Loss of RNASEK Terminates Egg Cylinder Development by Impairing Lysosomal Function Associated With V‐ ATPase in Mouse

Although RNASEK is defined as a subunit of V-ATPase, how it regulates the V-ATPase and relevant physiological functions remains largely uncharacterized. Utilizing a homozygous RNASEK knockout mouse, we demonstrate that the null function of RNASEK leads to catastrophic developmental failure at the egg cylinder stage (E5.5). Rnasek^-/- embryos exhibit pronounced lysosomal dysfunction and multilineage proliferation arrest, accompanied by the hallmarks of senescence, including elevated p21, reduced Ki67 and EdU incorporation as well as increased γH2AX foci, which are evident in vitro embryo culture as well. Unexpectedly, despite increased V0/V1 subunit colocalization, lysosomal alkalization, proteolytic failure, and autophagic flux blockade collectively indicate that the loss of RNASEK promotes malfunctional V-ATPase assembly. Pharmacological restoration of lysosomal acidity via EN6 partially mitigates senescence and extends the developmental window. These findings demonstrate that RNASEK regulates lysosomal function via V-ATPase and is required for egg cylinder development in the mouse embryo. Loss of RNASEK promotes premature senescence of multiple cell lineages, terminating early embryonic development.