B31D-01 INVITED 08:00h
Biosynthetic Pathways, Gene Replacement and the Antiquity of Life
Our understanding of the evolution and diversity of life during the Archean has been profoundly influenced by geological signatures including microfossils, stromatolites, and fractionated carbon and sulfur isotopes. These biosignatures have been extensively debated on the grounds of competing abiotic processes. Hopane and squalene biomarkers have been taken as evidence that by 2700 Ma, all three domains of life had emerged, that cyanobacteria, the `inventors' of oxygenic photosynthesis, had diverged from their bacterial ancestors, heralding the impending oxidation of Earth's atmosphere, and that O2 was already available, as it is required explicitly in modern organisms for the synthesis of squalene. We argue that, based on corollaries present throughout enzymology, the oxidization of squalene during the Archean could just as likely have been carried out by an anaerobic enzyme that either remains to be characterized or that may have been totally lost as eukaryotes assumed primarily aerobic lifestyles. The transition from an anoxic to an oxic world had unfathomable consequences for early life; molecular oxygen and its derivatives went from being poisonous in an obligately anaerobic world, to assuming a central role in the subsequent development of complex, macroscopic life. Though the aftermath was global in scale, the biological transition to aerobiosis was effected at the molecular level, which included the evolution of a wealth of new enzymes, the extensive modification of many existing enzymes, and the `rewiring' of central biochemical pathways.
B31D-02 08:20h
Can geological systems be used to calibrate rates of microbial genome evolution?
An important challenge is to develop absolute time calibrations for molecular-level evolutionary processes. This is especially difficult for free-living microorganisms, which lack an interpretable fossil record. Absolute rates of genome change would enable connection of (for example) environmental perturbations documented in the geologic record to organismal responses. Thus, the stimuli for speciation and mechanisms of speciation could be directly linked. It may be possible to apply comprehensive environmental genomic studies in carefully chosen geological settings in order to tackle this problem. For example, Tyson et al. (2004) assembled sequence data from a low diversity biofilm community from an acidic ecosystem to reconstruct near complete and partial genomes for the dominant strain populations. As sequence data from many individuals were assembled into each composite genome, the dataset provides information about strain-level diversity. Ongoing growth in sequencing capability makes application of this approach to increasingly complex ecosystems feasible. Comparisons among groups of genomes from organisms separated by different evolutionary distances can reveal information about the relative rates at which different forms of genomic change occur. For example, the number of changes in gene content and gene order in the genomes of members of a strain population may be small, but the placement and types of mobile constituents (e.g., transposases and prophage) may differ significantly. Each type of genome change can be quantified (e.g., number of nucleotide polymorphisms, amino acid changes, gene duplications, gene loss/gain, gene order shuffling). These data also can be collected for organisms from the same lineage that diverged earlier (e.g., for species or genera). Statistics for each type of genome change yield semi-independent measures of evolutionary distance or relative rates of genomic change. Absolute time calibration is all that is needed to convert these data to real rates. Genomic studies could be carried out in geologic systems for which a date of onset of an environmental change conducive to colonization (e.g., the appearance of a hot spring or onset of acidification) can be determined. If it is assumed that at many of the current microorganisms descended from the colonists, then evolutionary rates can be estimated. Such analyses could be conducted at multiple sites of the same type to verify findings. Comparisons involving geographically separated sites would also provide information about the rates and pathways of microbial dispersal. The coupling of genomics with geochronology appears to be a logical extension of palaeontological studies, and should advance understanding of biological evolution over Earth history.
B31D-03 08:40h
On the Significance of Bacterial triterpenic Biomarkers in Sediments
Triterpenic biomarkers are ubiquitous in the organic matter of sediments. Bacterial contribution is essential for several series. Despite the numerous investigations performed over the last decades, little is known about the distribution of triterpenoids in Eubacteria. An updated survey of triterpene distribution in Eubacteria points out a much broader diversity of the structures than expected ten years ago. Hopanoids characterized by their C35 skeleton resulting from a carbon/carbon linkage between the triterpene hopane skeleton and a D-ribose derivative are the most frequent ones. Their distribution cannot be readily interpreted and may result from lateral gene transfer. Many groups, such as strict anaerobes, are underrepresented in the screenings, mainly because of the complex techniques required for their growth. Most of the bacterial hopanoids belong to the (17$\alpha$H,21$\beta$H) series, corresponding to the stereochemistry of hopanoid biomarkers from non-mature sediments. (17$\beta$H,21$\alpha$H)- and especially (17$\alpha$,21$\beta$H)-hopanoids are derived from the former series via diagenesis and maturation of the organic matter. Both series were, however, recently found in widespread soil bacteria ({\it Frankia} spp., {\it Geodermatophilus} spp.) questioning at least partially their significance as maturation indicators. Quasi-hopanoids with the gammacerane skeleton were first found in ciliate protozoa. They are also present in high concentrations in the phylogenetically related bacteria {\it Rhodopseudomonas palustris} and all {\it Bradyrhizobium} spp. In all closely investigated hopanoid producing bacteria, a complex mixture of triterpene hydrocarbons accompanied in small amounts hop-22(29)-ene. They include pentacyclic triterpenes (rearranged hopenes, fernenes) as well as tetracyclic triterpenes (dammaradienes, euphadienes) and result from a lack of strict control of the cyclization process by the squalene/hopene cyclase. Triterpenoids related to sterol biosynthesis (lanosterol, cycloartenol) have been found in a few scattered taxa. Their contribution to the sediment biomarkers is not obvious. Finally, the reactions leading from biohopanoids to the geohopanoids are poorly known. Only evidence for abiotic degradation rather than for biological degradation is available.
B31D-04 INVITED 09:00h
Search for the Evolution of Steroid Biosynthesis in the Geological Record
To study the evolution of the structure of organisms we can directly examine fossilized shells, skeletons and petrified cells. In contrast, for the tentative reconstruction of the phylogeny of biosynthetic pathways, such as steroid anabolism, we rely entirely on the comparative molecular biology of living organisms. Thus, without fossil evidence, the times in geological history when successive steps of a metabolic pathway evolved remain particularly elusive. Molecular clocks of genes coding for the enzymes involved in a biosynthetic pathway might provide a rough guess when a natural product first appeared in geological time, but they are intrinsically unreliable without calibration points in the distant past. However, it might be possible to trace the evolutionary history of some biosynthetic pathways directly in the geological record by searching for hydrocarbon biomarkers of anabolic intermediates. Biomarkers are molecular fossils of natural products. They often retain the diagnostic carbon skeleton of their biological precursor and remain stable over hundreds of millions of years enclosed in organic-rich sedimentary rocks. Sterane hydrocarbons are particularly abundant biomarkers and potentially suitable for the search of biosynthetic intermediates. Steranes are the fossil equivalents of functionalized steroids found in eukaryotes and certain bacteria. The biosynthesis of typical eukaryotic steroids such as cholesterol (C$_{27}$), ergosterol (C$_{28}$) and sitosterol (C$_{29}$) from the acyclic precursor squalene (C$_{30}$) involves more than 20 enzymatic steps. The most crucial steps include modification of the carbon skeleton by removal of several methyl groups from the ring system and addition of alkyl groups to the steroid side chain. The evolution of this complex pathway must have occurred over geologically significant periods of time and likely involved several preadaptive intermediates that represented structurally less derived but fully functional lipids. Thus, if a molecular corollary of `ontogeny recapitulates phylogeny' applies, it might be possible to detect a sequence of increasingly modified fossil steroids in the geological record and to create a time frame for the evolution of this fundamental biosynthetic pathway. Here we present first results of an extensive search for the fossil remains of evolutionary intermediate steroids in sedimentary successions of Precambrian age.
B31D-05 09:15h
The Evolution of Sterol Biosynthesis in Bacteria: In Situ Fluorescence Localization of Sterols in the Nucleoid Bacterium {\it Gemmata obscuriglobus}
The biosynthesis of sterols is generally regarded as a eukaryotic process. The first enzymatic step in the production of sterols requires molecular oxygen. Therefore, both the origin of eukaryotes and the evolution of sterol biosynthesis were thought to postdate the rise of oxygen in earth's atmosphere, until Brocks et al. discovered steranes in rocks aged 2.7 Ga (1). Many prokaryotes produce hopanoids, sterol-like compounds that are synthesized from the common precursor squalene without the use of molecular oxygen. However, a few bacterial taxa are also known to produce sterols, suggesting this pathway could precede the rise of oxygen (2, 3). Recently, we discovered the shortest sterol-producing biosynthetic pathway known to date in the bacterium {\it Gemmata obscuriglobus} (4). Using genomic searches, we found that {\it Gemmata} has the enzymes necessary for synthesis of sterols, and lipid analyses showed that the sterols produced are lanosterol and its isomer parkeol. {\it Gemmata} is a member of the Planctomycetes, an unusual group of bacteria, all of the known species of which contain intracellular compartmentalization. Among the Planctomycetes, {\it Gemmata} uniquely is the only prokaryote known to contain a double-membrane-bounded nuclear body (5). Since sterols usually are found in eukaryotes, and {\it Gemmata} has a eukaryote-like nuclear organelle, we investigated the location of the sterols within {\it Gemmata} to postulate whether they play a role in stabilization of the nuclear membrane and control of genomic organization. We used the sterol-specific fluorescent dye Filipin III in conjunction with fluorescent dyes for internal and external cellular membranes in order to determine whether the sterols are located in the nuclear body membrane, external membrane, or both. We found that sterols in {\it Gemmata} are concentrated in the internal membrane, implying that they function in maintaining this unusual cellular component. It is notable that {\it Gemmata} also produce hopanoids, suggesting that they acquired the ability to produce sterols for a specialized function related to their nuclear membrane. 1. Brocks, J.J., et al., Science 285:1033-36 (1999). 2. Bird, C.W., et al., Nature 230:473-74 (1971). 3. Bode, H.B., et al., Mol. Microbiol. 47:471-81 (2003). 4. Pearson, A., et al., Proc. Natl. Acad. Sci. USA 100:15352-57 (2003). 5. Fuerst, J.A. and R.I. Webb, Proc. Natl. Acad. Sci. USA 88:8184-88 (1991).
B31D-06 09:30h
Reconstruction Of Ancient Food-Chain Dynamics From the Isotopic Composition of Molecular Fossils
Changes in the isotopic compositions of bulk sedimentary carbon are commonly used to infer past changes in the global carbon cycle. When both inorganic and organic carbon are analyzed, coevolving changes have revealed aspects of the dynamical interactions between these two bulk reservoirs. Of at least as much interest, however, are the interactions {\it within} the organic reservoir. Here we show how such interactions can be inferred from the changing isotopic compositions of specific organic compounds (i.e., ``molecular fossils'' or ``biomarkers''). Previous work has identified particular compounds whose isotopic compositions covary with primary (i.e., photosynthetically derived) and secondary (i.e., not primary) organic carbon as a whole. We model the evolution of these isotopic compositions as carbon flow on an elementary food chain. Two results are obtained. First, the prediction of a particular steady-state representation of the data provides a method for estimating the relative burial fluxes of primary and secondary sedimentary organic carbon. Second, the temporal fluctuations of the difference between primary and secondary isotopic compositions is related to community-level metabolism. Changes in the accumulation of such ``metabolic isotope effects'' have previously been related to changing food-chain lengths. A quantitative analysis suggests, however, that changes in the metabolic isotope effect itself---due, for example, to changing oxygen concentrations---provide a more plausible cause of such fluctuations. Preliminary studies of biomarkers traversing the Permian-Triassic extinction and the Cenomanian-Turonian oceanic anoxic event appear to confirm these predictions.
B31D-07 09:45h
Ancient DNA, climatic change, and loss of genetic diversity in an endemic Patagonian mammal
Understanding the response of animal populations to climatic change is essential for the future maintenance of biodiversity. One question that remains difficult to answer, and is particularly important to conservation, is how animals respond over time scales relevant to evolutionary change. Ancient DNA provides a unique opportunity to track animal response to Holocene climate change and to study species replacement patterns and genetic diversity over time. We used ancient DNA to compare response to climatic change in two species, C. sociabilis and C. haigi, over the last 8,000 years. Our study site, Cueva Traful, is a late-Holocene raptor roost in Parque Nacional Nahuel Huapi, Argentina. A lack of genetic diversity in modern C. sociabilis populations is indicative of past bottleneck events and a previous ancient DNA study found that it had remained genetically identical for at least 1000 years in the face of climatic change and human disturbance. Since Cueva Traful goes back further in time, our first goal was to examine genetic diversity in order to place a longer term historical perspective on the modern bottleneck. The second goal was to compare changes in genetic diversity in C. sociabilis to C. haigi a closely related species that may respond differently to climatic change. The use of ancient DNA presents unique challenges due to low copy number, environmental damage to template, and high contamination risk. Despite these challenges, ancient DNA provides a unique perspective on evolutionary history.