Genetics - Lecture 7, 8, 9

Front 1

Gene expression is mediated by _______ that act on ________.

Which one is cis, which trans?

Back 1

Gene expression is mediated by FACTORS (protein, RNA molecules or complexes) that work on ELEMENTS (present ON the DNA, RNA or protein).

FACTORS act in TRANS (exert effects on many different molecules)

ELEMENTS act in CIS (direct action on only the sequence where present)

Front 2

Do mutations in Trans acting factors or Cis acting elements have more widespread effects?

Back 2

Factors acting in TRANS have more widespread effects bc trans mutations can act on many molecules.

Front 3

Are Trans-acting factors or cis-acting elements usually the cause of inherited diseases?

Back 3

Cis acting elements are usually the culprit.

Trans acting factors are so widespread that they are lethal and not viable.

Front 4

What is a promoter?

What binds to this? What does this binding do?

Back 4

cis-acting sequences in DNA that are adjacent to the transcribed region.

The trans-acting factors usually bind to this. This binding influences the ability of RNA polymerase to bind and work.

Front 5

What trans acting factors specifically act on RNA Pol II to cause it to make mRNA?

Back 5

General Transcription Factors (TF's)
ex) TFIID and TFIIB
AND
Gene Regulatory Proteins

Front 6

What are two types of TF's? Which one binds first?

Back 6

TFIID and TFIIB.

TFIID binds FIRST to the TATA box. TFIIB binds next. They both stay on the promoter while RNAP II works.

Front 7

What are some examples of gene regulator proteins that can help activate transcription by RNAP II?

Back 7

enhancers = if loop in >1000 bp long. can be downstream. (must activate)

upstream elements = can repress or activate.

nuclear receptors = bind small hydrophobic molecules w their effector.

Front 8

What are two things that all activator gene regulatory proteins contain?

Back 8

1. DNA-binding domain - recognize specific DNA sequences (8-10 bp's long)

2. an activation domain = interacts with TF's

Front 9

What is an effector domain?

What are some examples of an effector domain?

Back 9

An effector domain is a portion of a gene regulatory protein that alters its ability to activate transcription in response to a signal.

ex) binding of steroid hormone, phosphorylation by protein kinase.

Front 10

A gene coding for what enzyme is one example of regulation through combinatorial control?

Back 10

Phosphoenolpyruvate Carboxykinase.

GRE, CRE and HFN4 all play a role in regulation of expression here.

Front 11

What is one example of a nuclear receptor?

what is it's function?

Back 11

steriod hormone receptors are a class of nuclear receptors.

help an effector and ligand bind.

Front 12

What is the difference between a nuclear receptor and a cell surface receptor?

Back 12

Nuclear receptor (like steroid receptors) are on the nucleus - must be made out of hydrophobic materials. DIRECT action at nuclear site.
Cell surface receptors get to the nucleus through 2nd messenger systems like cAMP.

Front 13

Clotting factor IX gene has binding sites for which two nuclear receptors?

What is special about these sites?

Back 13

Clotting factor IX has an androgen receptor and HNF4a.

They overlap!

Front 14

Mutation in which nuclear receptors cause Hemophelia B?

Which hemophelia goes away after puberty and why?

Back 14

Androgen Receptor and HNF4a.

Leyden shows remission bc this mutation only affects the HNF4a part of the site.

Front 15

Which form of Hemophelia B does not show remission after puberty?

Back 15

Brandenberg version does not show remission because the mutation here is in the overlapping portion of HNF4a and androgen receptors. This means that post-pubertal increase in androgens won't help.

Front 16

What does a mutation in HNF4a cause?

Back 16

MODY

Front 17

What enzyme methylates the complement strand of Methylated DNA?

What base becomes methylated in either case?

Back 17

Maintenance Methyltransferase

A CYTOSINE = but the cytosine must preceed a guanine.
5'- CpG -3'

Front 18

What is imprinting?

Back 18

When epigenetic modifications dictate whether a gene is expressed or not NOT the actual DNA code for the gene. Also - this follows maternal or paternal pattern.

Front 19

What is an example of a gene that is imprinted where the paternal allele is actually expressed by methylation.

Back 19

Igf2

Front 20

What enzyme acetylates Lysine?

How does this affect expression?

Back 20

Histone Acetyltransferase (HATS)

Neutralizes charge on Lysine = destabilizes histone/DNA form (heterochromatic) which ACTIVATES transcription.

Front 21

What enzyme phosphorylates serine or threonine?

How does this affect expression?

Back 21

Histone Kinases

Adds - charge = destabilizes histone/DNA (heterochromatic) state which ACTIVATES transcription.

Front 22

What enzyme methylates Lysine?

How does this affect transcription?

Back 22

Histone Methyltransferase

this BLOCKS the acetylation of lysine...which stabilizes the heterochromatic state = INACTIVATES transcription

Front 23

Which AA's get acetylated in histone modification?

Effect?

Back 23

Lysine

Activation of transcription

Front 24

Which AA's get phosphorylated during histone modification?

Effect?

Back 24

Serine/Threonine

Activates transcription

Front 25

Which AA's get methylated during histone modification?

Effect?

Back 25

Lysine (arginine can too)

INACTIVATION of transcription -
because it BLOCKS the acetylation of Lysine.

Front 26

What is a nucleosome?

Back 26

About 140 bp's of DNA wraps around octamers of histones to form a little packet called a nucleosome. This is a storage form of DNA.

Front 27

Is the methylation of Cytosine reversible?

When does this process usually cause repression of a gene?

Back 27

The methylation of cytosine is reversible.

Usually causes repression of a gene when it is located on or near the gene's promoter.

Front 28

What are the 4 core histones?

What is important structurally about these?

Back 28

H2A, H2B, H3 and H4

They have N-terminal tails that are rich in Lys and Arg (+), that can react with DNA backbone phosphate (-) groups.

Front 29

Histone tails on the four core histones tails contribute to the stability of the nucleosome. What effect does this have on transcription?

Back 29

A stable nucleosome favors a HETEROCHROMATIC state. No transcription occuring. Pol II and transcriptional activators are blocked.

Front 30

Acetylation and methylation of what amino acid are competing reactions?

Back 30

LYSINE

acetylation - ACTIVATES transcription
methylation- blocks acetylation

Front 31

What inheritance pattern does the Sickle Cell Disease follow?

Back 31

autosomal recessive

Front 32

What kind of inheritance pattern does Cystic Fibrosis follow?

What gene is mutated to cause CF? What exactly is mutated about the gene?

Back 32

autosomal recessive

Cystic Fibrosis Transmembrane Regulator (CFTR)
- a Phe group is deleted.

Front 33

What three organs does CF primarily affect?

Back 33

Lungs, Pancreas, Intestine

Front 34

What test is used for people with CF that do have deltaF508?

What percent of people with CF have this mutation?

Back 34

ASO test. 70% do have this mutation

Front 35

What is an RFLP marker?

What is an example of when you might use this?

Back 35

RFLP markers take advantage of natural variations in DNA that happen to be linked to a gene or it's mutated form. You can digest an RFLP and use the results to find carriers/affected or unaffected in a family - depending on the circumstance.
Can use this in finding mutated CFTR gene in affected families.

Front 36

Why do you need tight linkage bt an RFLP and a mutant gene for this marker to be useful?

Back 36

If the two are not tightly linked - there is a greater chance for meiotic recombination which could unlink them.

Front 37

Give an example of a test used to detect Sickle Cell Dz in adults.

Back 37

Non-denaturing protein gel electrophoresis of Hb (the HgA will migrate faster toward + pole.

Front 38

What kind of test is necessary to detect Sickle Cell Disease in a fetus?

Back 38

DNA test necessary from amniotic fluid or chorionic villus sample.

Takes advantage of the fact that SCD mutation also destroys a sequence specific endonuclease called MstII - will cause HbS gene to clip into two fragments.

Front 39

If MstII is clipped into two fragments only - does the baby have HbS?

What if the baby is a carrier?

Back 39

yes.

Baby will have fragments of 3 different lengths, one whole fragment = HbA and two tinier fragments = HbS

Front 40

What test looks for the different lengths of MstII in SCD?

This test is used for ___?

Back 40

Southern Blot test - DNA

Front 41

What does TFIID bind to first?

What is the function of this TF?

Back 41

TFIID binds to the TATA box first...then TFIIB binds and then that whole complex is bound by RNA pol II.

This is all necessary for transcription.

Front 42

What is increased in response to increase in epinephrine or glucagon?

Back 42

increase in cAMP which increases CRE which increases transcription.

Front 43

What is increased in response to increase in cortisol?

Back 43

GRE is increased. This is a gene regulatory protein effector - serves to increase transcription.