5q− syndrome is a myelodysplastic syndrome (MDS) subtype that is associated with defective erythroid differentiation and an increased risk of developing acute myeloid leukaemia. Patients with 5q− syndrome have a somatic monoallelic deletion of a region that encompasses 40 genes on chromosome 5q. To identify the causative gene(s), Golub and colleagues knocked down the expression of each of these 40 genes separately using lentiviral expression of short hairpin RNAs (shRNAs) in human CD34+ haematopoietic progenitor cells. By assessing the ability of these shRNA-expressing cells to differentiate into the erythrocyte lineage they showed that knockdown of
RPS14
(resulting in ∼60% reduction in protein expression) significantly recapitulated the phenotypes of 5q− syndrome. Furthermore, simultaneous knockdown of other genes within the 5q− region did not synergize with RPS14 knockdown, indicating that RPS14 haploinsufficiency alone is sufficient to cause the 5q− phenotypes. In addition, genome-wide expression profiling of CD34+ adult bone marrow cells infected with control or RPS14 shRNAs revealed that RPS14 knockdown generated a gene signature that correlated with abrogated erythroid differentiation and sensitivity to lenalidomide — the only drug that is effective for MDS patients with 5q deletions. Next, the authors excluded the possibility of biallelic inactivation of RPS14 by sequencing the remaining allele from 32 patients with 5q− syndrome, indicating that haploinsufficiency of RPS14 is the underlying genetic defect of 5q− syndrome.
So, what does RPS14 do? RPS14 is a conserved component of the 40S ribosomal subunit and it is required for 18S pre-rRNA processing in Saccharomyces cerevisiae. Therefore, the authors assessed the ratios of rRNA transcripts from erythroleukaemic cells expressing RPS14 shRNAs and showed that the ratio of 30S:18S rRNAs was increased 4–9-fold and that the formation of the 40S subunit was abrogated. Similarly, this ratio was altered in bone marrow cells from patients with 5q− syndrome. Crucially, the authors were able to rescue the erythroid differentiation defects by lentiviral expresssion of RPS14 in CD34+ cells derived from frozen bone marrow mononuclear cells obtained from 5q− syndrome patients, but not from cells derived from MDS patients without the 5q deletion, confirming that partial loss of RPS14 accounts for 5q− syndrome.
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