Eif6 Dosage Alleviates Activation of the Tp53 Pathway in Sbds-Deficient Cells: A Mechanism for Somatic Genetic Rescue in Shwachman-Diamond Syndrome
Authors: Oyarbide Usua , Bezzerri Valentino , Staton Morgan , Boni Christian , Shah Arish , Cipolli Marco , Calo Eliezer , Corey Seth J 
Published: December 2024

Shwachman-Diamond syndrome (SDS) is characterized by skeletal abnormalities, pancreatic insufficiency, and neutropenia, with an increased risk of developing myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Nearly all cases of SDS are caused by biallelic mutations in the SBDS gene, which is critical for ribosome assembly. SBDS interacts with EFL1 to remove EIF6 from the 60S ribosomal subunit, facilitating ribosome formation in the cytoplasm.

Research using mouse and zebrafish models has shown that Sbds deficiency activates the tumor suppressor protein Tp53, leading to developmental abnormalities and tissue-specific defects. TP53 biallelic mutations have been found in SDS patients with MDS/AML, suggesting a survival advantage for these mutations. SBDS deficiency is also linked to acquired deletions of chromosome 20q, where the EIF6 gene is located, or somatic mutations in EIF6. These changes, which have been termed somatic genetic rescue, are thought to compensate for the ribosome assembly defect in SDS and lower the risk of MDS/AML. Yet the mechanism of somatic genetic rescue has not been characterized.

We created sbds-null zebrafish that exhibited Eif6 accumulation, changes in levels of ribosome proteins, and activation of Tp53 pathways. We have generated an eif6 knockout (KO) line which died earlier (~7-10 days post-fertilization) than the sbds KO line (~15 days post-fertilization). To determine the role of a Eif6 dose effect, we generated zebrafish mutants with low Eif6 protein expression (10% of the wildtype). Surprisingly, we observed that these eif6 hypomorph mutants survived to adulthood. Polysome profiling revealed significant reductions in the 80S monosomes and 40S ribosomal subunits in the eif6 KO at 5 dpf. However, the eif6 hypomorph mutants displayed polysome profiles similar to the eif6 wild type.

We crossed the eif6 KO or hypomorph mutants with sbds-null fish and analyzed their phenotypic and molecular characteristics. In the sbds-null background, expression of eif6+/- significantly but partially extended their survival at 15 days post fertilization. The eif6 hypomorph mutants also increased larvae survival at 20 days post fertilization with Mendelian ratios. Low Eif6 levels did not rescue neutropenia in Sbds-deficient zebrafish. Notably, the eif6 hypomorph mutants in sbds-null background reduced the expression of Tp53-dependent targets (e.g., cdkn1a, bax, and puma). SDS patient-derived cell lines also showed accumulation of EIF6, TP53, and CDKN1A (p21). Knocking down EIF6 significantly decreased CDKN1A mRNA levels. These observations support the hypothesis that low levels of Eif6 mitigate Tp53 pathway activation and partially alleviate cellular stress in Sbds-deficient cells.

These findings highlight the complex relationships between SBDS, EIF6, TP53, and stress responses in the pathogenesis of SDS. EIF6 dosage determines the degree of intracellular stress mediated by TP53, which may explain the somatic genetic rescue in SDS and the decreased risk for malignant transformation. Understanding these mechanisms will aid in developing therapeutic strategies for SDS. In addition, this partial dosage effect suggests additional stress pathways that are unlikely to be TP53-dependent. We are currently identifying TP53-independent mechanisms.