiled coil with interdigitating leucines

Front 39

What steps occur between transcription and translation?

Back 39

1. 5' Capping
2. 3' Maturation: cleavage and polyadenylation
3. splicing
4. transport of RNA to cytoplasm
5. stabilization/destabilization of RNA

Front 40

List the intron classes and their distributions

Back 40

Group I - common in organelles, nuclear rRNA genes of lower eukaryotes, some prokaryotes
Group II -common in organelles, also in some prokaryotes and archaea
Nuclear mRNA-ubiquitous in eukaryotes
Nuclear tRNA-some eukaryotes

Front 41

Which intron classes display RNA-catalyzed splicing? What are these RNA called?

Back 41

I and II (and possibly nmRNA)
Ribozymes

Front 42

What was the first self-splicing intron to be discovered? Who discovered it and when?

Back 42

Tetrahymena group I 26s rRNA gene.
T. Cech, 1981

Front 43

What RNA structures are seen in group I introns?

Back 43

1. G-C pairs
2. Stacked helices
3. Long-range base pairing
4. Triple helices (3 strands)

Front 44

What are the two types of protein splicing factors for group I introns?

Back 44

1. intron encoded - promote splicing of only the intron that encoded it
2. nuclear encoded - splice organellar introns

Front 45

What are the relative abundances of the major RNAs?

Back 45

rRNA (90%)
tRNA (5%)
mRNA (2%)
SRP RNA, snRNA, snoRNA, miRNA (3%)

Front 46

What is RNA splicing?

Back 46

The removal of IVS's that interrupt the coding region of a gene. Excision is accompanied by precise ligation of exons.

Front 47

Describe the experiment involving R-loop analysis. What was discovered as a result of this experiment?

Back 47

1. Cloned genomic DNAs of a few highly expressed genes were hybridized to RNA fractions
2. Visulized by EM
3. Loops form from RNA annealing to template strand and displacing coding strand

Led to discovery of split genes because internal loops were observed.

Front 48

What are the relationships between the four intron classes?

Back 48

1. Each has a distinctive structure
2. The chemistry of splicing (transesterfication rxns) for group I, II and nmRNAs is similar
3. Splicing pathway for group II and nmRNA is similar
4. Splicing of groups I, II and possibly nmRNA is RNA-catalyzed

Front 49

What are the characteristics of self-splicing introns?

Back 49

- require Mg+ for proper folding
- conserved secondary structure but not primary sequence
- Group I require guanosine nucleotide in first step

Front 50

What are the functions of an RNA cap?

Back 50

1.Protection from some ribonucleases
2.Enhanced translation
3.Enhanced transport from nucleus
4.Enhanced splicing of first intron for some pre-mRNAs

Front 51

What are the functions of the Poly-A tail?

Back 51

1. promote mRNA stability
2. enhance translation

Front 52

What is the mechanism for polyadenylation of pre-mRNA?

Back 52

1. Transcription extends beyond mRNA end
2.Transcript is cut at 3’ end of what will become the mRNA (in green)
3.PolyA Polymerase adds ~250 As to 3’ end
4.“Extra” RNA (in red) degraded

Front 53

What are the proteins that promote formation of the commitment complex in humans and in yeast?

Back 53

humans: SR proteins SC35 and SF2
yeast: BBP binds to U1 and Mud2p, helping to define the intron prior to splicing

Front 54

What model shows regulation of alternative splicing?

Back 54

Sex determination in drosophila

Front 55

What is Trans-splicing? In what organism was it first discovered?

Back 55

Intermolecular splicing of pre-mRNAs. Trypanosomes

Front 56

What is the function of Cap in splicing?

Back 56

Cap stimulates splicing of the first intron in a multi-intron pre-mRNA

Front 57

What is the function of CBP in splicing?

Back 57

CBP may be important for spliceosome formation in vivo on the first intron

Front 58

What effect does the poly-A tail have on splicing?

Back 58

Promotes the splicing of the last intron

Front 59

What is RNA Editing?

Back 59

Any process, other than splicing, that results in a change int eh sequence of a RNA transcript such that it differs from the sequence of the DNA template.