Hemoglobinopathies

Question 1

Hemoglobin alpha cluster

Answer 1

We have four copies of alpha both of which are preceded by fetal zeta (5’)

Chromosome 16

Question 2

Hemoglobin beta cluster

Answer 2

epsilon
Ggamma
Agamma
delta 
beta

Question 3

How many copies of beta?

Answer 3

two total (one on each homolog 11)

Question 4

Embryonic hemoglobin?

Answer 4

zeta2epsilon2
alpha2epsilon2
zeta2gamma2

Question 5

fetal hemoglobin?

Answer 5

alpha2gamma2

Question 6

adult hemoglobin?

Answer 6

95% alpha2beta2

3.5% alpha2delta2

Question 7

Globin switching during development

alpha cluster?

Answer 7

5th week switch from zeta to alpha

Question 8

Globin switching during development

beta cluster?

Answer 8

Switch from epsilon to gamma in-utero

Switch from gamma to beta at birth

Question 9

Locus control region

Answer 9

10 kb upstream - Regulates globin transcription of beta cluster
Both timing and level

Question 10

What happens if we delete LCR of beta cluster?

Answer 10

Hispanic epsilongammadeltabeta thalassemia –> have much lower expression of beta isoforms

Question 11

Sickle Cell Disease

Answer 11

Single gene disease

Single point mutation at codon 6 in exon 1 of beta globin

Question 12

Sickle cell disease

Amino acid substitution

Answer 12

Glutamine for valine

Question 13

Hemaglobin C disease

Answer 13

Same codon affected as sickle cell disease but different mutation (effects first position of codon and sickle effects second)- results in glutamine to lysine transition

Question 14

Sickle cell and hemoglobin C inheritance pattern?

Answer 14

Autosomal recessive

Question 15

Clinical manifestiation of hemoglobin C?

Answer 15

decrease solubility of hemoglobin - tends to lyse red blood cells

Question 16

Diagnosing sickle cell: DNA diagnosis

Answer 16

Mst II restriction enzyme normally cleaves 5’cctnagg-3’ but the sickle cell mutation changes the a to t and thus it is not recognized by the restriction enzyme - leading to a single heavier band in sickle

Question 17

How can we distinguish sickle cell from Hemoglobin C with DNA diagnosis?

Answer 17

Hemoglobin C mutation does not effect restriction site of MstII, so Mst II will still cleave the DNA and yield two bands

Question 18

Weight of Hb bands?

Answer 18

1. 15
2. 20
3. 35 if uncleaved

Question 19

Can we diagnosis hemoglobin diseases with electrophoresis

Answer 19

Yes, because HbS (sickle) has been changed from a negative charge (glutamate) to valine - so we would expect that it would not travel as far to the anode

HbC the glutamate has beeen changed to lysine (a positive charge) so we would expect it to travel the least distance

In order of migration, from farthest to shortest distance traveled we would expect
HbA
HbS
HbC

Question 20

Hemoglobin oxygenate form

Answer 20

The heme is planar when oxygen is bound - via Fe - which is also bound to His

Question 21

Hemoglobin deoxygenated form

Answer 21

The heme is domed (nonplanar/tense) when O2 is not bound

Question 22

Hemoglobin Kempsey

Answer 22

High oxygen affinity
Less O2 to tissues
Overproduction of blood cells leads to polycythemia

Question 23

Hemoglobin Kansas

Answer 23

Low oxygen affinity
Lower O2 level in RBC
Cyanosis

Question 24

Thalassemia are due to?

Answer 24

Globin level imbalance

Question 25

Alpha thalassemias are usually caused by ____ where as beta thalassemias are usually caused by _____

Answer 25

Alpha are usually caused by deletions Beta are usually caused by point mutations

Question 26

Alpha thalassemia alpha beta gamma

Answer 26

low or zero alpha | beta and gamma in excess and preicipitates

Question 27

alpha thalassemia onset?

Answer 27

both fetal and postnatal

Question 28

Alpha thalassemia usually caused by?

Answer 28

deletions in alpha globin genes

Question 29

Beta thalassemia alpha beta

Answer 29

low or zero beta globin | alpha globin in excess and precipitates

Question 30

Beta thalassemia onset?

Answer 30

Postnatal only

Question 31

What usually causes beta thalassemia?

Answer 31

Point mutations in the beta globin gene

Question 32

What less commonly causes beta thalassemias?

Answer 32

Deletions in the LCR or the Beta gene cluster

Question 33

If you have alpha thalassemia what do you have too much of, relatively speaking?

Answer 33

You have a lot more beta in adults and beta tends to form homotetramers You can also form gamma tetramers

Question 34

If you have beta thalassemia, what do you have too much of, relatively speaking?

Answer 34

You have a lot more alpha, which form alpha tetramers

Question 35

What happens when you have homotetramers in alpha or beta thalassemia?

Answer 35

Homotetramers do not carry oxygen well and tend to precipitate causing hemolysis

Question 36

Three primary consequences of thalassemia (globin level imbalance)

Answer 36

Hemolysis Microcytosis Hypochromia

Question 37

Microcytosis

Answer 37

Smaller RBCs because total protein is less

Question 38

Hypochromia

Answer 38

poor oxygen binding gives rise to paler color of RBC

Question 39

``` Alpha thalassemia Globin Level Phenotype alpha alpha / apha apha - alpha alpha / alpha ____ alpha alpha / ____ ____ alpha____/ alpha_____ alpha____ /____ ______ ___________________ ```

Answer 39

Globin level Symptoms 100% normal 75% silent 50% alpha thalassemia 1 trait 50% alpha thalassemia 2 trait 25% HbH 0% fetal death alpha thalassemia trait - mild anemia HbH4 (B4) severe anemia gamma 4 - fetal death = hydrops fetalis - not sufficient to carry infant to term

Question 40

When do alpha thalassemia start exerting effect?

Answer 40

early fetal development through adulthood, because... alpha is turned on in fetal development (5wk) and stays on throughout life - so if you have less alpha expression you effect fetus and adult

Question 41

Alpha-thalassemia - high risk groups

Answer 41

alpha-thal-1 allele (- -) Southeast Asia alpha thal-2 allel (alpha -) Africa/Meditteranean/Asia

Question 42

Dangers of alpha-thal-1

Answer 42

In alpha-thal-1 both copies of alpha are deleted on the same chromosome Thus, if two carriers get together and have a baby - and their fetus happened to receive both (- -) alleles --> hydrops fetalis (- - / - -) (gamma4)

Question 43

What would happen in an alpha-thal-1- allele (- -) and single deletion (alpha - ) came from a mating? Which population is at increased risk?

Answer 43

HbH disease (alpha - ) (- - ) (B4) Southeast Asia

Question 44

Even though both alpha thalassemia 1 and alpha thalassemia 2 have the same level of alpha expression (50%), why is alpha thalassemia 1 more worrisome?

Answer 44

Because the deletions are on the same chromosome In alpha thalassemia 2, the deletions are on opposite chromosomes, so there is no worry that there could be hydrops fatalis

Question 45

What is hemogoblin H?

Answer 45

When you have HbH (beta4)

Question 46

When do beta thalassemia start to exert their effect?

Answer 46

after birth - because beta globin genes not turned on until after birth

Question 47

How long does it take for beta thalassemias to show up

Answer 47

It takes a few months after birth because RBC turnover is 120 days, so you need around that much time to turnover the fetal gamma to beta

Question 48

Thalassemia major

Answer 48

severe enough you need treatment

Question 49

Thalassemia minor

Answer 49

mild symptoms or no symptoms

Question 50

Beta-o heterozygote -->

Answer 50

50% beta | Beta thalassemia minor

Question 51

Beta+ heterozgote -->

Answer 51

>50% because you mutated allele makes some | Beta thalassemia minor

Question 52

Beta-o homozygote -->

Answer 52

Beta thalassemia major

Question 53

Beta+ / Beta-o

Answer 53

Beta thalassemia major

Question 54

Beta thalassemia causing mutations can result from

Answer 54

Basically anything that can go wrong has gone wrong ``` Transcription splicing cap site rna cleavage frameshift nonsense unstable deletion ``` upstream downstream intron exon

Question 55

Complex beta thalassemias

Answer 55

Caused by larger chunk of genes deleted (beta cluster combo) or LCR

Question 56

LCR deletion -->

Answer 56

Hispanic thalassemia

Question 57

Hereditary persistance of fetal hemoglobin expression (HPFH)

Answer 57

Doesnt usually take out coding sequence in beta cluster but take out sequence further downstream that causes HPFH

Question 58

HPFH: mechanism (2) and implication

Answer 58

1. large deletion downstream may bring downstream enahncer closer to gamma locus that will keep expression elevated after birth 2. point mutation in gamma promoter where suppressor would bind may prevent suppression can manipulate to jack up expression gamma to help treat (sub) for beta thalassemia

Question 59

Inhibition of gamma repression

Answer 59

BCL11A is required to repress gamma - we could potentially target BCL11A via miRNA inhibition gene silencing

Question 60

Partial trisomy 13 and fetal hemoglobin

Answer 60

miRNA15a and 16-1 are on chromosome 13, people with partial trisomy 13 have fetal hemoglobin because these miRNA inhibit an inhibitor of gamma - leaving gamma active.