Hemoglobinopathies Flashcards
Hemoglobin alpha cluster
We have four copies of alpha both of which are preceded by fetal zeta (5’)
Chromosome 16
Hemoglobin beta cluster
epsilon Ggamma Agamma delta beta
How many copies of beta?
two total (one on each homolog 11)
Embryonic hemoglobin?
zeta2epsilon2
alpha2epsilon2
zeta2gamma2
fetal hemoglobin?
alpha2gamma2
adult hemoglobin?
95% alpha2beta2
3.5% alpha2delta2
Globin switching during development
alpha cluster?
5th week switch from zeta to alpha
Globin switching during development
beta cluster?
Switch from epsilon to gamma in-utero
Switch from gamma to beta at birth
Locus control region
10 kb upstream - Regulates globin transcription of beta cluster
Both timing and level
What happens if we delete LCR of beta cluster?
Hispanic epsilongammadeltabeta thalassemia –> have much lower expression of beta isoforms
Sickle Cell Disease
Single gene disease
Single point mutation at codon 6 in exon 1 of beta globin
Sickle cell disease
Amino acid substitution
Glutamine for valine
Hemaglobin C disease
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
Sickle cell and hemoglobin C inheritance pattern?
Autosomal recessive
Clinical manifestiation of hemoglobin C?
decrease solubility of hemoglobin - tends to lyse red blood cells
Diagnosing sickle cell: DNA diagnosis
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
How can we distinguish sickle cell from Hemoglobin C with DNA diagnosis?
Hemoglobin C mutation does not effect restriction site of MstII, so Mst II will still cleave the DNA and yield two bands
Weight of Hb bands?
- 15
- 20
- 35 if uncleaved
Can we diagnosis hemoglobin diseases with electrophoresis
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
Hemoglobin oxygenate form
The heme is planar when oxygen is bound - via Fe - which is also bound to His
Hemoglobin deoxygenated form
The heme is domed (nonplanar/tense) when O2 is not bound
Hemoglobin Kempsey
High oxygen affinity
Less O2 to tissues
Overproduction of blood cells leads to polycythemia
Hemoglobin Kansas
Low oxygen affinity
Lower O2 level in RBC
Cyanosis
Thalassemia are due to?
Globin level imbalance
Alpha thalassemias are usually caused by ____ where as beta thalassemias are usually caused by _____
Alpha are usually caused by deletions
Beta are usually caused by point mutations
Alpha thalassemia
alpha
beta
gamma
low or zero alpha
beta and gamma in excess and preicipitates
alpha thalassemia onset?
both fetal and postnatal
Alpha thalassemia usually caused by?
deletions in alpha globin genes
Beta thalassemia
alpha
beta
low or zero beta globin
alpha globin in excess and precipitates
Beta thalassemia onset?
Postnatal only
What usually causes beta thalassemia?
Point mutations in the beta globin gene
What less commonly causes beta thalassemias?
Deletions in the LCR or the Beta gene cluster
If you have alpha thalassemia what do you have too much of, relatively speaking?
You have a lot more beta in adults and beta tends to form homotetramers
You can also form gamma tetramers
If you have beta thalassemia, what do you have too much of, relatively speaking?
You have a lot more alpha, which form alpha tetramers
What happens when you have homotetramers in alpha or beta thalassemia?
Homotetramers do not carry oxygen well and tend to precipitate causing hemolysis
Three primary consequences of thalassemia (globin level imbalance)
Hemolysis
Microcytosis
Hypochromia
Microcytosis
Smaller RBCs because total protein is less
Hypochromia
poor oxygen binding gives rise to paler color of RBC
Alpha thalassemia Globin Level Phenotype alpha alpha / apha apha - alpha alpha / alpha \_\_\_\_ alpha alpha / \_\_\_\_ \_\_\_\_ alpha\_\_\_\_/ alpha\_\_\_\_\_ alpha\_\_\_\_ /\_\_\_\_ \_\_\_\_\_\_ \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_
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
When do alpha thalassemia start exerting effect?
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
Alpha-thalassemia - high risk groups
alpha-thal-1 allele (- -) Southeast Asia
alpha thal-2 allel (alpha -) Africa/Meditteranean/Asia
Dangers of alpha-thal-1
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)
What would happen in an alpha-thal-1- allele (- -) and single deletion (alpha - ) came from a mating?
Which population is at increased risk?
HbH disease
(alpha - ) (- - ) (B4)
Southeast Asia
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?
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
What is hemogoblin H?
When you have HbH (beta4)
When do beta thalassemia start to exert their effect?
after birth - because beta globin genes not turned on until after birth
How long does it take for beta thalassemias to show up
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
Thalassemia major
severe enough you need treatment
Thalassemia minor
mild symptoms or no symptoms
Beta-o heterozygote –>
50% beta
Beta thalassemia minor
Beta+ heterozgote –>
> 50% because you mutated allele makes some
Beta thalassemia minor
Beta-o homozygote –>
Beta thalassemia major
Beta+ / Beta-o
Beta thalassemia major
Beta thalassemia causing mutations can result from
Basically anything that can go wrong has gone wrong
Transcription splicing cap site rna cleavage frameshift nonsense unstable deletion
upstream
downstream
intron
exon
Complex beta thalassemias
Caused by larger chunk of genes deleted (beta cluster combo) or LCR
LCR deletion –>
Hispanic thalassemia
Hereditary persistance of fetal hemoglobin expression (HPFH)
Doesnt usually take out coding sequence in beta cluster but take out sequence further downstream that causes HPFH
HPFH: mechanism (2) and implication
- large deletion downstream may bring downstream enahncer closer to gamma locus that will keep expression elevated after birth
- 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
Inhibition of gamma repression
BCL11A is required to repress gamma - we could potentially target BCL11A via miRNA inhibition gene silencing
Partial trisomy 13 and fetal hemoglobin
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.
