Enzymological analysis of the tumor suppressor A-C1 reveals a novel group of phospholipid-metabolizing enzymes

Naoki Shinohara,Toru Uyama,Xing-Hua Jin,Kazuhito Tsuboi,Takeharu Tonai,Hitoshi Houchi,Natsuo Ueda

doi:10.1194/jlr.m015081

Abstract

A-C1 protein is the product of a tumor suppressor gene negatively regulating the oncogene Ras and belongs to the HRASLS (HRAS-like suppressor) subfamily. We recently found that four members of this subfamily expressed in human tissues function as phospholipid-metabolizing enzymes. Here we examined a possible enzyme activity of A-C1. The homogenates of COS-7 cells overexpressing recombinant A-C1s from human, mouse, and rat showed a phospholipase A½ (PLA½) activity toward phosphatidylcholine (PC). This finding was confirmed with the purified A-C1. The activity was Ca²⁺ independent, and dithiothreitol and Nonidet P-40 were indispensable for full activity. Phosphatidylethanolamine (PE) was also a substrate and the phospholipase A₁ (PLA₁) activity was dominant over the PLA₂ activity. Furthermore, the protein exhibited acyltransferase activities transferring an acyl group of PCs to the amino group of PEs and the hydroxyl group of lyso PCs. As for tissue distribution in human, mouse, and rat, A-C1 mRNA was abundantly expressed in testis, skeletal muscle, brain, and heart. These results demonstrate that A-C1 is a novel phospholipid-metabolizing enzyme. Moreover, the fact that all five members of the HRASLS subfamily, including A-C1, show similar catalytic properties strongly suggests that these proteins constitute a new class of enzymes showing PLA½ and acyltransferase activities.

Highlights

A-C1 protein is the product of a tumor suppressor gene negatively regulating the oncogene Ras and belongs to the HRASLS (HRAS-like suppressor) subfamily
We examined a possible secretion of recombinant human A-C1 into the culture medium by measuring phospholipase A1/2 (PLA1/2) activity
Our present studies revealed that this protein is capable of catalyzing phospholipase A1 (PLA1)/2-like hydrolysis, PE N-acylation, and lyso PC O-acylation

Summary

Introduction

A-C1 protein is the product of a tumor suppressor gene negatively regulating the oncogene Ras and belongs to the HRASLS (HRAS-like suppressor) subfamily. Because all the other members of the HRASLS subfamily show phospholipidmetabolizing activities and because histidine-30 and cysteine-119 of A-C1 correspond to the catalytic dyad of LRAT [26] (Fig. 1), we hypothesized that A-C1 functions as an enzyme involved in phospholipid metabolism. The primary structures of A-C1 proteins were composed of 168 (human), 167 (mouse), and 167 (rat) amino acid residues, respectively (Fig. 2A).

Results

Conclusion