SBDSR126Trescues survival ofsbds−/−zebrafish in a dose-dependent manner independently of Tp53

Abstract

Defects in ribosomal biogenesis profoundly affect organismal development and cellular function, and these ribosomopathies produce a variety of phenotypes. One ribosomopathy, Shwachman–Diamond syndrome (SDS) is characterized by neutropenia, pancreatic exocrine insufficiency, and skeletal anomalies. SDS results from biallelic mutations inSBDS, which encodes a ribosome assembly factor. Some individuals express a missense mutation,SBDSR126T, along with the common K62X mutation. We reported that thesbds-null zebrafish phenocopies much of SDS. We further showed activation of Tp53-dependent pathways before the fish died during the larval stage. Here, we expressedSBDSR126Tas a transgene in thesbds−/−background. We showed that one copy of theSBDSR126Ttransgene permitted the establishment of maternal zygoticsbds-null fish which produced defective embryos withcdkn1aup-regulation, a Tp53 target involved in cell cycle arrest. None survived beyond 3 dpf. However, two copies of the transgene resulted in normal development and lifespan. Surprisingly, neutropenia persisted. The surviving fish displayed suppression of female sex differentiation, a stress response in zebrafish. To evaluate the role of Tp53 in the pathogenesis ofsbds−/−fish phenotype, we bred the fish with a DNA binding deficient allele,tp53M214K. Expression of the loss-of-functiontp53M214Kdid not rescue neutropenia or survival insbds-null zebrafish. Increased expression ofcdkn1awas abrogated in thetp53M214K/M214K;sbds−/−fish. We conclude that the amount of SBDSR126Tprotein is important for development, inactivation of Tp53 fails to rescue neutropenia or survival in thesbds-null background, andcdkn1aup-regulation was dependent on WTtp53. We hypothesize that additional pathways are involved in the pathophysiology of SDS.