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Explain glucokinase gene mutations causing early onset type 2 diabetes in some subjects

(MODY) but chronic hypoglycemia in other subjects.(15 marks)

Glucokinase GK) is an

enzyme that phosphorylates glucose to glucose 6-phosphate (G6P) with a high S0.5

of 8mM. (2 marks)

This very low affinity for glucose and lack of inhibition by

the reaction product, G6P, results in the ability to sense glucose

concentration within a cell, because the concentration of G6P formed is simply

proportional to blood glucose concentration in the presence of the GLUT2

glucose transporter. (3 marks)

In the beta cell of the pancreas, this biochemistry is

linked directly to insulin secretion. (1 mark)

In the liver it controls glycogen synthesis (because G6P

activates glycogen synthase even in the absence of insulin). (2 marks)

A mutation in the GK

gene which increases the S0.5 or lowers the Vmax of the enzyme results in

diabetes, because the threshold for glucose sensing is raised. (2 marks).

A mutation which lowers the Km and or increases the Vmax of

the enzyme causes chronic hypoglycemia. (2 marks)

More than 200 such SNP mutations have been identified in

man, accounting for a significant percentage of MODY. )1 mark)

Mutation is rare in adult forms of Type 2 diabetes.

Epigenetic control of GK expression may be more important in adult cases of

Type2 diabetes. (2 marks)

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What evidence supports the view that Type 2 Diabetes

(T2D) has a genetic basis? (7 marks).

Why is DNMT3, but not DNMT1 considered to be of significance in the proposed

epigenetic etioligy of T2D (17 marks).

T2D clearly runs in families, 40% increased lifetime

risk of developing the disease if one parent has T2D and a 70% increased risk

if 2 parents are T2D (3 marks).

Concordance in identical twins is higher than

in non-identical twins (2 marks)

identical twins have an identical

genome, but non-identical twins have similar ones, and carrying

those identical genes underlies the increased risk (2 marks).

DNMT1 can methylate only hemi-methylated DNA

DNMT3

can methylate both unmethylated DNA and hemi-methylated DNA (3 marks).

DNMT3 can be involved in establishing new patterns of DNA

methylation in genes

DNMT1 can only copy an established DNA methylation

pattern (2 marks).

Increased DNA methylation of specific areas of genes

mostly promoters is associated with gene-silencing (2

marks),

Degree of methylation in the DNA alters the battery of genes

expressed and therefore the cell type or efficacy in a particular tissue or

specifically switching on or off a specific gene within a cell type (2 marks).

Important processes during fetal development when the cell lineages,

tissues and organ structure are becoming established (2 marks).

Strong relationship between indices of fetal growth, such as birth weight,

and later onset of adult diseases such as T2D (2 marks),

Changes in the DNA methylation pattern in the developing fetus may render it

more or less susceptible to adult onset diseases (2 marks). which is fetal programming of adult disease and is clearly established in the case of

T2D (2 marks).

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The incidence of Tay-Sachs disease in Ashkenazi Jews is approximately 1 / 2,000.

A. What is the estimated frequency of heterozygotes?

q2 = 1 / 2,000

square root of q = 1 / 45

p = 1-1/45 = 44/45

p = 44 / 45

2pq = 2 x 44/45 x 1/45

2pq = 88 / 2,025 ≈ 1 / 23

How was the

karyogram obtained?

Giemsa staining after arrest in metaphase using colcemid.

Describe the

translocation, as precise

as possible, using

standard nomenclature

Translocation between chromosomes 10 and 15.

Nomenclature: 46,XX,t(10q:15q)

More precisely: 46,XX,t(10:15)(q26:q26)

Describe a strategy to analyse the target cells in various individuals from the

family. (Spinal muscular atrophy)

Spinal motoneurons from the probands are not accessible directly. They

can be obtained using induced pluripotent stem cells: fibroblasts are

extracted from a skin sample of each proband, reprogrammed to

pluripotency, and differentiated into motoneurons with a defined protocol.

Compare figures 2a, 2b, 2c. Can plastin 3 play a role in the development of SMA?

What can you learn about the expression patterns of PLS3? Discuss a strategy for

pharmacological intervention.

The expression of PLS3 is regulated in a cell-type specific manner. As

suspected, plastin 3 could play a role in the development of SMA as a

modifier: it is strongly expressed in motoneurons from probands without

SMA inspite of SMN1 mutations but weakly in their siblings with SMA.

Timely augmentation of PLS3 expression may attenuate the disease.

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Describe the preparation of the karyotype.

Cells are grown in culture and treated with colcemid or colchicin to disrupt the spindle

formation and arrest cell cycle. Cell are detached with trypsin if they are adherent cells,

fixative is added and aliquots are transferred on a slide. Chromosomes are treated with

trypsin for denaturation and stained with Giemsa to obtain a dark and light banding pattern.

Chromosomes are studied with a light microscope and sorted with a computer software.

Would you expect the mother to show a disorder based on the karyotype? Why?

The mother has a balanced translocation, i.e. an exchange of chromosomal material

between chromosomes 1 and 22, therefore the total amout of DNA is unchanged and the

karyotype is probably not associated with disease.

What is the most probable type of chromosomal aberration in the daughter?

Balanced translocations in a parent can lead to offspring with unbalanced translocations.

Each chromosome segregates independently, i.e. the altered chromosomes 1 and 22 might

not segregate together.

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Difference between genome and exome?

Genome is all the bases of the cells DNA

Exome is a subgroup of the genome only involved in expression as mRNA

What happens to cells with DNMT3A?

DNMT3A is a de-novo DNA methylator

It methylates DNA that hasn't been methylated before

Participates in epignetic processes with cells

Decreasing DNMT3A would reduce DNA methylation in regions of DNA that can be epigentically modified.

Glucokinase

converts glucose into glucose-6-phosphate

dependent on ATP

is mostly expressed in the liver

only responds to high blood glucose concentrations

expressed less in the pancreas part of glucose senor

Glucokinase is a key enzyme for uptake of glucose and glycogen synthesis in the liver

Liver GK makes sure more glucose is converted to glucose-6-phosphate when glycemia increases

increased glucose-6-phosphate-increases glycogen synthesis-more glucose uptake.

Mutation Glucokinase

causes MODY because it is needed for glucose uptake and insulin release

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