SBDS-Deficient Cells Have an Altered Homeostatic Equilibrium due to Translational Inefficiency Which Explains their Reduced Fitness and Provides a Logical Framework for Intervention

Abstract

Ribosomopathies are a family of inherited disorders caused by mutations in genes necessary for ribosomal function. Shwachman-Diamond Bodian Syndrome (SDS) is an autosomal recessive disease caused, in most patients, by mutations of the SBDS gene. SBDS is a protein required for the maturation of 60S ribosomes. SDS patients present exocrine pancreatic insufficiency, neutropenia, chronic infections, and skeletal abnormalities. Later in life, patients are prone to myelodisplastic syndrome and acute myeloid leukemia (AML). It is unknown why patients develop AML and which cellular alterations are directly due to the loss of the SBDS protein. Here we derived mouse embryonic fibroblast lines from an SbdsR126T/R126T mouse model. After their immortalization, we reconstituted them by adding wild type Sbds. We then performed a comprehensive analysis of cellular functions including colony formation, translational and transcriptional RNA-seq, stress and drug sensitivity. We show that: 1. Mutant Sbds causes a reduction in cellular clonogenic capability and oncogene-induced transformation. 2. Mutant Sbds causes a marked increase in immature 60S subunits, limited impact on mRNA specific initiation of translation, but reduced global protein synthesis capability. 3. Chronic loss of SBDS activity leads to a rewiring of gene expression with reduced ribosomal capability, but increased lysosomal and catabolic activity. 4. Consistently with the gene signature, we found that SBDS loss causes a reduction in ATP and lactate levels, and increased susceptibility to DNA damage. Combining our data, we conclude that a cell-specific fragile phenotype occurs when SBDS protein drops below a threshold level, and propose a new interpretation of the disease.

High levels of eukaryotic Initiation Factor 6 (eIF6) are required for immune system homeostasis and for steering the glycolytic flux of TCR-stimulated CD4+ T cells in both mice and humans

“…Extracts were then centrifuged at 10,000g for 10 min at 4 °C. The acid supernatant was neutralized with K 2 CO 3 and used for luminometric determination of ATP (ATP determination kit, Molecular Probes) using the method of Lundin (Lopez-Lluch et al, 2006) as modified in (Calamita et al, 2017).…”

Shwachman-Diamond Syndrome Protein SBDS Maintains Human Telomeres by Regulating Telomerase Recruitment

“…Accumulating evidence strongly suggests a ribosomal function of SBDS, as human SBDS could interact with certain ribosomal subunits and play an important role in ribosome maturation (Calamita et al, 2017; Huang and Shimamura, 2011). A recent report further supports the idea that SDS belongs to ribosomopathies because defective ribosome synthesis and protein translation insufficiency was observed in an SBDS-deficient mouse model (Tourlakis et al, 2015).…”

“…For example, two of the most frequent disease-associated mutations are 183-184TA/→CT and 258+2T/→C, which result in two premature SBDS-truncated proteins with an in-frame stop codon (K62X) and a frameshift mutation (84Cfs3), respectively (Boocock et al, 2003). SBDS has been implicated in mRNA translation and ribosome biogenesis (Calamita et al, 2017; Ganapathi et al, 2007). Previous reports also suggest that SBDS may contribute to telomere regulation, because SDS and SBDS -mutated aplastic anemia (AA) patients have shorter telomeres (Calado et al, 2007; Thornley et al, 2002).…”

The eukaryotic Initiation Factor 6 (eIF6) regulates ecdysone biosynthesis by modulating translation inDrosophila

“…In Drosophila, we found that an overexpression of eIF6 leads to a reduction of the free 60S pool in eye imaginal discs, consistent with eIF6 biochemical activity. Such reduction could imply lower general translation, due to less availability of 60S subunits, as in the case of Sbds mutants 39 . Conversely, 60S could be already engaged with 40S into active translating 80S, thus heightening general translation.…”