Oculocutaneous Albinism in Griscelli syndrome via the MYO5A Gene
- Summary and Pricing
- Clinical Features and Genetics
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The great majority of tests are completed within 20 days.
Predicting clinical sensitivity for the MYO5A gene is challenging due to genetic heterogeneity of Oculocutaneous albinism and the limited number of cases reported with MYO5A-associated Griscelli syndrome. Gross deletions and duplications will not be detected by sequencing.
Oculocutaneous albinism (OCA) is an inherited disorder caused by deficiency in melanin synthesis that results in hypopigmentation of the skin, eyes, and hair. If the phenotype is mainly restricted to the eyes and the optic system, it is referred to as ocular albinism (OA) (Gargiulo et al. 2011). The reduction or complete absence of melanin pigment in the developing eye leads to foveal hypoplasia and misrouting of the optic nerves in the affected individuals (Oetting and King 1999). The eye and optic system abnormalities that are common to all types of albinism are nystagmus, photophobia, strabismus, moderate to severe impairment of visual acuity, reduced iris pigment with iris translucency, reduced retinal pigment with visualization of the choroidal blood vessels on ophthalmoscopic examination, refractive errors and altered visual evoked potentials (VEP). The degree of skin and hair hypopigmentation varies with the type of OCA, but the ocular phenotype does not change (Lewis 2012). To date, four types of non-syndromic OCA (type I-IV, based on gene involved) have been described. Their prevalence varies among different populations (Lewis 2013).
Clinical findings of hypopigmentation of the skin and hair, in addition to the characteristic ocular symptoms are present in many syndromic disorders for e.g., Griscelli syndrome (GS) (Ménasché et al. 2000; Pastural et al. 1997). MYO5A-associated GS is described as characteristic albinism with a severe developmental delay and mental retardation that occurs early in life (Ménasché et al. 2000).
Mutations in MYO5A gene, which encodes an actin-based molecular motor protein Myosin Va (MyoVa) are associated with recessive GS (Pastural et al. 1997). It has been reported that the MYO5A encoded protein has a role in neuron function (Miyata et al. 2011). Myosin Va is recruited on to diverse organelles, such as melanosomes and secretory vesicles and is involved in short-range axonal/dendritic transport (Langford 2002; Brown et al. 2004), which explains the developmental delay in the GS patients. Very few causative mutations (nonsense and gross insertions and deletions) with limited cases have been reported in this gene (Human Gene Mutation Database).
For this NextGen test, the full coding regions plus ~10 bp of non-coding DNA flanking each exon are sequenced for the gene listed below. Sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization kit, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for any regions not captured or with insufficient number of sequence reads. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.
Indications for Test
All patients with symptoms suggestive of Oculocutaneous albinism with developmental delay are candidates.
|Official Gene Symbol||OMIM ID|
- Genetic Counselor Team - firstname.lastname@example.org
- Madhulatha Pantrangi, PhD - email@example.com
- Brown JR, Stafford P, Langford GM. 2004. Short-range axonal/dendritic transport by myosin-V: A model for vesicle delivery to the synapse. J. Neurobiol. 58: 175–188. PubMed ID: 14704950
- Gargiulo A, Testa F, Rossi S, Iorio V Di, Fecarotta S, Berardinis T de, Iovine A, Magli A, Signorini S, Fazzi E. 2011. Molecular and clinical characterization of albinism in a large cohort of Italian patients. Investigative Ophthalmology & Visual Science 52: 1281–1289. PubMed ID: 20861488
- Human Gene Mutation Database (Bio-base).
- Langford GM. 2002. Myosin-V, a versatile motor for short-range vesicle transport. Traffic 3: 859–865. PubMed ID: 12453149
- Lewis RA. 2012. Oculocutaneous Albinism Type 2. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301410
- Lewis RA. 2013. Oculocutaneous Albinism Type 1. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301345
- Ménasché G, Pastural E, Feldmann J, Certain S, Ersoy F, Dupuis S, Wulffraat N, Bianchi D, Fischer A, Deist F Le, Saint Basile G de. 2000. Mutations in RAB27A cause Griscelli syndrome associated with haemophagocytic syndrome. Nat. Genet. 25: 173–176. PubMed ID: 10835631
- Miyata M, Kishimoto Y, Tanaka M, Hashimoto K, Hirashima N, Murata Y, Kano M, Takagishi Y. 2011. A Role for Myosin Va in Cerebellar Plasticity and Motor Learning: A Possible Mechanism Underlying Neurological Disorder in Myosin Va Disease. Journal of Neuroscience 31: 6067–6078. PubMed ID: 21508232
- Oetting WS, King RA. 1999. Molecular basis of albinism: mutations and polymorphisms of pigmentation genes associated with albinism. Hum. Mutat. 13: 99–115. PubMed ID: 10094567
- Pastural E, Barrat FJ, Dufourcq-Lagelouse R, Certain S, Sanal O, Jabado N, Seger R, Griscelli C, Fischer A, Saint Basile G de. 1997. Griscelli disease maps to chromosome 15q21 and is associated with mutations in the myosin-Va gene. Nat. Genet. 16: 289–292. PubMed ID: 9207796
NextGen Sequencing using PG-Select Capture Probes
We use a combination of Next Generation Sequencing (NGS) and Sanger sequencing technologies to cover the full coding regions of the listed genes plus ~20 bases of non-coding DNA flanking each exon. As required, genomic DNA is extracted from the patient specimen. For NGS, patient DNA corresponding to these regions is captured using an optimized set of DNA hybridization probes. Captured DNA is sequenced using Illumina’s Reversible Dye Terminator (RDT) platform (Illumina, San Diego, CA, USA). Regions with insufficient coverage by NGS are covered by Sanger sequencing. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.
For Sanger sequencing, Polymerase Chain Reaction (PCR) is used to amplify targeted regions. After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.
Patient DNA sequence is aligned to the genomic reference sequence for the indicated gene region(s). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories, listed below, per ACMG Guidelines (Richards et al. 2015).
(1) Pathogenic Variants
(2) Likely Pathogenic Variants
(3) Variants of Uncertain Significance
(4) Likely Benign Variants
(5) Benign, Common Variants
Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (http://www.hgvs.org). Rare variants and undocumented variants are nearly always classified as likely benign if there is no indication that they alter protein sequence or disrupt splicing.
As of March 2016, 6.36 Mb of sequence (83 genes, 1557 exons) generated in our lab was compared between Sanger and NextGen methodologies. We detected no differences between the two methods. The comparison involved 6400 total sequence variants (differences from the reference sequences). Of these, 6144 were nucleotide substitutions and 256 were insertions or deletions. About 65% of the variants were heterozygous and 35% homozygous. The insertions and deletions ranged in length from 1 to over 100 nucleotides.
In silico validation of insertions and deletions in 20 replicates of 5 genes was also performed. The validation included insertions and deletions of lengths between 1 and 100 nucleotides. Insertions tested in silico: 2200 between 1 and 5 nucleotides, 625 between 6 and 10 nucleotides, 29 between 11 and 20 nucleotides, 25 between 21 and 49 nucleotides, and 23 at or greater than 50 nucleotides, with the largest at 98 nucleotides. All insertions were detected. Deletions tested in silico: 1813 between 1 and 5 nucleotides, 97 between 6 and 10 nucleotides, 32 between 11 and 20 nucleotides, 20 between 21 and 49 nucleotides, and 39 at or greater than 50 nucleotides, with the largest at 96 nucleotides. All deletions less than 50 nucleotides in length were detected, 13 greater than 50 nucleotides in length were missed. Our standard NextGen sequence variant calling algorithms are generally not capable of detecting insertions (duplications) or heterozygous deletions greater than 100 nucleotides. Large homozygous deletions appear to be detectable.
Interpretation of the test results is limited by the information that is currently available. Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.
When Sanger sequencing does not reveal any difference from the reference sequence, or when a sequence variant is homozygous, we cannot be certain that we were able to detect both patient alleles. Occasionally, a patient may carry an allele which does not amplify, due to a large deletion or insertion. In these cases, the report will contain no information about the second allele. Our Sanger and NGS Sequencing tests are generally not capable of detecting Copy Number Variants (CNVs).
We sequence all coding exons for each given transcript, plus ~20 bp of flanking non-coding DNA for each exon. Test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions or any currently uncharacterized alternative exons.
In most cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.
Our ability to detect minor sequence variants due to somatic mosaicism is limited. Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.
Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR.
Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes from whole blood). Test reports contain no information about the DNA sequence in other cell-types.
We cannot be certain that the reference sequences are correct.
Rare, low probability interpretations of sequencing results, such as for example the occurrence of de novo mutations in recessive disorders, are generally not included in the reports.
We have confidence in our ability to track a specimen once it has been received by PreventionGenetics. However, we take no responsibility for any specimen labeling errors that occur before the sample arrives at PreventionGenetics.
myPrevent - Online Ordering
- The test can be added to your online orders in the Summary and Pricing section.
- Once the test has been added log in to myPrevent to fill out an online requisition form.
- A completed requisition form must accompany all specimens.
- Billing information along with specimen and shipping instructions are within the requisition form.
- All testing must be ordered by a qualified healthcare provider.
(Delivery accepted Monday - Saturday)
- Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
- For small babies, we require a minimum of 1 ml of blood.
- Only one blood tube is required for multiple tests.
- Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
- During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
- In cold weather, include an unfrozen ice pack in the shipping container as insulation.
- At room temperature, blood specimen is stable for up to 48 hours.
- If refrigerated, blood specimen is stable for up to one week.
- Label the tube with the patient name, date of birth and/or ID number.
(Delivery accepted Monday - Saturday)
- Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
- For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
- DNA may be shipped at room temperature.
- Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
- We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.
(Delivery preferred Monday - Thursday)
- PreventionGenetics should be notified in advance of arrival of a cell culture.
- Culture and send at least two T25 flasks of confluent cells.
- Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
- Send specimens in insulated, shatterproof container overnight.
- Cell cultures may be shipped at room temperature or refrigerated.
- Label the flasks with the patient name, date of birth, and/or ID number.
- We strongly recommend maintaining a local back-up culture. We do not culture cells.