Cardiolipin membrane domains in prokaryotes and eukaryotes.

Cardiolipin (CL) plays a key role in dynamic organization of bacterial and mitochondrial membranes. CL forms membrane domains in bacterial cells, and these domains appear to participate in binding and functional regulation of multi-protein complexes involved in diverse cellular functions including cell division, energy metabolism, and membrane transport. Visualization of CL domains in bacterial cells by the fluorescent dye 10-N-nonyl acridine orange is critically reviewed. Possible mechanisms proposed for CL dynamic localization in bacterial cells are discussed. In the mitochondrial membrane CL is involved in organization of multi-subunit oxidative phosphorylation complexes and in their association into higher order supercomplexes. Evidence suggesting a possible role for CL in concert with ATP synthase oligomers in establishing mitochondrial cristae morphology is presented. Hypotheses on CL-dependent dynamic re-organization of the respiratory chain in response to changes in metabolic states and CL dynamic re-localization in mitochondria during the apoptotic response are briefly addressed

May the Force Be With You: Unfolding Lipid-Protein Interactions By Single-Molecule Force Spectroscopy

In this issue of Structure, Serdiuk et al. report the use of single-molecule force microscopy to establish a role for phosphatidylethanolamine in promoting the native fold of lactose permease, thereby preventing it from populating a functionally defective, nonnative conformation (Serdiuk et al., 2015)

Adenine nucleotide-dependent regulation of assembly of bacterial tubulin-like FtsZ by a hypermorph of bacterial actin-like FtsA.

Cytokinesis in bacteria depends upon the contractile Z ring, which is composed of dynamic polymers of the tubulin homolog FtsZ as well as other membrane-associated proteins such as FtsA, a homolog of actin that is required for membrane attachment of the Z ring and its subsequent constriction. Here we show that a previously characterized hypermorphic mutant FtsA (FtsA*) partially disassembled FtsZ polymers in vitro. This effect was strictly dependent on ATP or ADP binding to FtsA* and occurred at substoichiometric levels relative to FtsZ, similar to cellular levels. Nucleotide-bound FtsA* did not affect FtsZ GTPase activity or the critical concentration for FtsZ assembly but was able to disassemble preformed FtsZ polymers, suggesting that FtsA* acts on FtsZ polymers. Microscopic examination of the inhibited FtsZ polymers revealed a transition from long, straight polymers and polymer bundles to mainly short, curved protofilaments. These results indicate that a bacterial actin, when activated by adenine nucleotides, can modify the length distribution of bacterial tubulin polymers, analogous to the effects of actin-depolymerizing factor/cofilin on F-actin

Biochemical characterization of the initial steps of the Kennedy pathway in Trypanosoma brucei:the ethanolamine and choline kinases

Note related output below contains correction of this paper.Ethanolamine and choline are major components of the trypanosome membrane phospholipids, in the form of GPEtn (glycero-phosphoethanolamine) and GPCho (glycerophosphocholine). Ethanolamine is also found as an integral component of the GPI (glycosylpliosphatidylinositol) anchor that is required for membrane attachment of cell-surface proteins, most notably the variant-surface glycoproteins. The de novo synthesis of GPEtn and GPCho starts with the generation of phosphoethanolamine and phosphocholine by ethanolamine and choline kinases via the Kennedy pathway. Database mining revealed two putative C/EKs (choline/ethanolamine kinases) in the Trypanosoma brucei genome, which were cloned, overexpressed, purified and characterized. TbEK 1 (T brucei ethanolamine kinase 1) was shown to be catalytically active as an ethanolamine-specific kinase, i.e. it had no choline kinase activity. The K values for ethanolamine and ATP were found to be 18.4 +/- 0.9 and 219 29 mu M respectively. TbC/EK2 (T brucei choline/ethanolamine kinase 2), on the other hand, was found to be able to phosphorylate both ethanolamine and choline, even though choline was the preferred substrate, with a K-m 80 times lower than that of ethanolamine. The K. values for choline, ethanolamine and ATP were 31.4 +/- 2.6 mu M, 2.56 +/- 0.31 mu M and 20.6 +/- 1.96 mu M respectively. Further substrate specificity analysis revealed that both TbEK1 and TbC/EK2 were able to tolerate various modifications at the amino group, with the exception of a quaternary amine for TbEK1 (choline) and a primary amine for TbC/EK2 (ethanolamine). Both enzymes recognized analogues with substituents oil C-2, but substitutions oil C-1 and elongations of the carbon chain were not well tolerated.Publisher PDFPeer reviewe

Modulation of Myocardial Mitochondrial Mechanisms during severe Polymicrobial Sepsis in the Rat

Background: We tested the hypothesis that 5-Hydroxydecanoic acid (5HD), a putative mitoKATP channel blocker, will reverse sepsis-induced cardiodynamic and adult rat ventricular myocyte (ARVM) contractile dysfunction, restore mitochondrial membrane permeability alterations and improve survival. Methodology/Principal Findings: Male Sprague-Dawley rats (350-400 g) were made septic using 400 mg/kg cecal inoculum, ip. Sham animals received 5% dextrose water, ip. The Voltage Dependent Anion Channels (VDAC1), Bax and cytochrome C levels were determined in isolated single ARVMs obtained from sham and septic rat heart. Mitochondria and cytosolic fractions were isolated from ARVMs treated with norepinephrine (NE, 10 µmoles) in the presence/absence of 5HD (100 µmoles). A continuous infusion of 5HD using an Alzet pump reversed sepsis-induced mortality when administered at the time of induction of sepsis (-40%) and at 6 hr post-sepsis (-20%). Electrocardiography revealed that 5HD reversed sepsis-induced decrease in the average ejection fraction, Simpsons+m Mode (53.5±2.5 in sepsis and 69.2±1.2 at 24 hr in sepsis+5HD vs. 79.9±1.5 basal group) and cardiac output (63.3±1.2 mL/min sepsis and 79.3±3.9 mL/min at 24 hr in sepsis+5HD vs. 85.8±1.5 mL/min basal group). The treatment of ARVMs with 5HD also reversed sepsis-induced depressed contractility in both the vehicle and NE-treated groups. Sepsis produced a significant downregulation of VDAC1, and upregulation of Bax levels, along with mitochondrial membrane potential collapse in ARVMs. Pretreatment of septic ARVMs with 5HD blocked a NE-induced decrease in the VDAC1 and release of cytochrome C. Conclusion: The data suggest that Bax activation is an upstream event that may precede the opening of the mitoKATP channels in sepsis. We concluded that mitoKATP channel inhibition via decreased mitochondrial membrane potential and reduced release of cytochrome C provided protection against sepsis-induced ARVM and myocardial contractile dysfunction. © 2011 Chopra et al

The translocator maintenance protein Tam41 is required for mitochondrial cardiolipin biosynthesis

The mitochondrial inner membrane contains different translocator systems for the import of presequence-carrying proteins and carrier proteins. The translocator assembly and maintenance protein 41 (Tam41/mitochondrial matrix protein 37) was identified as a new member of the mitochondrial protein translocator systems by its role in maintaining the integrity and activity of the presequence translocase of the inner membrane (TIM23 complex). Here we demonstrate that the assembly of proteins imported by the carrier translocase, TIM22 complex, is even more strongly affected by the lack of Tam41. Moreover, respiratory chain supercomplexes and the inner membrane potential are impaired by lack of Tam41. The phenotype of Tam41-deficient mitochondria thus resembles that of mitochondria lacking cardiolipin. Indeed, we found that Tam41 is required for the biosynthesis of the dimeric phospholipid cardiolipin. The pleiotropic effects of the translocator maintenance protein on preprotein import and respiratory chain can be attributed to its role in biosynthesis of mitochondrial cardiolipin

Mutations in the CDP-Choline Pathway for Phospholipid Biosynthesis Bypass the Requirement for an Essential Phospholipid Transfer Protein

SEC14p is the yeast phosphatidylinositol (PI)/phosphatidylcholine (PC) transfer protein, and it effects an essential stimulation of yeast Golgi secretory function. We now report that the SEC14p localizes to the yeast Golgi and that the SEC14p requirement can be specifically and efficiently bypassed by mutations in any one of at least six genes. One of these suppressor genes was the structural gene for yeast choline kinase (CKI), disruption of which rendered the cell independent of the normally essential SEC14p requirement. The antagonistic action of the CKI gene product on SEC14p function revealed a previously unsuspected influence of biosynthetic activities of the CDP-choline pathway for PC biosynthesis on yeast Golgi function and indicated that SEC14p controls the phospholipid content of yeast Golgi membranes in vivo