Chapter 11: General Transcription Factors in Eukaryotes

Question 1

TFIIIC

Answer 1

600kD protein w/ 6 subunits and “linker” region (dumbell shaped) that can contact 2 internal promoters simultaneously and can stretch to span large distances (required for all class III transcription)

Question 2

GC-rich box/region

Answer 2

located in genes lacking TATA box upstream of transcription start site

Question 3

determined by Reinberg et al to be important elongation factor using radio-labelled nucleotides in vitro

-prevents RNA polymerase from sustained pausing at specific pause sites in DNA & triggers proofreading by stimulatig RNase enzyme to remove RNA & resume transcription

Answer 3

TFIIS (x2)

Question 4

TAF250 & TAF150 (x3)

-other functions=acetyltransferase or protein kindase enzyme that phosphorylates other proteins

Answer 4

essential for TFIID binding to promoter; can bind initiators and DPEs without binding TATA box: can function as coactivators b/c can stimulate transcription in TATA box/TBP absence (recruit TBP and other TAFs to complex)

-other functions?

Question 5

UBF (upstream binding factor) (x3)

-binds to upstream promoter element (UPE)

Answer 5

bends the DNA to help SL1 bind to the core promoter element by functioning as an assembly factor (coactivator)

• composed of 2 polypeptides, sufficient to stimulate rRNA transcription
• binds to?

Question 6

pre-initiation complex

Answer 6

promoter with RNA Pol & trxn factors assembled there

-tightly bound, initiates open promoter complex (DNA melted)

Question 6

TFIIIB (x3)

Answer 6

orients polymerase at start site and tightly binds DNA at upstream promoter, promoting several rounds of transcription

-required for all classIII gene expression

Question 6

Sentenac et al

Answer 6

observed TFIIIC attachment to different sizes of tRNA genes using SEM

Question 6

zinc finger protein with 2 cysteines/histidines that binds the 5S rRNA internal promoter with TFIIIC, triggering TFIIIB binding to upstream promoter; essential for 5S rRNA transcription

Answer 6

TFIIIA

Question 7

observed TFIIIC attachment to different sizes of tRNA genes using SEM

Answer 7

Sentenac et al

Question 8

bends the DNA to help SL1 bind to the core promoter element by functioning as an assembly factor (coactivator)

• composed of 2 polypeptides, sufficient to stimulate rRNA transcription
• binds to?

Answer 8

UBF (upstream binding factor)

-binds to upstream promoter element (UPE)

Question 9

composed of 30 and 70kD peptides that tightly/specifically bind RNA polymerase and prevents nonspecific DNA binding by polymerase

-other function?

Answer 9

TFIIF

-can prevent short pausing at pause sites

Question 10

TFIIE

Answer 10

enhances TFIIH kinase activity

Question 11

TFIIIA (x4)

Answer 11

zinc finger protein with 2 cysteines/histidines that binds the 5S rRNA internal promoter with TFIIIC, triggering TFIIIB binding to upstream promoter; essential for 5S rRNA transcription

Question 11

promoter with RNA Pol & trxn factors assembled there

-tightly bound, initiates open promoter complex (DNA melted)

Answer 11

pre-initiation complex

Question 13

TAFs (TBP-associated proteins)

Answer 13

14 ID’d by precipitating TBP and western blotting (8 in Drosophila, 12 in yeasts)

-function=interact with promoter initiator, downstream promoter element (DPE), and gene-specific transcription factors

Question 14

TBP (universal transcription factor) (x4)

-structure=saddle-shaped; function=opens up DNA and bends TATA box

Answer 14

38/43kD protein rich in basic amino acids (giving positive charge) which binds to the minor groove within the TATA box by mutating nucleotides in both major and minor grooves; recruits TAFs to assemble preinitiation complex

-structure/function?

Question 15

binds to GC domains and recruits TAF-110 (130 human) which functions as bridge for TAF250-150. These then recruit TBP to form preinitiation complex and begin transcription

Answer 15

Sp1 (an activator)

Question 16

TFIIS (x3)

Answer 16

determined by Reinberg et al to be important elongation factor using radio-labelled nucleotides in vitro

-prevents RNA polymerase from sustained pausing at specific pause sites in DNA & triggers proofreading by stimulatig RNase enzyme to remove RNA & resume transcription

Question 17

located in genes lacking TATA box upstream of transcription start site

Answer 17

GC-rich box/region

Question 18

TFIIF (x2)

-can prevent short pausing at pause sites

Answer 18

composed of 30 and 70kD peptides that tightly/specifically bind RNA polymerase and prevents nonspecific DNA binding by polymerase

-other function?

Question 19

contains TATA-Box binding protein (TBP) and TBP-associated proteins/factors (TAFs)

Answer 19

TFIID

Question 19

found evidence of pre-formed preinitiation comlex: RNA polymerase holoenzyme

-contains?

Answer 19

Kornberg & Young

-contains TFIIB, A, F, & H

Question 20

essential for TFIID binding to promoter; can bind initiators and DPEs without binding TATA box: can function as coactivators b/c can stimulate transcription in TATA box/TBP absence (recruit TBP and other TAFs to complex)

-other functions?

Answer 20

TAF250 & TAF150

-other functions=acetyltransferase or protein kindase enzyme that phosphorylates other proteins

Question 21

complete preinitiation complex; TFIIE then TFIIH bind DABPolF complex

Answer 21

DABPolFEH complex

Question 22

DA complex recruits TFIIB -function=recruits TFIIF & RNA pol2 which join complex upstream of start site

Answer 22

DAB complex -function?

Question 22

ID'd essential class III transcription factors (TFIIIA, B, and C)

Answer 22

Roeder et al

Question 23

discovered SL1 requirement by incubating in increasing concentration with rRNA promoters and measuring transcription levels - also demoed that UBF greatly enhances transcription w/ rRNA upstream promoter element - analyzed SL1 protein complex with immunoprecipitation

Answer 23

R. Tijan et al (x3)

Question 24

35kD protein w/ 2 key domains: one interacts with TBP (of TFIID), the other with RNA polymerase/TFIIF near transcription start site

Answer 24

TFIIB

Question 25

Key featuers: 1. class I genes 2. class II genes 3. class III genes

Answer 25

1. utilize UBF as key assembly factor (lack TATA box) 2. may use Sp1 or TAF250-150 3. utilize TFIIIC to bind TFIIIB and help assemble preinitiation complex

Question 26

14 ID'd by precipitating TBP and western blotting (8 in Drosophila, 12 in yeasts) -function=interact with promoter initiator, downstream promoter element (DPE), and gene-specific transcription factors

Answer 26

TAFs (TBP-associated proteins)

Question 28

developed specific binding sequence of class II proteins and enzymes

Answer 28

Reinberg et al

Question 30

TFIID

Answer 30

contains TATA-Box binding protein (TBP) and TBP-associated proteins/factors (TAFs)

Question 31

600kD protein w/ 6 subunits and "linker" region (dumbell shaped) that can contact 2 internal promoters simultaneously and can stretch to span large distances (required for all class III transcription)

Answer 31

TFIIIC

Question 32

core biding protein that recruits polymerase1 - complex composed of TATA-box binding protein (TBP) and 3 unique TAFs: TAF110, 63, & 48 - binds?

Answer 32

SL1 -binds core promoter element

Question 34

Kornberg & Young -contains TFIIB, A, F, & H

Answer 34

found evidence of pre-formed preinitiation comlex: RNA polymerase holoenzyme -contains?

Question 35

1. utilize UBF as key assembly factor (lack TATA box) 2. may use Sp1 or TAF250-150 3. utilize TFIIIC to bind TFIIIB and help assemble preinitiation complex

Answer 35

Key featuers: 1. class I genes 2. class II genes 3. class III genes

Question 36

orients polymerase at start site and tightly binds DNA, promoting several rounds of transcription

Answer 36

TFIIIB

Question 37

SL1 (x2) -binds core promoter element

Answer 37

core biding protein that recruits polymerase1 - complex composed of TATA-box binding protein (TBP) and 3 unique TAFs: TAF110, 63, & 48 - binds?

Question 39

R. Tijan et al (x3)

Answer 39

discovered SL1 requirement by incubating in increasing concentration with rRNA promoters and measuring transcription levels - also demoed that UBF greatly enhances transcription w/ rRNA upstream promoter element - analyzed SL1 protein complex with immunoprecipitation

Question 40

DAB complex -function?

Answer 40

DA complex recruits TFIIB -function=recruits TFIIF & RNA pol2 which join complex upstream of start site

Question 41

38/43kD protein rich in basic amino acids (giving positive charge) which binds to the minor groove within the TATA box by mutating nucleotides in both major and minor grooves; recruits TAFs to assemble preinitiation complex -structure/function?

Answer 41

TBP (universal transcription factor) -structure=saddle-shaped; function=opens up DNA and bends TATA box

Question 42

TFIIB (x2)

Answer 42

35kD protein w/ 2 key domains: one interacts with TBP (of TFIID), the other with RNA polymerase/TFIIF near transcription start site

Question 43

TFIIH (x3) - function=enhance DNA melting and moving polymerase across target gene promoter - demoed by ability to separate 41 base strand from phage M13 DNA

Answer 43

large complex protein w/ 9 subunits; functions as protein kinase b/c can phosphorylate RNA Pol2 w/in its CTD while it's bound to DNA (requires ATP) - function? - also contains protein helicase activity (capable of unwinding DNA) required for transcription b/c allows polymerase to clear promoter and elongate new RNA strand (requires ATP) demoed by?

Question 44

large complex protein w/ 9 subunits; functions as protein kinase b/c can phosphorylate RNA Pol2 w/in its CTD while it's bound to DNA (requires ATP) - function? - also contains protein helicase activity (capable of unwinding DNA) required for transcription b/c allows polymerase to clear promoter and elongate new RNA strand (requires ATP) demoed by?

Answer 44

TFIIH (x2) - function=enhance DNA melting and moving polymerase across target gene promoter - demoed by ability to separate 41 base strand from phage M13 DNA

Question 45

Reinberg et al

Answer 45

developed specific binding sequence of class II proteins and enzymes

Question 47

Sp1 (an activator)

Answer 47

binds to GC domains and recruits TAF-110 (130 human) which functions as bridge for TAF250-150. These then recruit TBP to form preinitiation complex and begin transcription

Question 48

first one formed in preinitiation complex; TFIID and TFIIA bind TATA box

Answer 48

DA complex

Question 49

DA complex

Answer 49

first one formed in preinitiation complex; TFIID and TFIIA bind TATA box

Question 50

enhances TFIIH kinase activity

Answer 50

TFIIE

Question 51

DABPolFEH complex

Answer 51

complete preinitiation complex; TFIIE then TFIIH bind DABPolF complex

Question 52

Roeder et al

Answer 52

ID'd essential class III transcription factors (TFIIIA, B, and C)