20.03.07 AD - dominant negative effect

Question 1

What are dominant negative variants

Answer 1

• Variants that impair the function of the protein encoded by the mutated gene and also, due to protein-protein interactions, reduce the function of the protein encoded by the normal gene.
• Only seen in heterozygotes

Question 2

What has likely evolved to protect from dominant negative effects of abnormal truncated proteins

Answer 2

Nonsense-mediated decay.

Question 3

Which proteins are particularly susceptible to dominant negative effects

Answer 3

-Proteins with multimeric structures, where they are dependent on oligomerisation for activity.

Question 4

Review of non-syndromic sensorineural hearing loss

Answer 4

• Mutations in GJB2 (connexin 26) or GJB6 (connexin 30)
• Gap junction proteins expressed in human cochlea.
• 6 connexins oligomerise (can be homopolymeric or heteropolymeric) to form a hemi channel (connexon). Connexons align symmetrically to those of neighbouring cells to form gap junctions.
• Involved in potassium ion recycling in ear.

Question 5

Molecular genetics of non-syndromic sensorineural hearing loss (why AD and AR inheritance)

Answer 5

• Most GJB2 variants are recessive
• Dominant missense variants M34T or R75W, produce full length structurally abnormal Connexin 26 molecules. These form plaques with wildtype Cx26 and Cnx30 to form connexons with impaired permeability to potassium ions.

Question 6

Review of Osteogenesis imperfecta (OI)

Answer 6

• 90% patients have mutations in COL1A1 or COL1A2
• Encode type 1 procollagen chains.
• Fibrillar collagens are major structural proteins of connective tissue and consist of triple helices of homo or hetero-trimeric polypeptide chains.
• 3 preprocollagen chains associate and wind into a triple helix.

Question 7

Disease mechanism for Osteogenesis imperfecta

Answer 7

• Haploinsufficiency (OI type 1). Null variants in COL1A1. Decrease in amount of type 1 collagen. Mild phenotype
• Dominant negative (OI type 2,3,4). 80% mutations replace glycine residues in triple helical domain of COL1A1 or COL1A2. Produce abnormal molecules that disrupt triple helix formation. Severe disease. Most severe if mutations are close to C-terminus (where helix starts assembling)

Question 8

Review osteopetrosis

Answer 8

• Inherited metabolic bone disorder that results from ineffective osteoclast-mediated bone reabsorption.
• Mutations in CLCN7 gene.
• Chloride channel protein 7, regulates pH of osteoclasts.
• Has two c-terminal CBS domains thought to facilitate formation of channel dimers
• AR and AD inheritance

Question 9

Disease mechanism for osteopetrosis

Answer 9

• Autosomal recessive= Null variants lead to loss of chloride channel function
• Autosomal dominant= variants cause incomplete inactivation of chloride channel so reduced conductance. More variants causing AD form are in CBS-domains. Target for siRNA to silence affected allele. Less severe than AR form.

Question 10

Review myotonia congenita

Answer 10

• Most common genetic skeletal muscle ion channelopathy.
• Characterised by muscle stiffness and inability to contract after voluntary contraction.
• Mutations in CLCN1- encoding skeletal muscle voltage-gated chloride channel.
• Exists as a homodimer

Question 11

Disease mechanism for myotonia congenita

Answer 11

• Channel helps stabilise the cell’s electrical charge, to prevent muscles from contracting abnormally.
• Autosomal recessive= Loss of function through NMD (nonsense vars), impaired transport to membrane or inability to form dimers.
• Autosomal dominant= Inactive channel dimers contain WT and mutant CLCN1 protein. Less common and less severe than AR. Dominant variants are more common at dimer interface.

Question 12

Disease mechanism for TP53

Answer 12

• Functional inactivation of tumour suppressor p53 is a feature of human cancer.
• p53 is recruited to sites of DNA damage and acts as a transcription factor to change gene expression to promote apoptosis or cell cycle arrest.
• 74% of p53 mutations are missense and occuring in DNA-binding domain.
• p53 variants may cause cancer by exerting a dominant negative effect on wild-type p53. p53 dimers form a tetramer which binds to DNA. Mutant and WT tetramers have impaired DNA binding or transcriptional activity.

Question 13

Review marfan syndrome

Answer 13

• Connective tissue disorder where morbidity and mortality results from aortic aneurysm and dissection.
• Mutations in FBN1 gene
• Encodes fibrillin-1 which has a structural role in the wall of large arteries.

Question 14

Disease mechanism in marfan syndrome

Answer 14

• Haploinsufficient= Nonsense/ frameshift variants that lead to NMD. Decreased amounts of fibrillin-1 leads to thinner matrix in the vasculature, decreasing aortic wall strength. Increased risk of cardiovascular death than dominant negative variants. More responsive to Losartan therapy for inhibition of aortic root dilatation.
• Dominant negative (usually cysteine substitutions). Variants result in a stable but altered variant protein. Disturbed interaction between WT and mutant proteins, leading to disorganised extracellular matrix. Leads to more ectopia lentis.