NF2/Merlin and Neurofibromatosis 2

March 25, 2003By: Caroline Clendenin

Neurofibromatosis: An Overview

• NF is the most common neurological disorder caused by a single gene.

• It is subdivided into two distinct types: NF1 and NF2, which are caused by different genes on different chromosomes.

• NF1 is more common, affecting 1:4,000 births throughout the world; while NF2 affects about 1:40,000 people worldwide.

• These diseases affect more than 100,000 Americans—making NF more prevalent than Cystic Fibrosis, hereditary Muscular Dystrophy, Huntington’s Disease, and Tay Sachs combined.

Neurofibromatosis: An Overview:

• Both forms of NF are inherited by autosomal dominant mechanisms.

• NF can also arise from spontaneous mutations in sperm or eggs.

• In this presentation we will focus on NF2 and its role in the cell.

NF2: Signs and Symptoms • Bilateral tumors on the eighth cranial nerve• Problems with balance, secondary to dysfunction of the vestibular

nerve • Hearing loss and tinnitus, secondary to dysfunction of the auditory

nerve • Schwannomas—tumors arising from Schwann cells• Symptoms of schwannomas depend on location: those on cranial

nerves affect only the head and neck unless they grow and put pressure on the brainstem—which can be life threatening.

• Those on nerves as they exit the spinal cord may cause numbness in a region of the body or weakness and numbness in the legs if the tumor places pressure on the spine, itself.

• Peripheral schwannomas rarely cause neurological symptoms, but may be disfiguring.

• Facial weakness• Headache

NF2: Diagnosis• Bilateral tumors of eighth

cranial nerve OR• A parent, sibling, or child with

NF2 and a unilateral tumor on the eighth cranial nerve OR

• A parent, sibling or child with NF2 and two of the following:– Glioma– Meningioma– Neurofibroma– Schwannoma– Early onset cataract

Clinical Onset of NF2• The identification of the NF2 gene product allows

genetic testing for families with documented cases of NF2 in order to confirm diagnosis.

• Prenatal testing by amniocentesis and chorionic villi sampling is also available for those at risk.

• The mean age of onset is in the early 20’s.• Those who present earlier with clinical symptoms

tend to have more rapid, aggressive tumors with a greater number of tumors present.

• Those who present later exhibit a less severe disease course with fewer tumors.

Treatments:• At present, treatments

are aimed at controlling the symptoms.

• Diagnostic technologies such as MRI and CT can reveal tumors as small as a few millimeters in diameter, thus allowing early treatment.

• Surgical removal of the tumors—may result in hearing loss.

• Partial removal of tumors• Radiation therapy

(radiosurgery)• Watchful waiting—if

tumors are not progressing rapidly or if the patient has other serious medical issues.

The NF2 Gene:What is its normal function?

• This tumor suppressor gene was identified by positional cloning and loss of heterozygosity studies.

• The protein encoded by NF2 is called merlin (or schwannomin).

• It is a member of the ERM (ezrin, radixin, moesin) family of cytoskeleton:membrane linker proteins. Bretscher, et. al.

In what tissues is NF2 found?

• In human and fetal mouse tissues, it is found in:– Heart– Brain– Liver– Spleen– Skeletal muscle– Kidney

• In adult mice NF2 is NOT found in:– Spinal cord– Brain– Liver– Pancreas

What does it do?• NF2 gene product is

essential for extraembryonic development just prior to gastrulation.

• Homozygous mutant mouse embryos die between day 6.5 and day 7.

• They show collapsed extraembryonic region and an absence of organized extraembryonic ectoderm.

• Conditional homozygous NF2 knockout mice show characteristics of NF2 disorder.

Giovannini, et. al.

The NF2 Gene:Domains

• Three domains:1. FERM2. alpha helix3. C-terminus

• It is missing N-terminal actin binding domain that is present in all other ERM proteins.

Reed & Gutmann

Conformations of NF2

• “closed” vs. “open”• Binding sites for

interactions with CD44 and HRS (hepatocyte growth factor-regulated tyrosine kinase substrate)

• Proposed mechanism of interaction.

Gutmann

Uncertainties:

Although the molecular basis of merlin’s growth-suppressing activity is not known, the recent identification of merlin-interacting proteins, and advances in understanding the molecular function of related ERM proteins, suggest that merlin may function to control the localization of signaling molecules through its effects on cytoskeletal reorganization.

Model of Activation of ERM Proteins and Merlin

• For ERM proteins, phosphorylation of c-terminal residue mediated by the Rho pathway induces transition from inactive oligomers to active monomers.

• For merlin, the Rac pathway mediates phosphorylation of a c-terminal residue which causes inactivation of its tumor suppressor activity.

• Dephosphorylation of both ERM and merlin allows interdomain association to occur.

Gautreau, et. al.

How does merlin mess up?

• As a tumor suppressor, merlin is affected by Knudson’s Two Hit Hypothesis. Both copies of the gene must be mutated to remove its function.

• Generally, mutations in NF2 involve deletions that cause the formation of truncated proteins.

• As with Rb, a person with a mutated copy of the NF2 gene (NF2+/NF2-) will almost invariably develop acoustic neuromas associated with neurofibromatosis 2.

• The NF2 mutation is about 98% penetrant.

Merlin Mutations in Mice

• Mutant mice exhibited formation of metastatic osteosarcomas, whereas in humans tumor growth is usually benign schwannomas which are localized.

• This difference is probably due to the fact that that p53 is highly linked to the NF2 gene in mice, but not in humans.

McClatchey, et. al.

The Future of NF2:It is hoped that through these studies of molecular genetics that nonsurgical or pharmacologic treatments can be employed to slow and suppress the growth of tumors associated with the neurofibromatoses.

Summary:• NF2 is a tumor suppressor gene that encodes the

protein merlin.• Merlin is a member of the ERM family of

membrane:cytoskeletal linker proteins.• It requires “two hits” to eliminate its function on the

cellular level, although the NF2 disorder is inherited in an autosomal dominant fashion, similar to Rb.

• Mice have been used to study this protein’s function.

• Although the specific pathway for this gene is not fully understood, it is presumed that it acts on cytoskeletal reorganization to cause tumors.