Miller Syndrome via the DHODH Gene

Clinical Features and Genetics

Clinical Features

Miller syndrome (OMIM# 263750, also called postaxial acrofacial dystosis, Genee-Wiedemann syndrome, and Wildervanck-Smith syndrome) is characterized by dysmorphic craniofacial features with postaxial limb deformities (Donnai, D. et al. J Med Genet 24(7):422-425, 1987). The most common features include severe micrognathia, cleft lip and/or palate, absence of fifth fingers and toes, syndactyly, abnormal bones in the forearms and lower legs, coloboma of the eyelids, and supernumerary nipples. Other dysmorphic features include downward slanting palpebral fissures, malar hypoplasia, malformed ears and a broad nasal ridge. The less common features include abnormalities of the heart, kidneys, genitalia, or gastrointestinal tract.  Patients may show delayed speech development due to hearing impairment, but usually their intelligence is normal. Other syndromes such as Pierre-Robin, Treacher-Collins, and Franschetti-Klein share facial features with Miller syndrome.

Citations

Genetics

Miller syndrome is inherited in an autosomal recessive manner and is caused by mutations in the DHODH gene. DHODH (OMIM#126064, dihydroorotate dehydrogenase) encodes a mitochondrial protein located on the outer surface of the inner mitochondrial membrane which converts dihydroorotate to orotic acid in pyrimidine production. To date, 16 unique mutations have been documented in HGMD (Human Gene Mutation Database): missense (14/16); nonsense (1/16), and a small deletion (1/16). No large deletions/insertions have been reported (Ng et al. Nat Genet 42(1): 30-35, 2010; Rainger  et al. Hum Mol Genet 21(18):3969-3983, 2012).

Citations

Testing Strategy

The DHODH protein is coded by exons 1 to 9 of the DHODH gene on chromosome 16q22. Testing involves PCR amplifications from genomic DNA and bidirectional Sanger sequencing of the coding exons and ~20bp of adjacent noncoding sequences.  We will also sequence any single exon (test#100) or pair of exons (test#200) in family members of patients with known mutations or to confirm research results.

Indications for Test

Candidates for this test are patients with symptoms consistent with Miller syndrome and the family members of patients who have known DHODH mutations.

 

Gene(s)

Offical Gene Symbol OMIM Id
DHODH

 

Gene References

Genomic mRNA Protein CCDS
NC_000016.9 NM_001361.3 NP_001352.2 CCDS42192.1
126064

Disease(s)

Name OMIM Id
Miller Syndrome 263750

Related Tests

Contacts

Genetic Counselors

Geneticist(s)

Methods and Pricing

Test Methods

Bi-directional Sanger Sequencing

Test Number Test Price CPT Code
1019 DHODH Sanger Sequencing $750 81479
100 DHODH Targeted Familial Mutations - Single Exon Sequencing $250 81479
200 DHODH Targeted Familial Mutations - Double Exon Sequencing $370 81479
300 DHODH Targeted Familial Mutations - Triple Exon Sequencing $440 81479

Test Procedure

As required, DNA is extracted from the patient specimen using a 5 Prime ArchivePure DNA Blood Kit. PCR is used to amplify the indicated exons plus additional flanking intronic or other non-coding sequence. After cleaning of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. Products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. Sequencing is performed separately in both the forward and reverse directions.

Clinical Sensitivity

Analytical sensitivity should be high because almost all of the documented DHODH mutations are point mutations, which are expected to be detected by direct sequencing of genomic DNA. Rainger et al (2012) identified compound heterozygous mutations in three out of eight unrelated families with  Miller syndrome (Rainger  et al. Hum Mol Genet 21(18):3969-83, 2012).

Citations

Analytical Validity

As of July 2013, we compared 7.3 megabases of Sanger DNA sequence generated at PreventionGenetics to NextGen sequence generated in other labs. We detected only 4 errors in our Sanger sequences, and these were all due to allele dropout during PCR. For Proficiency Testing, both external and internal, in the 9.5 years of our lab operation we have Sanger sequenced roughly 2,000 PCR amplicons (~ 1 megabase). No errors have been identified.

Our Sanger sequencing is capable of detecting virtually all nucleotide substitutions within the PCR amplicons. Similarly, we detect essentially all heterozygous or homozygous deletions within the amplicons. Homozygous deletions which overlap one or more PCR primer annealing sites are detectable as PCR failure. Heterozygous deletions which overlap one or more PCR primer annealing sites are usually not detected (see Analytical Limitations). All heterozygous insertions within the amplicons up to about 100 nucleotides in length appear to be detectable. Larger heterozygous insertions may not be detected. All homozygous insertions within the amplicons up to about 300 nucleotides in length appear to be detectable. Larger homozygous insertions may masquerade as homozygous deletions (PCR failure).

Analytical Limitations

In exons where our sequencing did not reveal any variation between the two alleles, we cannot be certain that we were able to PCR amplify both of the patient’s alleles. Occasionally, a patient may carry an allele which does not amplify, due for example to a deletion or a large insertion. In these cases, the report contains no information about the second allele.

Similarly, our sequencing tests have almost no power to detect duplications, triplications, etc. of the gene sequences.

In most cases, only the indicated exons and roughly 20 bp of flanking non-coding sequence on each side are analyzed. Test reports contain little or no information about other portions of the gene, including many regulatory regions.

In nearly all 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 for example 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 and cycle sequencing.

Unless otherwise indicated, the sequence data that we report are based on DNA isolated from a specific tissue (usually leukocytes). Test reports contain no information about gene sequences in other tissues.

Turnaround Time

Maximum of 40 days, although many tests are completed in 2-3 weeks.

Version: 1.1
Last Updated 06/24/2013