We describe five patients carrying identical 1.65 Mb deletions of 16p13.11 encompassing 15 genes. In addition, we found five patients carrying apparently reciprocal duplications of this same region. The rearrangement breakpoints are located in low copy repeats implying that non-allelic homologous recombination between these flanking LCRs mediates these rearrangements. Three further patients presenting with larger atypical rearrangements showed breakpoints that also mapped to clusters of LCR16.
Despite careful parental analysis, observations of rearrangements of 16p13.11 make it difficult to distinguish between disease-causing events (generally presumed to be de novo) and benign variants (that do not contribute to a phenotype). Two deletions and one duplication were found to be inherited from an apparently normal parent, and another duplication was inherited from a clinically mildly abnormal parent. Conversely, we also found a duplication that occurred de novo. Previous studies have reported 16p13.11 deletions occurring both de novo and by inheritance from a normal parent, while duplications were reported to be inherited from normal parents in two families.11 22 23
One interpretation of these observations could be that rearrangements of this region are benign variants, and that the observed phenotypes are coincidental with the presence of the imbalance. However, one case–control study reported a significantly higher incidence of the del16p13.11 in patients with MR/MCA, implying that the del16p13.11 is a risk factor contributing to the MR/MCA phenotype. The duplication is present in equal frequency in the normal and the patient population, indicating this variant is compatible with a normal phenotype. The identification of this dup16p13.11 in a patient with fragile X (patient 9) in our study lends support to this hypothesis. Recently, Ullmann et al11
reported three carriers of dup16p13.11 in a cohort of 182 people with autism. Because the duplication was found in 5 out of 2014 normal participants in our study, the duplication seems overrepresented in the autistic population (p
0.023, Fisher exact test). More studies are needed strengthen this association.
Three of the four adult patients with 16p13.11 deletions had both microcephaly and seizures, and the two fetuses with the deletions had brain anomalies, one of which also had relative microcephaly. The same three adult patients also have small stature. Small stature and dysmorphic features were also a feature of one of the patients described by Ullman et al
In contrast, our fourth adult patient and a previously described deletion carrier22
had normal head circumference (55.3 cm), while one previously reported patient had macrocephaly.11
Although the typical 16p13.11 duplication may be a benign variant, it is striking that four of our duplication carriers not only presented with MR but also behavioural problems. Similarly, members of two of three families with duplications reported by Ullman et al11
had similar behavioural problems. Although we do not have data on the behavioural phenotypes of all patients with MR/MCA tested using the arrays, the occurrence of this type of behaviour in this patient population seems higher than average, and hence, it seems plausible that dup16p13.11 carriers may have a predisposition for aggressive behaviour.
To date, most genomic imbalances have been classified as either benign or pathogenic and most microdeletion syndromes are presumed to be well-defined clinical conditions. However, even well-known genomic disorders can be phenotypically heterogeneous and more variable than originally thought, owing to incomplete penetrance or variable expression. The variability of the del22q11 phenotype originally led to their different clinical classification as DiGeorge syndrome (heart and thymus defects) (OMIM 188400), Sprintzen syndrome (speech difficulties) or velocardiofacial syndrome (VCFS) (conotruncal anomaly face) (OMIM 192430). Recently, several reports have been published about atypical patients with 22q11 deletions.24 25
Equally, in dup22q11 carriers, the phenotype may range from severe MR through to completely unaffected, and various minor developmental anomalies are noted.26 27
In addition, 22q11 duplications can be inherited from apparently normal parents.26 28
Therefore, it is possible that 16p13.11 duplications might also be causative and the heterogeneous phenotype of our patients explained in part by (1) the unbiased selection criteria, (2) variability due to other genetic or possibly environmental determinants, (3) incomplete penetrance, (4) variable expression or (5) unmasking of recessive alleles.29
In particular, owing to the presence of del16p13.11 in a normal parent of one of our probands, we set out to test the latter hypothesis. Because microcephaly was observed in two of the three adult patients with typical deletions and in one of the two fetuses with deletion, NDE1
was considered an excellent candidate gene for this phenotype. NDE1
is strongly expressed in brain,30
and forms complexes with LIS1
, a dosage-sensitive gene that is crucial for neuronal migration and cerebral development, and that is known to underlie Miller–Dieker lissencephaly syndrome (OMIM 247200). Furthermore, Nde1
-null mice show microcephaly.31
However, sequencing of all exons of NDE1
in four patients with deletions did not reveal any mutation on the remaining allele, suggesting that this is not the mechanism responsible for the phenotype in our patients. Another plausible candidate gene is NTAN1
(asparagine-specific N-terminal amidase). Mouse models deficient for this enzyme showed alterations in activity, social behaviour and memory.32
However although this region is reported in the Redon database (http://projects.tcag.ca/variation/
) to have CNVs, learning and memory defects and aberrant social behaviour could not be excluded among controls and therefore the NTAN1
gene remains a good candidate gene.
Imprinting is a mechanism that could potentially explain the presence of these rearrangements in unaffected relatives.11
However, there are no known imprinted genes on chromosome 16 (http://www.geneimprint.com/site/genes-by-species
). Furthermore, for both deletions and duplications of 16p we observed the inheritance of these imbalances from normal parents through both the maternal and paternal germ lines, indicating that imprinting does not significantly influence patient phenotype of this disorder.
In addition to the common 1.65 Mb rearrangement observed in the 10 patients reported here, 3 “atypical” chromosomal imbalances either overlapping or flanking this common region were found in patients with MR/MCA. Breakpoints for all three imbalances were also located within LCR16 sequences. However, the atypical 3.4 Mb duplication seems to be mediated by different LCR16s compared with the atypical deletion. Therefore, the complex structure of the LCR16s in this region seems to be involved in generating a variety of different chromosomal rearrangements. The finding of variably sized rearrangements on chromosome 16p is similar to that observed for other recurrent genomic disorders, such as the Prader–Willi/Angelman syndrome,33
and the 15q24 deletion syndrome,35
in which recombination within alternate LCRs can result in recurrent deletions and duplications of different sizes.
In conclusion, we report a novel genomic disorder likely caused by NAHR between copies of LCR16. Although in some cases this is inherited from a normal parent, we found a strong association of the deletion with developmental disorders. Reciprocal duplications were observed as both inherited and de novo events, and were also identified in several controls, suggesting that the duplication by itself confers either no phenotype at all or a range of phenotypes of varying severity. Alternatively, the duplication may require additional predisposing factors to have a phenotypic effect. Our findings have important implications for genetic counselling. Traditionally, chromosomal imbalances inherited from a normal parent were considered benign, while de novo chromosomal imbalances were considered pathogenic. Although our results suggest that the inherited 16p13.11 deletion is likely causal for the phenotype, the clinical significance of both de novo and inherited duplications remains uncertain and they may be benign variants. The study of additional patients and normal individuals with 16p13.11 rearrangements is required to reinforce this hypothesis and to obtain better insight in the potential pathology associated with the observed microdeletion and microduplication events.