MTC - 17 - Transcription Factor

-_- where is DPE and what is its consensus sequence

+28 to +32, GGGCC

what does FOXA2 do? where is it?

- regulates fatty acid oxidation and ketogenesis during fasting - nucleus but incytosol if phosphorylated)

what does FOX01 do? where is it?

-regulates gluconeogenesis -nucleus (but in cytosol if phosphorylated by phosphokinase B PKB)

mechanism of glucorticoid receptor

1. 2 HSP and 2 glucocorticoid receptors are associated 2. 2 cortisol comes in and kick out HSP 3. activated receptors dimerize, move into nucleus and bind to coactivator (glucocorticoid response element GRE) 4. increase gene transcription

mechanism for thyroid hormone receptor

1. T3 binds receptor 2. thyroid receptor conjugated to RXR 3. converts HDAC domain to HAC domain (histone deacetylase > histone acetylase) 4. histone is aectylated and release DNA

what is the signaling pathways for insulin-FOX regulation?

1. insulin binds insulin receptor IR 2. IRS2 activates PKB 3. PKB moves into nucleus and phosphorylates FOX01 on insulin response element IRE 4. FOX01 leaves nucleus (inhibits gluconeogenesis)

what does effect metformin have on AMPK?

1. it activates LKB1 (liver kinase B1) 2. LKB1 phosphorylate AMPK 3. AMPK sequesters TORC2 from nucleus 4. TORC2 cannot combine with CREB (inhibiting gluconeogenesis) (TORC2 = Transducer of regulated CREB )

-_- 3 factors determining transcription factor DNA binding specificity

1. repeat sequence 2. base between 2 repeats 3. orientation of the repeats

4 types of nuclear receptors

1. steroid homodimer 2. RXR heterodimers (retinoid x receptor) 3. dimer receptors 4. monomer receptors

-_- structure of a ligand binding domain on a hormone nuclear receptor

12 α-helices 2 β-turns arranged in 3 layers (helices 3, 5, 10 & 11 form a hydrophobic binding pocket)

what is the zinc in the zinc finger stabilized by?

4 cysteine or histidine (via chelation)

(quiz) Phosphorylation is one of the posttranslational modifications in the regulation of transcriptional activity. Which transcription factor is translocated to nucleus upon phosphorylation to activate target gene expression? And what is its major target gene? A. CREB:Phosphoenolpyruvate carboxykinase B. PPARd:Carnitine Palmitoyltransferase 1 C. Foxo1:Glucose-6-Phophatase D. SREBP1:Fatty Acid Synthase E. GLUT4:Glucokinase

A. CREB:Phosphoenolpyruvate carboxykinase

-_- what half sites do steroid receptors recognize?

AGAACA

-_- what half sites do RXR/TR heterodimer receptors react with? what steroid receptor also recognizes this half site?

AGGTCA estrogen

PPARα targets what to cause what

Acyl Coa A syntase & CPT 1 (Carnitine palmitoyltransferase I) gene fatty acid oxidation to increase serum triglyeride level

(quiz) Which of the followings are activated by binding of lipophilic molecules? A. Forkhead box proteins B. Peroxisome proliferator activated receptors C. Sterol regulatory element binding proteins D. Signal transducer and activator of transcription E. Cyclic AMP response element binding protein

B. Peroxisome proliferator activated receptors

what are 3 types of direct repats?

DR1 - AGGTCAnAGGTCA DR2 - AGGTCAnnACCTCA DR3 - ACCTCAnnnnACCTCA

-_- what happens to RXR/RAR heterodimer when it binds to the direct repeat DR1 site and DR5 sites?

DR1: blocks transcription (RAR on top, RXR bottom) DR5: initiates transcription (RXR on top, RAR bottom)

what motif is a forkhead transcription factor, how does it bind?

FOX protein winged helix that binds to the consensus sequence as a monomer or dimer

-_- with insulin resistance, where will FOX01 and FOX02 be?

FOX01 will stay in nucleus; FOX02 will be in cytoplasm (pt will have lots of glucose made and fatty acid not oxidized)

what is FOXA2 sensitive to?

IRS1 and 2 (insulin receptor substrate 1 and 2)

-_- 3 monomeric receptors

NGFI-B SF-1 (splicing factor 1) FTF (fetoprotein transcription factor)

what does CRE signal the transcription of?

PEPCK (Phosphoenolpyruvate carboxykinase - used in gluconeogenesis pathway) (mnemonic like pepsi kinase, like sugar)

-_- what are the 6 RXR heterodimer nuclear receptors?

PPAR RXR/PPAR thyroid RXR/TR vitamin D3 RXR/VDR farnesoid (bile acid) RXR/FXR retinoic acids RXR/RAR oxysterols RXR/LXR

-_- how does PPAR action on PUFA anti-inflammatory?

PUFA is involved in inflammation

-_- 3 homodimeric receptors

RXR HNF4 (hepatocyte nuclear factor 4) COUPTFs (Chicken ovalbumin upstream promoter-transcription factor)

what is SCAP?

SCAP = sterol responsive activating protein

what is SREBP? what is its motif?

SREBP = sterol responsive element binding protein; basic helix-loop-helix leucine zipper transcription factor

(quiz) Which SREBP is activated by cleavage in the Golgi and functions to induce the transcription of HMG-CoA synthase, HMG-CoA reductase, squalene synthase and LDL receptor?

SREBP-2

(review) which of the following resides in the ER membrane in intact form? A. PPAR gamma B. SREBP1c C. FOX01 D. CREB E. GR

SREBP1c

4 domains of nuclear hormones

TAD: transactivating domain (binds coactivator) e.g. AF1 DBD: DNA binding domain - binds hormone response element LBD: ligand binding domain NLS: nuclear localization signal

-_- what 3 consensus sequences are in the core promoter region

TATA box initiator DPE (downstream promoter element) (important for TBP binding)

-_- what happens to helix 12 after binding

a conformation change that reveals the nuclear receptor (NR) box (1. conformation change causes a reposition of H12 forming an active AF-2 region 2. the active AF-2 region recruits co-activator via its LxxLL (i.e. nuclear receptor NR box) 3. H12 is packed into the binding pocket and forms an additonal H-bond with the ligand to stabilize the conformation)

-_- what does a coactivator usually do?

acetylates histones (opens the site for general transcription factors to bind to the core promter)

-_- function of a nuclear hormone receptor

activated by ligand bind to DNA response element activate transcription factor (e.g. estrogen receptor)

(review) which on of this is not a DNA binding motif? A. beta sheet B. zinc finger C. leucine zipper D. helix-turn-helix

beta-sheet

-_- what does the core promoter region contain

binding site for general (basal) transcription factors

-_- what does insulin do?

block FOX01 - no gluconeogenesis block FOXA2 - no fatty acid oxidation

what does TFiiA do

blocks inhibitor binding

does leucine zipper function as homodimer or heterodimer?

both

what does PEPCK do?

catalyzes enzymatic reactions for gluconeogenis in liver

what are helix-loop-helix mostly involved in?

cell differentiation (helix-loop-helix resembles a leucine zipper)

-_- what is recruited as a result of the activation of a transcription factor?

coactivators

what is the zinc in zinc finger stablized by

cysteine

PPAR agonists are used to improve homeostatis and insulin sensitivity in pt with (3)

diabetes fatty liver high lipid - hyperlipidemia

what are the 2 types of DNA response element sequence?

direct repeat palindrome

-_- what are the 5 steroid receptors?

glucocorticoid GR/GR mineralocorticoid MR/MR progesterone PR/PR androgen AR/AR estrogen ER/ER (AGGTCA) ! steroid receptors are all homodimers, all palindrome

Forkhead box proteins are involved in many physiological homeostasis mechanisms like

glucose and lipid metabolism

FOX01 is known to play a big role in

hepatic glucose production at insulin resistant state

if zinc fingers bind to a dimer palindrome of estrogen receptors what does this do to specificity

increases it

what are the 2 types of palindromes

inverted (mirrored then translated <-- -->) everted (mirrored then translated --> <--)

what kind of transcription factor is CREB (i.e. what motif and how sequence does it bind)?

leucine zipper that bind palindrome (cAMP response element CRE) as a homodimer (CREB = cAMP response element-binding protein)

what do PPARγ

lipoprotein lipase & GLUT4 gene for lipogenesis to decrease serum glucose and fatty acid, increase insulin sensitivity, increase adipose tissue number

where do zinc fingers bind

major groove of DNA (side chains may interact with the minor groove)

-_- is initiator the actual transcription start site?

no, it can also be upstream or downstream

where does a helix turn helix bind?

one helix fits into the major groove and 2 other lay parallel across it

-_- what IRS gene expression does insulin resistance block?

only IRS1

what is PPAR

peroxisome proliferator activated receptor

what does TFiiH do

phosphorylates C-domain of RNA polymerase for transcription

what are the ligands for PPARδ

polysaturated fatty acid (PUFA) synthetic agonists

what are the ligands for PPARα

polyunsaturated fatty acid (PUFA) fibrate drugs (hyperlipidemia Rx - clofibrate, bezafibrate, fenofibrate)

what are the ligands for PPARγ

polyunsaturated fatty acid (PUFA) thiazolidinodione (TZD) and avandia (for Type II diabetes)

what does TFiiB do (2)

recognize and bind BRE (transcription factor IIB recognition element i.e. signal upstream of TATA box); recruit TFiiF and TFiiE

-_- what does TFiiE and F do?

recruit RNA polymerase

-_- what is a NR box

sequence on the nuclear receptor that recruits co-activators (leucine-leucine-xx-leucine; like what happens when transcription factor is activated)

(review) how is the transcription factor SREBP1c released from ER membrane? A. phosphorylation B. hydrophobic ligand binding C. cAMP D. inhibit degradation E. site specific clevage

site specific clevage

what does SREBP2 do?

takes HMG-CoA reductace and LDL-receptor to regulate cholesterol synthesis and uptake (liver homeostasis)

what does SREBP1 do?

takes stearoyl CoA desaturase and fatty acid synthase (SCD-1 and FAS) for fatty acid synthesis (fatty liver and hypertiglyceridemia)

(review) nuclear hormone receptors are A. membrane bound B. localize to cytosol by insulin signaling C. only present in nucleus under normal condition D. transcripstion factor

transcription factor

what do PPARδ target to cause what

uncoupling protein, lipoprotein lipase and CPT1 gene for increasing FA oxidation and converting Type II to Type I muscle to increase exercise endurance

how does leucine zipper binds DNA

via its basic amino acid (+ charge lysine & arginine) interacting with the negatively charged DNA

when is gene transcription activated?

when a transcription factor binds to a DNA response element (unique cis-element in the proximal promoter region)

-_- 4 DNA binding motifs

zinc finger (most common; monomer or dimer) leucine zipper (dimer) helix-turn-helix helix-loop-helix (dimer) (motif: how transcription factors bind to DNA)

3 types of PPAR

α - liver δ - muscle γ - adipose tissue