# PIKI-1, a class II PI 3-kinase, functions in endocytic trafficking

## TL;DR

This study identifies a role for the PIKI-1 lipid kinase in endocytic trafficking in C. elegans, showing how it interacts with other proteins to regulate intracellular transport.

## Contribution

The paper reveals a novel connection between lipid modification and endocytic trafficking regulation in C. elegans.

## Key findings

- PIKI-1 is responsible for PI(3,4)P2 production in C. elegans epidermis.
- Reduced PIKI-1 affects early endosome size and protein composition.
- Inhibition of HIPR-1 suppresses nekl molting defects.

## Abstract

Cellular membrane trafficking, including endocytosis and exocytosis, is a complex process coordinated by trafficking-associated proteins, cargo molecules, the cytoskeleton, and membrane lipids. The NIMA-related kinases NEKL-2 (human NEK8/9) and NEKL-3 (human NEK6/7) are conserved regulators of membrane trafficking in Caenorhabditis elegans that are required for the completion of molting. Using a genetic approach, we isolated reduction-of-function mutations in piki-1 that suppress nekl-associated molting defects. piki-1 encodes the sole predicted C. elegans Class II PI 3-kinase (PI3K), a relatively understudied class of lipid modifiers that contribute to the production of PI 3-phosphate (PI(3)P) and PI 3,4-bisphosphate (PI(3,4)P2). Using genetically encoded lipid sensors, we found that PIKI-1 was responsible for the production of PI(3,4)P2 in the C. elegans epidermis but played only a minor role in contributing to PI(3)P levels. Consistent with this, both PI(3,4)P2 and PIKI-1 partially colocalized to early endosomes, and reduction of PIKI-1 affected the size and protein composition of early endosomal compartments marked by RAB-5, EEA-1, and SNX-1. Reduced PIKI-1 also led to increased tubulation of endosomal compartments associated with recycling or the degradation of cellular debris. In contrast to studies using mammalian cell culture, PIKI-1 was largely dispensable for clathrin-mediated endocytosis in the worm epidermis, a polarized epithelium. Notably, reduction of PIKI-1 function mitigated defects in early endosomes associated with the depletion of NEKL-2. We propose that reduction of PIKI-1 function may suppress nekl molting defects by partially restoring endocytic trafficking function within a subset of compartments, including the early endosome. We also show that inhibition of HIPR-1, an ortholog of the mammalian PI(3,4)P₂-binding proteins, HIP1 and HIPR1, suppresses nekl molting defects, consistent with a model that loss of PIKI-1 alters the binding of endocytic regulators in a manner that partially compensates for the loss of NEKL-2 activity.

The uptake of materials from outside the cell and their subsequent delivery to specific intracellular locations are essential for cell function and survival. Two of the mechanisms that control this complex intracellular pathway involve the modification of proteins and of lipids, processes that are highly conserved across species. In this study, we used the model organism Caenorhabditis elegans, which is highly amenable to cell biological and genetic approaches, to establish a novel connection between these two regulatory mechanisms and demonstrate the importance of lipid modifications in maintaining the normal functioning of intracellular transport. Our results also provide insights into the fundamental cellular functions of proteins associated with human disease including cancer and metabolic disease.