6 - human genetics

Question 1

how do nucleotide repeats induce disease?

Answer 1

nucleotide repeats are very unstable

the repeats impair the structure of the protein → loss of function

there is no correlation between number of repeats and onset of disease

Question 2

how can positional cloning be used to identify human disease-associated genes?

Answer 2

positional cloning is a laboratory technique used to locate the position of a disease-associated gene on a chromosome

the gene of interest is mapped very precisely

used for SINGLE-GENE DISORDERS

Question 3

how do multifactorial gene disorders arise

Answer 3

variants often benign on their own

when multiple variants exist, they can combine and become deleterious, resulting in disease

Question 4

how do you identify genes associated with multifactorial disorders?

Answer 4

GENOME WIDE ASSOCIATION STUDIES

observational study of a genome-wide set of genetic variants across different individuals

identify variants associated with a particular trait/disease

Question 5

what is aneuploidy

Answer 5

an organism or cell having a chromosome number that is not an exact multiple of the haploid

Question 6

list the structural abnormalities that result in chromosomal disorders

Answer 6

inversions
insertions
deletions
duplications
translocations

Question 7

T or F

mutations in different genes always have different phenotypic effects

Answer 7

FALSE

mutations in different genes can have the same phenotypic effects

Question 8

what are the consequences of missing an X chromosome in a germ cell VS a somatic cell

Answer 8

GERM CELL:
loss of X-chromosome is inherited by all cells
leads to Turner syndrome (short stature, delayed puberty, learning disabilities, heart defects)

SOMATIC CELL:
loss of X-chromosome leads to mosaic
complementation from normal cells means symptoms of Turner syndrome aren’t as penetrant

Question 9

how do chromosomal translocations create gene fusion products?

use the philadelphia chromosome as an example

Answer 9

breakpoints at:
– bcr (housekeeping gene) on chromosome 22
– c-abl (tyrosine kinase which regulates cell-cycle proteins) on chromosome 9

start (amino terminus) of bcr fuse to end (carboxyl terminus) of c-abl to create gene fusion product

this particular gene fusion product is called the Philadelphia chromosome → very common in leukaemia

start of bcr gene contains promoter → gene product regulated by bcr ∴ is constantly expressed in white blood cells

gene product also has activity of c-abl → cells constantly proliferate → cancer

Question 10

how do chromosomal translocations create insertions?

use berkitts lymphoma as an example

Answer 10

berkitts lymphoma occurs between chromosome 8 and chromosomes 2, 14 or 22 (most commonly 8 and 14)

breakpoints between 8 and 14 result in the c-myc gene inserting onto the end of chromosome 14

c-myc becomes under control of Ig promoter and enhancers = constant expression

Ig gene produces antibodies:
– expressed heavily by white blood cells
– very important for cell cycle
– constant expression = constant proliferation

Question 11

what is the difference between tumour suppressor genes and oncogenes
give examples for both

Answer 11

tumour suppressor genes = when mutated, fail to suppress pathways for tumour development
e.g. p53, BRCA1

oncogenes = when mutated, actively promote pathways for tumour development

e. g. RAS, MYC
* note: non-mutated oncogene = proto-oncogene

Question 12

what happens when cancer genes are mutated in germ-line cells?

Answer 12

when cancer genes* are mutated in germ-line, there is a predisposition to develop cancer

• mutation is most often loss of function in tumour suppressor gene

two-hit hypothesis:
cancer develops only if a second mutation in somatic cells knocks out the function of the remaining wild-type allele

Question 13

in gleevac effective in treating leukaemia

Answer 13

gleevac blocks the active site on the abl protein in the bcr-abl complex

abl is now unable to phosphorylate/activate proteins

∴ no production of myeloid leukaemia

Question 14

are B-raf inhibitors effective in treating leukaemia

Answer 14

no

B-raf inhibitors do prevent cell proliferation but only act on a single pathway

∴ tumour able to develop using a different pathway

effective drugs target multiple pathways

Question 15

how can mutation databases be used to identify cancer-associated genes?

Answer 15

within individual cancer there are often mutation in the same pathway (similar genes affected)

COSMIC databases collates mutations to see which genes / what types of mutations are commonly effected in different cancers

can learn how different types of cancers function

Question 16

what is bioinformatic analysis used for?

Answer 16

bioinformatics is the study of genetic and other biological information using computer and statistical techniques

used to analyse:
– sequence mutations
– sequence motifs found in other disease-associated genes
– gene co-expression
– transcriptional networks
– functional genomics

Question 17

how can bioinformatic predictions be validated?

Answer 17

use protein or RNA analysis to determine when and where a gene is expressed

1) use antibodies to stain for particular proteins/RNA OR add GFP to gene sequence at carboxy or amino terminal (depends on structure). these methods allow one to characterise exact location and distribution of proteins throughout cell
2) insert strong promoter into gene of interest so it is overexpressed. observe phenotype or monitor RNA expression to discern effects
3) use RNA interference to block gene expression (RNA cleavage or transcript degradation). observe phenotype when gene doesn’t function to discern effects

Question 18

describe process of somatic gene therapy

Answer 18

bone marrow stem cells are taken

a retroviral vector is prepared containing WT copy of mutant gene

retrovirus infects stem cells —> WT incoorperated into host genome

still have copy of defective gene, but now have copy of functional gene

cells are grown and expanded in lab until sufficient quantity is reached

cells injected back into patient

Question 19

describe process of germline gene therapy

Answer 19

guideRNA targets enzyme to particular location in genome

creates ds break in DNA at location

non-homologous repair often results in small insertion or deletion —> random mutation

mutation used to knock-out target gene

can also use donor DNA to insert at location i.e. a transgene or WT copy

must be inserted at very particular time in development (too late, result in mosaic)

Question 20

pros and cons of germline vs somatic gene therapy

Answer 20

GERMLINE
pros:
• corrects every cell in body
• no mutant allele present if successful
cons: 
• ethical issues re. genetic engineering
• very expensive
• doubts on effectiveness

SOMATIC:
pros:
• can treat patients at any age
cons:
• mutant allele remains in some cells
• may not recover full physiological function
• chance of death