Cell polarity is fundamental to differentiation and function of most cells. Studies in mammalian epithelial cells have revealed that the establishment and maintenance of cell polarity depends upon cell adhesion, signaling networks, the cytoskeleton, and protein transport. Atypical protein kinase C (PKC) isotypes PKCζ and PKCλ have been implicated in signaling through lipid metabolites including phosphatidylinositol 3-phosphates, but their physiological role remains elusive. In the present study we report the identification of a protein, ASIP (atypical PKC isotype–specific interacting protein), that binds to aPKCs, and show that it colocalizes with PKCλ to the cell junctional complex in cultured epithelial MDCKII cells and rat intestinal epithelia. In addition, immunoelectron microscopy revealed that ASIP localizes to tight junctions in intestinal epithelial cells. Furthermore, ASIP shows significant sequence similarity to Caenorhabditis elegans PAR-3. PAR-3 protein is localized to the anterior periphery of the one-cell embryo, and is required for the establishment of cell polarity in early embryos. ASIP and PAR-3 share three PDZ domains, and can both bind to aPKCs. Taken together, our results suggest a role for a protein complex containing ASIP and aPKC in the establishment and/or maintenance of epithelial cell polarity. The evolutionary conservation of the protein complex and its asymmetric distribution in polarized cells from worm embryo to mammalian-differentiated cells may mean that the complex functions generally in the organization of cellular asymmetry.

Epidermal integrity is essential for animal development and survival. Here, we demonstrate that TSP-15, a member of the tetraspanin protein family, is required for epithelial membrane integrity in Caenorhabditis elegans. Reduction of tsp-15 function by mutation or by RNA interference elicits abnormalities of the hypodermis, including dissociation of the cuticle and degeneration of the hypodermis. Lethality during molting often results. Examination of GFP transgenic animals, genetic mosaic analysis and rescue assays revealed that TSP-15 functions in hyp7, a large syncytium that composes most of the hypodermis. Assays with a membrane-impermeable dye or leakage analysis of a hypodermal-specific marker indicate that the barrier function of the hypodermal membrane is impaired owing to the loss or reduction of TSP-15. These results indicate that TSP-15 functions in the maintenance of epithelial cell integrity.

Chronic exposure to nicotine leads to long-term changes in both the abundance and activity of nicotinic acetylcholine receptors, processes thought to contribute to nicotine addiction. We have found that in Caenorhabditis elegans, prolonged nicotine treatment results in a long-lasting decrease in the abundance of nicotinic receptors that control egg-laying. In naive animals, acute exposure to cholinergic agonists led to the efficient stimulation of egg-laying, a response mediated by a nicotinic receptor functionally expressed in the vulval muscle cells. Overnight exposure to nicotine led to a specific and long-lasting change in egg-laying behavior, which rendered the nicotine-adapted animals insensitive to simulation of egg-laying by the nicotinic agonist and was accompanied by a promoter-independent reduction in receptor protein levels. Mutants defective in the gene tpa-1, which encodes a homolog of protein kinase C (PKC), failed to undergo adaptation to nicotine; after chronic nicotine exposure they remained sensitive to cholinergic agonists and retained high levels of receptor protein in the vulval muscles. These results suggest that PKC-dependent signaling pathways may promote nicotine adaptation via regulation of nicotinic receptor synthesis or degradation.

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The tpa-1 gene mediates the action of tumor-promoting phorbol esters in the nematode Caenorhabditis elegans. A genomic fragment that constitutes a portion of the tpa-1 gene was cloned by Tc1 transposon tagging and was used as a probe to screen a nematode complementary DNA library. One of the isolated complementary DNA clones had a nucleotide sequence that predicts a polypeptide of 526 amino acids. The predicted amino acid sequence revealed that the predicted tpa-1 protein sequence is highly similar to protein kinase C molecules from various animals, including man.