Reduced EIF6 dosage attenuates TP53 activation in models of Shwachman-Diamond syndrome

Shwachman-Diamond syndrome (SDS) is characterized by neutropenia, exocrine pancreatic insufficiency, and bony abnormalities with an increased risk of myeloid neoplasia. Almost all cases of SDS result from biallelic mutations in SBDS. SBDS interacts with EFL1 to displace EIF6 from the 60S ribosomal subunit. Released EIF6 permits the assembly of ribosomal large and small subunits in the cytoplasm. Decreased EIF6 levels due to haploinsufficiency or missense mutations which lead to decreased protein expression may provide a somatic genetic rescue and anti-leukemic effects. We observed accumulation of EIF6 protein in sbds knockout (KO) zebrafish models, confirmed in patient-derived tissues, and correlated with changes in ribosome proteins and TP53 pathways. The mechanism of action for this adaptive response is unknown. To address this, we generated an eif6 zebrafish KO line which do not survive past 10 days post fertilization. We also created two mutants with low Eif6 expression, 5-25% of the wildtype levels, that can survive until adulthood. We bred them with sbds-null strains and analyzed their phenotype and biochemical properties. Low Eif6 levels reduced Tp53 pathway activation but did not rescue neutropenia in Sbds-deficient zebrafish. Further studies elucidating the interplay between SBDS, EIF6, TP53, and cellular stress responses offer promising insights into SDS pathogenesis, somatic genetic rescue, and therapeutic strategies. RNA was extracted from pools of 8–9 individually genotyped larvae at 10 dpf using TRIzol. Three pools of eif6-/- or wildtype from the same clutch were compared. RNA quality was determined by Bioanalyzer (Agilent), and eif6 mRNA expression was measured by RT-qPCR. RNA-Seq library preparation and sequencing and mapping of 3 pools of eif6-/- and 3 pools of eif6+/+ were performed by the Beijing Genome Institute (4). Based on the FPKM values (39), we used EBSeq R package for differentially expressed gene detection between eif6-/- and eif6+/+ (fold change > 2, and P < 0.05).