Regulation of Macromolecular Biosynthesis in a Mutant of Escherichia coli Defective in Membrane Phospholipid Biosynthesis

Abstract

Nucleic acid and protein synthesis were studied in temperature-sensitive mutants defective in phospholipid synthesis. The defect is due to a single mutation in glycerol 3-phosphate acyltransferase (EC 2.3.1.15). The results show that at the restrictive temperature not only does phospholipid synthesis cease, but DNA, RNA, and protein synthesis also cease. Active transport continues, however, indicating that the cells do not become leaky or lose their energy supply. These results suggest that phospholipid synthesis is coupled to DNA, RNA, and protein synthesis.Since lipid synthesis, and consequently membrane synthesis, is necessary for growth of cells, it is reasonable that cells would have regulatory mechanisms to couple the synthesis of lipids with the synthesis of macromolecules in order to maintain balanced cell growth. The availability of mutants in lipid biosynthesis has helped to elucidate the contribution of phosl)holipids to membrane structure and function (1). In particular, one apparent result was that protein synthesis and cell growth could continue for a significant amount of time in the absence of lipid synthesis. When an unsaturated fattyacid auxotroph of Escherichia coli was deprived of unsaturated fatty acids, growth, as well as DNA, RNA, and protein synthesis, continued for several hours (2-5). Similar results were obtained when 3-decynoyl-N-acetylcysteamine was used to inhibit unsaturated fatty-acid synthesis (4, 6, 7). Bacillus subtilis (8-10), Staphylococcus aureus (11), and E. coli (12) glycerol auxotrophs have been studied. When these mutants were deprived of glycerol, net phospholipid synthesis stopped immediately. However, macromolecular synthesis and cell growth continued for some time.Recently this laboratory has described a temperaturesensitive mutant of E. coli, CV15, that does not grow or make phospholipid at the restrictive temperature (13). The defect is due to a single mutation in glycerol 3-phosphate acyltransferase (EC 2.3.1.15), which catalyzes the first reaction unique to phospholipid biosynthesis. In this paper, detailed studies of the synthesis of phospholipid, DNA, RNA, and protein are reported for mutants with a thermolabile glycerol 3-phosphate acyltransferase. The results show that DNA, RNA, and protein synthesis do not continue at the restrictive temperature; the shutoff of the synthesis of these macromolecules parallels the shutoff of phospholipid synthesis. Active transport continues, however, indicating that the cells do not become leaky or lose their energy supply. MATERIALS AND METHODSBacterial Strains, Mfedia, and Growth Conditions. CV15 and its parent, strain 8, were described previously (13).Abbreviations: TMG, methyl-l-thio-j-D-galactopyranoside; IPTG, isopropyl-l-thio-,f-D-galactopyranoside.

Biogenesis of membrane lipids: mutants of Escherichia coli with temperature-sensitive phosphatidylserine decarboxylase.

“…1 and 2). Until recently, the only phospholipid mutants available were affected early in the pathway at the level of phosphatidic acid synthesis (3)(4)(5)(6)(7). Reports have recently appeared, however, describing the isolation of mutants of Escherichia coli defective in some early stage of the formation of phosphatidylethanolamine, the predominant phospholipid in E. coli (8)(9)(10).…”

Identification of a Gene Encoding Adenylate Kinase Involved in Antifungal Activity Expression of the Biocontrol Strain Burkholderia pyrrocinia CH-67

“…Adenylate kinase plays a key role in the interconversion of the constituents of the adenine nucleoside pool, catalyzing the reversible reaction ATP + AMP to ADP (Glaser et al, 1975). Regulation of adenylate kinase activity may provide the cell with a mechanism for controlling macromolecular biosynthesis such as membrane phospholipids and regulating cell growth (Goelz and Cronan, 1982;Huss and Glaser, 1983).…”

Isolation and characterization of adenylate kinase (adk) mutations in Salmonella typhimurium which block the ability of glycine betaine to function as an osmoprotectant

“…Most adk mutations in E. coli have been isolated as temperature-sensitive defects (9,20,29), suggesting that Adk is an essential enzyme. Shift of the adk mutants to the nonpermissive temperature results in rapid decline in the ATP pool, increase in AMP, and cessation of DNA, RNA, and protein synthesis (20,21).…”

“…Shift of the adk mutants to the nonpermissive temperature results in rapid decline in the ATP pool, increase in AMP, and cessation of DNA, RNA, and protein synthesis (20,21). However, there are a number of biochemical and genetic studies which indicate an even more complex role for Adk besides the synthesis of ADP or the maintenance of the equilibrium among the adenylates.…”

“…However, there are a number of biochemical and genetic studies which indicate an even more complex role for Adk besides the synthesis of ADP or the maintenance of the equilibrium among the adenylates. Several adk mutants have emerged from selections designed to yield mutants with temperature-sensitive lesions in lipid synthesis (20,32). Single mutations in the adk gene were found to be able to cause temperature sensitivity in sn-glycerol-3-phosphate acetyltransferase, which is a membrane-bound enzyme catalyzing one of the early steps of phospholipid synthesis.…”