Abstract
Hopanoids are sterol-like membrane lipids widely used as geochemical proxies for bacteria. Currently, the physiological role of hopanoids is not well understood, and this represents one of the major limitations in interpreting the significance of their presence in ancient or contemporary sediments. Previous analyses of mutants lacking hopanoids in a range of bacteria have revealed a range of phenotypes under normal growth conditions, but with most having at least an increased sensitivity to toxins and osmotic stress. We employed hopanoid-free strains of Methylobacterium extorquens DM4, uncovering severe growth defects relative to the wild-type under many tested conditions, including normal growth conditions without additional stressors. Mutants overproduce carotenoids–the other major isoprenoid product of this strain–and show an altered fatty acid profile, pronounced flocculation in liquid media, and lower growth yields than for the wild-type strain. The flocculation phenotype can be mitigated by addition of cellulase to the medium, suggesting a link between the function of hopanoids and the secretion of cellulose in M. extorquens DM4. On solid media, colonies of the hopanoid-free mutant strain were smaller than wild-type, and were more sensitive to osmotic or pH stress, as well as to a variety of toxins. The results for M. extorquens DM4 are consistent with the hypothesis that hopanoids are important for membrane fluidity and lipid packing, but also indicate that the specific physiological processes that require hopanoids vary across bacterial lineages. Our work provides further support to emerging observations that the role of hopanoids in membrane robustness and barrier function may be important across lineages, possibly mediated through an interaction with lipid A in the outer membrane.
Highlights
Hopanoids are pentacyclic triterpenoids present in some bacteria that are ubiquitous in sediments and sedimentary rocks
A permeability role for hopanoids was inferred in the alphaproteobacterium Zymomonas mobilis, which displayed an increased sensitivity to ethanol concentrations when hopanoid production was diminished by the addition of azasqualene, which inhibits the key enzyme squalene-hopene cyclase (SHC) that catalyzes the synthesis of hopanoids [7]
As a first stage in characterization of the M. extorquens DM4 shc mutant, its lipid content was compared to WT
Summary
Hopanoids are pentacyclic triterpenoids present in some bacteria that are ubiquitous in sediments and sedimentary rocks. These types of molecules are among the best documented organic geochemical biomarkers in the rock record, and have been detected in rocks at least 1.7 billion years old [1]. In the gammaproteobacterium Frateuria aurantia, hopanoid production increased with growth temperature [8]. In the Firmicute Alicyclobacillus (formerly Bacillus) acidocaldarius, the abundance of side-chain containing hopanoids is positively correlated with growth at increased temperature or decreased pH [9]. The authors of the latter study hypothesized that hopanoids help overcome the expected increased permeability and decreased stability of biological membranes under these conditions
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