Page 51 - Haematologica June

failure syndromes,46 we speculated that the hematologic remission in our patients was attained via genetic rever- sion (e.g. uniparental disomy). Sequencing of DNA in P1-3 either at multiple time points or after the acquisition of treatment independence did not reveal any changes in mutation load (Figure 2B).
Mutations in RPL15 recapitulate specific pre-rRNA processing defects found in RP depleted cells
Mutations in DBA-linked RP genes result in haploinsuf- ficiency of the encoded RP. Since the majority of RPs incorporate into pre-ribosomal particles in a progressive
manner concurrently with pre-rRNA maturation, a lack of one RP impairs pre-rRNA processing in an explicit way that is reproducible in most cell types.47 Online Supplementary Figure S2 illustrates the normal pre-rRNA processing pathway that begins with a single transcribed strand of 47S pre-rRNA that then undergoes a complex series of cleavages and trimmings to ultimately result in the mature 18S, 28S, and 5.8S strands of rRNA found in ribosomes. To investigate the functional consequences of the newly identified RP gene mutations reported here, EBV-immortalized lymphoblast cell lines (LCLs) were gen- erated and the pre-rRNA processing of these LCLs was
RPL15 mutations in DBA
A
B
Figure 2. Longitudinal mutations in RPL15 are identified in individuals diagnosed with Diamond-Blackfan anemia (DBA). (A) Clinical evolution of P1-P3 carrying truncating hotspot mutation RPL15 p.Tyr81*. Patient 1 manifest- ed with DBA after birth and after one transfu- sion achieved spontaneous remission at the age of six months. A relapse occurred five years later and after a short course of steroids the patient attained treatment inde- pendence. Patients 2 and 3 had a similar clin- ical course with hydrops fetalis and prenatal intrauterine transfusions, and achieved treat- ment independence either spontaneously or after one course of steroids. (B) Sanger sequencing results of the recurring mutation in RPL15 from initial diagnoses as well as post remission. Arrows indicate the inserted A nucleotide. FUP: follow up; BM: bone marrow; PB: peripheral blood; LCL: lymphoblastoid cell lines.
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