Aging, Dauer, and Stature Phenotypes Are Conferred by Structure‐Directed Missense Mutations in the Endogenous‐1/Phosphatidylinositol 3‐Kinase Catalytic Subunit

Abstract

ABSTRACT Phosphatidylinositol 3‐kinase (PI3K) integrates insulin/IGF signaling (IIS) and Ras inputs to control lifespan, metabolism and growth. Yet the organismal consequences of selective structural perturbations remain poorly understood. Using structure‐guided CRISPR/Cas9‐dependent genome editing, we dissected functions of AGE‐1, the sole Class IA PI3K catalytic subunit in. An endogenously tagged AGE‐1, containing a long flexible linker, epitope and fluorescent tag, retained full activity, enabling visualization of native protein dynamics in vivo. A likely constitutively activating E630K substitution, modeled on oncogenic p110α alleles, markedly shortened lifespan and enhanced Ras‐dependent induction of primary vulval precursor cell (VPC) fate, confirming evolutionary conservation of PI3K activation mechanisms that directly modulate longevity and development. Structural modeling further guided mutation of AGE‐1 residues predicted to mediate Ras binding. Surprisingly, a putative AGE‐1 variant defective in Ras association, together with a complementary Ras effector‐binding mutation, produced enlarged animals with reduced dauer formation. These phenotypes reveal a previously unrecognized Ras>PI3K signaling axis that restrains somatic growth and promotes entry into diapause, counter to canonical IIS models. Together, these structure‐informed alleles show that discrete PI3K structural perturbations can differentially uncouple lifespan, growth, and developmental outcomes in vivo. By combining structural modeling with genome editing in a tractable aging model, this work establishes a framework for dissecting conserved signaling enzymes at single‐residue resolution and uncovers unexpected organismal roles for PI3K structure in coordinating growth and longevity. Structure‐guided missense mutations in theAGE‐1/PI3Kinase catalytic subunit and its LET‐60/Ras‐binding interface reveal separable signaling outputs from the Ras‐bound PI3K complex. Enhanced canonical catalytic activity, conferred by a gain‐of‐function mutation modeled on oncogenic human p110⍺ PI3K variants, promotes aging, reproductive growth, and cell fate specification. In contrast, the Ras‐PI3K interaction selectively restrains animal growth and promotes dauer/diapause entry, demonstrating context‐dependent and functionally distinct outputs of PI3K signaling in vivo. graphical