RNAi-based biosynthetic pathway screens to identify in vivo functions of non-nucleic acid-based metabolites such as lipids.

The field of metabolomics continues to catalog new compounds, but their functional analysis remains technically challenging, and roles beyond metabolism are largely unknown. Unbiased genetic/RNAi screens are powerful tools to identify the in vivo functions of protein-encoding genes, but not of nonproteinaceous compounds such as lipids. They can, however, identify the biosynthetic enzymes of these compounds-findings that are usually dismissed, as these typically synthesize multiple products.

Vesicular sorting controls the polarity of expanding membranes in the C. elegans intestine.

Biological tubes consist of polarized epithelial cells with apical membranes building the central lumen and basolateral membranes contacting adjacent cells or the extracellular matrix. Cellular polarity requires distinct inputs from outside the cell, e.g.

Intracellular lumen extension requires ERM-1-dependent apical membrane expansion and AQP-8-mediated flux.

Many unicellular tubes such as capillaries form lumens intracellularly, a process that is not well understood. Here we show that the cortical membrane organizer ERM-1 is required to expand the intracellular apical/lumenal membrane and its actin undercoat during single-cell Caenorhabditis elegans excretory canal morphogenesis. We characterize AQP-8, identified in an ERM-1-overexpression (ERM-1[++]) suppressor screen, as a canalicular aquaporin that interacts with ERM-1 in lumen extension in a mercury-sensitive manner, implicating water-channel activity.

Clathrin and AP-1 regulate apical polarity and lumen formation during C. elegans tubulogenesis.

Clathrin coats vesicles in all eukaryotic cells and has a well-defined role in endocytosis, moving molecules away from the plasma membrane. Its function on routes towards the plasma membrane was only recently appreciated and is thought to be limited to basolateral transport. Here, an unbiased RNAi-based tubulogenesis screen identifies a role of clathrin (CHC-1) and its AP-1 adaptor in apical polarity during de novo lumenal membrane biogenesis in the C.

Apicobasal domain identities of expanding tubular membranes depend on glycosphingolipid biosynthesis.

Metazoan internal organs are assembled from polarized tubular epithelia that must set aside an apical membrane domain as a lumenal surface. In a global Caenorhabditis elegans tubulogenesis screen, interference with several distinct fatty-acid-biosynthetic enzymes transformed a contiguous central intestinal lumen into multiple ectopic lumens. We show that multiple-lumen formation is caused by apicobasal polarity conversion, and demonstrate that in situ modulation of lipid biosynthesis is sufficient to reversibly switch apical domain identities on growing membranes of single post-mitotic cells, shifting lumen positions.