Prokaryotic Hopanoids and other Polyterpenoid Sterol Surrogates

We have discovered a large and highly conserved RNA motif that typically resides in a noncoding section of a multigene messenger RNA in extremophilic Gram-positive eubacteria. RNAs of this class adopt an ornate secondary structure, are large compared with most other noncoding RNAs, and have been identified only in certain extremophilic bacteria. These ornate, large, extremophilic (OLE) RNAs have a length of Ϸ610 nucleotides, and the 35 representatives examined exhibit extraordinary conservation of nucleotide sequence and base pairing. Structural probing of the OLE RNA from Bacillus halodurans corroborates a complex secondary structure model predicted from comparative sequence analysis. The patterns of structural conservation, and its unique phylogenetic distribution, suggest that OLE RNA carries out a complex and critical function only in certain extremophilic bacteria.isoprenoid ͉ riboswitch ͉ ribozyme ͉ superoperon I n recent years, a large number of small noncoding RNAs (1-3) have been identified in eubacteria (4), archaea (5), and eukaryotes (6). In many instances, novel noncoding RNAs in bacteria have been discovered by using bioinformatics search strategies (e.g., refs. 7-13) that take advantage of the conserved nucleotide sequences and secondary structures of these functional RNAs. Another productive way to identify numerous noncoding RNAs from bacteria is by cloning and sequencing small RNAs isolated from cell extracts (e.g., refs. 13-16), and this approach is useful particularly for those examples that are not well conserved through evolution.RNAs such as tRNAs, rRNAs, and some ribozymes have noncoding functions that have long been known to be central to RNA processing and protein synthesis mechanisms. However, it seems possible that additional noncoding RNAs will be found that perform fundamental biochemical tasks that to date have been assumed to be the exclusive province of protein factors. A number of noncoding RNAs discovered recently have proven to participate as the key components of gene regulation systems (e.g., refs. 17-19). Additional examples of widespread noncoding RNAs in bacteria, such as 6S RNA (20-22), the dual-function tmRNA (23), and noncoding portions of messenger RNAs such as T-Box elements (24) and riboswitches (25-27) perform important gene control and molecular sensing tasks that are critical for cells to function normally. The existence of so many RNAs with atypical functions implies that some newly discovered noncoding RNAs might perform surprising and important roles in fundamental cellular processes. In this article, we describe features of a noncoding RNA element whose size, structural sophistication, and unique phylogenetic distribution are suggestive of a complex biological function. ResultsIdentifying Ornate, Large, Extremophilic (OLE) RNAs by Using Bioinformatics. We have previously used computer-aided search strategies that employ comparative sequence analysis (e.g., refs. 11 and 28-30) to identify noncoding RNAs whose nucleotide sequences and secondary structures are c...

Section: Discussionmentioning

confidence: 99%

Identification of a large noncoding RNA in extremophilic eubacteria

Puerta-Fernández

Barrick

Roth

et al. 2006

“…It is believed that OSC evolved from the hopanoid pathway predecessor, bacterial squalenehopene cyclase (SHC) (8,9). In eukaryotes, the initial sterols lanosterol and cycloartenol are merely biosynthetic intermediates; i.e., up to 20 additional steps are required for downstream conversion of lanosterol to the animal product, cholesterol.…”

Section: S Terol Biosynthesis Is Nearly Ubiquitous Among Eukaryotes;mentioning

confidence: 99%

Phylogenetic and biochemical evidence for sterol synthesis in the bacterium Gemmata obscuriglobus

Pearson

Budin

Brocks

2003

233

189

Sterol biosynthesis is viewed primarily as a eukaryotic process, and the frequency of its occurrence in bacteria has long been a subject of controversy. Two enzymes, squalene monooxygenase and oxidosqualene cyclase, are the minimum necessary for initial biosynthesis of sterols from squalene. In this work, 19 protein gene sequences for eukaryotic squalene monooxygenase and 12 protein gene sequences for eukaryotic oxidosqualene cyclase were compared with all available complete and partial prokaryotic genomes. The only unequivocal matches for a sterol biosynthetic pathway were in the proteobacterium, Methylococcus capsulatus, in which sterol biosynthesis is known, and in the planctomycete, Gemmata obscuriglobus. The latter species contains the most abbreviated sterol pathway yet identified in any organism. Analysis shows that the major sterols in Gemmata are lanosterol and its uncommon isomer, parkeol. There are no subsequent modifications of these products. In bacteria, the sterol biosynthesis genes occupy a contiguous coding region and possibly comprise a single operon. Phylogenetic trees constructed for both enzymes show that the sterol pathway in bacteria and eukaryotes has a common ancestry. It is likely that this contiguous reading frame was exchanged between bacteria and early eukaryotes via lateral gene transfer or endosymbiotic events. The primitive sterols produced by Gemmata suggest that this genus could retain the most ancient remnants of the sterol biosynthetic pathway. S terol biosynthesis is nearly ubiquitous among eukaryotes; conversely, it is almost completely absent in prokaryotes (1). As a result, the presence of diverse steranes in ancient rocks is used as evidence for eukaryotic evolution Ͼ2.7 billion years ago (2). However, the occasional presence of sterols in prokaryotes is poorly understood. Sterol production by bacteria previously has been demonstrated only in the Methylococcales (3, 4) and Myxobacteriales (5,6).Understanding the evolution of sterol biosynthesis is of significant interest to biochemistry, evolutionary biology, and the geosciences, because the only known biosynthetic pathway requires molecular oxygen. The first step in this pathway is the epoxidation of the hydrocarbon squalene, in which the addition of 1 ⁄2O 2 is catalyzed by the enzyme squalene monooxygenase (SQMO) (7). Unless there are other unknown enzymes or abiogenic reactions capable of producing squalene epoxide, this would require the prior evolution of oxygenic photosynthesis. For sterol biosynthesis to date to the last common ancestor, a biogenic or abiogenic peroxidation could be a potential mechanism, although this has not yet been demonstrated.Cyclization of squalene epoxide to form the initial sterol proceeds immediately through the action of a second enzyme, oxidosqualene cyclase (OSC). It is believed that OSC evolved from the hopanoid pathway predecessor, bacterial squalenehopene cyclase (SHC) (8, 9). In eukaryotes, the initial sterols lanosterol and cycloartenol are merely biosynthetic intermediates; i.e.,...

“…Permeability changes were monitored by measuring the swelling rates of large unilamellar vesicles (LUVs) following an osmotic shock in a stopped-flow spectrophotometer. This method had been developed to investigate the role of some prokaryotic terpenoids as membrane reinforcers (4)(5)(6)(7)(8). The kinetics of swelling was shown to be controlled by the water permeability of bilayers and to depend on the lipid composition of vesicles (5).…”

mentioning

confidence: 99%

Differential effects of plant sterols on water permeability and on acyl chain ordering of soybean phosphatidylcholine bilayers.

Schuler

Milon

Nakatani

et al. 1991

Self Cite

210

150

To gain some insight into the structural and functional roles of sterols in higher plant cells, various plant sterols have been incorporated into soybean phosphatidyicholine (PtdCho) bilayers and tested for their ability to regulate water permeability and acyl chain ordering. Sitosterol was the most efficient sterol in reducing the water permeability of these vesicles and stigmasterol appeared to have no significant effect.Vesicles containing 24fmethylcholesterol exhibited an intermediate behavior, similar to that of vesicles containing cholesterol. Cycloartenol, the first cyclic biosynthetic precursor of plant sterols, reduced the water permeability in a very effective way. Oftwo unusual plant sterols, 246-methylpollinastanol and 14a,24Udimethylcholest-8-en-3fi-ol, the former was found to be functionally equivalent to sitosterol and the latter was found to be relatively inefficient. 2H NMR experiments have been performed with oriented bilayers consisting of soybean PtdCho with sitosterol, stigmasterol, or 24-methylpoll nastanol. The results provided clear evidence that sitosterol and 24k-methylpolinastanol exhibit a high efficiency to order PtdCho acyl chains that closely parallels their ability to reduce water permeability. By contrast, stigmasterol shows a low efficiency for both functions. These results show that sitosterol and stigmasterol, two major 24-ethylsterols differing only by the absence or presence of the A22 double bond in the side chain, probably play different roles in regulating plant membrane properties; they also may explain why 9P,19-cyclopropylsterols behave as good surrogates of sitosterol.In mammalian cells, the bulk of free cholesterol is localized in the plasma membrane where it participates in the regulation of membrane fluidity and in the activity of many membrane-bound enzymes (1). Changes in cholesterol content have been shown to induce modifications in cellular functions, which depend on the dynamics of the cell-surface membrane such as the recognition of an external signal (1). In higher plant cells, the three typical phytosterols: sitosterol, and 24F-methylcholesterol [the mixture of 24R (i.e., campesterol) and 24S (i.e., 22-dihydrobrassicasterol) epimers] are also mainly concentrated in the plasma membrane (2). To investigate whether the different plant sterols play similar roles in higher plant cells, we started a study with well-defined model membrane systems prepared from soybean phosphatidylcholine (PtdCho), a major representative plant phospholipid, and various sterols in different molar ratios. Because of their high content in polyunsaturated fatty acyl chains (3), soybean PtdCho bilayers can be considered a valuable model of higher plant membrane. The present work deals with the effect of some plant sterols on water permeability and acyl chain ordering of these bilayers. Permeability changes were monitored by measuring the swelling rates of large unilamellar vesicles (LUVs) following an osmotic shock in a stopped-flow spectrophotometer. This method had been develo...