Functional characterization of multiple DICER1 mutations in an adolescent

M K Wu, L de Kock, L S Conwell, C J R Stewart, B R King, C S Choong, K Hussain, N Sabbaghian, I J MacRae, M R Fabian, W D Foulkes

DICER1 is an endoribonuclease which catalyzes the generation of mature 5p and 3p miRNAs. We present the detailed molecular characterization of three DICER1 mutations occurring in a child with multinodular goitre (MNG) and an ovarian sex cord-stromal tumor (OSCST), conditions associated with the DICER1 syndrome (OMIM 601200). We identified three DICER1 mutations affecting the RNase IIIb domain in the child: a novel germ-line DICER1 mutation (c.5441C>T) which does not affect the metal ion binding residues of RNase IIIb, and two different somatic mutations (5425G>T in the MNG and c.5125G>A in the OSCST), both situated at these metal ion binding residues. In vitro functional studies of the germ-line DICER1 (c.5441C>T) mutation showed that both 5p and 3p miRNAs were decreased, whereas the somatic mutation occurring in the MNG, c. 5425G>T, is predicted to function either as a missense (p.D1810Y) or a cryptic splice site (p.K1844fsX17). When the latter, no miRNAs are made. We also identified the previously studied c.5125G>A (p.D1709N) mutation in the OSCST. Following identification of the c.5125G>A, review of the original diagnosis of the OSCST (adult granulosa cell tumour) was revised to Sertoli-Leydig cell tumour, illustrating how molecular analysis can assist in the diagnosis of uncommon ovarian neoplasms. We also report the first instance of biallelic DICER1 mutations in a MNG. The three single base pair substitutions in DICER1 all affect the RNase IIIb domain, but have different effects on miRNA generation. This study demonstrates that detailed functional characterisation of DICER1 missense mutations may be required to determine pathogenicity.