X-Linked Disorders

Question 1

The HGP sequenced and identified about __ genes in each human

Answer 1

The HGP sequenced and identified about 20,500 genes in each human

Question 2

Each chromosome comes in __, inherited from each __

Answer 2

Each chromosome comes in pairs, inherited from each parent

Question 3

What is the essential first step in determining inheritance patterns ?

Answer 3

Investigate pedigree

Question 4

Define pedigree

Answer 4

• pictorial representation of a family medical history
• tracks inheritance pattern of disease

Question 5

A pedigree includes

Answer 5

• entire maternal and paternal lineage
• minimum of 3 generations
• illness, defects, conditions of each family member
• age (current, at diagnosis, at death)
• miscarriages/ stillbirths
• adoptions
• ethnicity

Question 6

Diffentiate genotype vs phenotype

Answer 6

Genotype:
- actual DNA sequence at a specific locus

Phenotype:
- physical manifestations of genotype (hair color, susceptibility to a disease)

Question 7

Allele vs wild-type vs variant

Answer 7

Allele:
- different forms of a gene

Wild-type:
- most common allele in a population

Variant:
- permanent alteration in a DNA sequence (mutation)

Question 8

List specific info in family history that may indicate X-linked genetic disorders

Answer 8

1. Multiple affected males in the maternal side
2. Especially neonatal/ child deaths
3. Mildly affected females (sisters mothers, maternal aunts)
4. No known risk factors

Question 9

X-linked recessive inheritance

Answer 9

• higher expression in males > females
• heterozygous females usually do not have phenotype
• X-linked disorders inherited from father to all daughters
• never transmitted from father to sons
• affected males within same family always related through females
• significant proportion due to new/ de novo variants in a gene on X-chromosome

Question 10

Characteristics of X-linked dominant inheritance

Answer 10

• more commonly expressed in females > males
• BUT females have milder phenotypic expressions of genetic disease
• affected males will have normal sons and affected daughters
• affected female have 50% risk of having children with genetic disease

Question 11

What is XIST ?

Answer 11

X-inactive specific transcript:
- non-coding untranslated RNA
- major effector of X-inactivation process
- causes chromatin condensation and inactivation = Barr body
- epigenetics change = changes gene but does not involve a base change

Question 12

Purpose of X-chromosome inactivation in females

Answer 12

• compensates for dosage of X-linked genes in females (XX) vs males (XY) = both males and females only have one active X-chromosome
• X-chromosome with variant is always silenced
• cells that transcribe normal allele compensates enough gene products for deficient variant cells
• deficient cells divide less efficiently, and are eventually overgrown by normal cells

Question 13

T or F: Random X-chromosome inactivation is a normal, expected process

Answer 13

TRUE; Random X-chromosome inactivation is a normal, expected process

Question 14

How does DNA methylation of X alleles evaluate X-chromosome Inactivation ?

Answer 14

• Human androgen receptor assay (HUMARA)
• Targets polymorphic short tandem repeat of the Xq-
  linked androgen receptor (AR) gene
• Methylation status of the AR on inactive X
  chromosome correlates to the whole X chromosome
  inactivation

Since Paternal X and maternal X have 50% probability of being
methylated and inactivated
- 1:1 ratio for X chromosome inactivation is expected
- deviations from this theoretical ratio = skewed X inactivation

Question 15

Describe Methylation-specific PCR

Answer 15

Two-step approach:

1. PCR with primers specific for methylated vs unmethylated DNA
2. Chemical modification of DNA with sodium bisulfite
   - treatment converts unmethylated cytosines into uracil = amplified as Thymine

Question 16

How does Expressed Polymorphisms of X alleles evaluate X-chromosome Inactivation ?

Answer 16

• not often used
• not clinically useful for blood

Question 17

Why are males hemizygous ?

Answer 17

Males have a single X chromosome

Question 18

Phenotype of MECP2 disorder

Answer 18

Progressive neurodevelopmental disorder:
- First year is normal, followed by rapid regression of development
- Repetitive, stereotypic hand movements
- Fits of screaming and inconsolable crying
- Seizures
- Acquired microcephaly

Question 19

Describe MECP2 gene and describe pathogenesis of disorder

Answer 19

“Methyl-CpG binding protein 2 gene:”
- located on Xq28
- encodes Chromatin-associated protein that activates/ represses transcription

• neurons cannot mature in MECP2 disorders

Question 20

How are MECP2 disorders diagnosed ?

Answer 20

• Sequencing and deletion/duplication analysis of MECP2
• 99% are de novo (a single occurrence in a family)

Question 21

Describe DMD gene and inheritance pattern

Answer 21

• Located on Xp21.2 - Xp.21.1
• encodes for Dystrophin protein (large muscle protein)
• X-linked recessive: Hemizygous; MOSTLY IN MEN or heterozygous pathogenic variants (in females) result in dystrophinopathies

Question 22

Describe DMD phenotype

Answer 22

Duchene Muscular Dystrophy:
- Delayed motor capabilities; waddling gait and difficulty doing active moments
- Wheelchair dependent by 12 years of age
- Few survive beyond 30
- Respiratory complications and progressive cardiomyopathy being common causes of death

Question 23

Describe Becker Muscular Dystrophy phenotype

Answer 23

• Later-onset skeletal muscle weakness
• Mean age of death = mid-40s
• Heart failure due to cardiomyopathy is the most common cause of death

Question 24

Diagnosis of DMD

Answer 24

• characteristic clinical findings
• elevated [CK]
• ID of a hemizygous pathogenic variant in DMD gene

Question 25

Describe ABCD1 gene and pathogenesis of disorder

Answer 25

- Located on **Xq28** - Encodes **ATP-binding cassette subfamily D member 1** - X-linked recessive: **Hemizygous** or **heterozygous** (females) with variant ABCD1 results in X-linked **adrenoleukodystrophy** - **Failure to transport fatty acids into peroxisome** results in **accumulation of long-chain fatty acids** - Affects the nervous system, adrenal cortex, and Leydig cells of the testes

Question 26

Describe purpose of ATP-binding cassette subfamily D member 1 protein ?

Answer 26

- Member of the **ATP-binding cassette (ABC) protein transporter** family - **Transports certain fatty acids into peroxisome**

Question 27

X-linked adrenoleukodystrophy in females

Answer 27

- **Adrenal function usually normal** - More than 20% of female (heterozygous) carriers develop **mild-to-moderate spastic paraparesis** in middle age or later Paraparesis: inability to move legs

Question 28

X-linked adrenoleukodystrophy in males

Answer 28

- Impaired adrenocortical function - Manifests between **4 to 8 years** of age - Initially resembles attention-deficit disorder/ hyperactivity and **progressively leads to cerebral disability**

Question 29

Diagnosis of X-linked adrenoleukodystrophy

Answer 29

- Suggestive clinical findings - **Elevated very long chain fatty acids (VLCFA)** - **Abnormal brain MRI** in boys - ID heterozygous ABCD1 pathogenic variant in girls

Question 30

Describe F8 gene and inheritance pattern of disorder

Answer 30

- Located on **Xq28** - Encodes **coagulation factor VIII** - X-linked recessive: **Hemizygous** or **heterozygous** (female) pathogenic variants in F8 cause **hemophilia A**

Question 31

What is FVIII ?

Answer 31

- Large plasma glycoprotein - Cofactor in blood coagulation cascade; activates FX - Binds von Willebrand factor during transport in blood

Question 32

Describe Hemophilia A phenotype in males

Answer 32

- Delayed or recurrent bleeding after injury Severe hemophilia A: - During first 2 years of age - Spontaneous joint bleeds or deep-muscle hematomas - Prolonged bleeding & excessive pain from minor injuries Mild hemophilia A: - Diagnosed later in life - Prolonged bleeding occurs from surgery or tooth extractions

Question 33

Hemophilia A in females

Answer 33

~ 30% of **heterozygous females** have **reduced FVIII clotting activity** and are at risk for bleeding - **Prolonged bleeding after major injuries**

Question 34

Describe the FMR1 gene and pathogenesis for disorder

Answer 34

- Located on **Xq27.3** - Encodes **Fragile X messenger ribonucleoprotein 1;** - a selective **RNA binding protein** for polyribosomes in translation (regulation and mRNA stability) - central role in neuronal development - Expanded **CCG repeats (>200)** = **excessive methylation of cytosines in promoter** of FMR1 gene = **failure to produce protein**

Question 35

Describe Fragile X syndrome phenotype

Answer 35

- **Most common heritable form of intellectual disability** - Neurodevelopment disorders - Seizures - Sleep disorders - Scoliosis Penetrance in females = 50-60% range NOTE: increases within generations

Question 36

Describe SLC6A8 gene and inheritance pattern for disorder

Answer 36

- Located on **Xq28** - Encodes **Solute carrier family 6 member 8** - **13 exons** (8.5 kb of genomic DNA) - X-linked recessive: **Hemizygous** or **heterozygous** (female) pathogenic variants in SLC6A8 = **creatine transporter deficiency**

Question 37

Describe SLC6A8 phenotype

Answer 37

- creatine transporter deficiency - Epilepsy - Developmental delay - Hyperactivity

Question 38

SLC6A8 variant diagnosis

Answer 38

- Heterozygous missense variant (c.1067G>T ) = (p.Gly356Val) in SLC6A8 - 50% of normal creatine uptake in fibroblasts - Chromosome analysis 46, XX - No skewed X-inactivation in peripheral blood

Question 39

What is SLC6A8 protein?

Answer 39

Solute carrier family 6 member 8: - **NaCl dependent** - **Transports creatine** in/ out of cells - For high energy requiring organs (brain and muscle retina)