Lipid remodeling in Rhodopseudomonas palustris TIE‐1 upon loss of hopanoids and hopanoid methylation

Abstract

The sedimentary record of molecular fossils (biomarkers) can potentially provide important insights into the composition of ancient organisms; however, it only captures a small portion of their original lipid content. To interpret what remains, it is important to consider the potential for functional overlap between different lipids in living cells, and how the presence of one type might impact the abundance of another. Hopanoids are a diverse class of steroid analogs made by bacteria and found in soils, sediments, and sedimentary rocks. Here, we examine the trade-off between hopanoid production and that of other membrane lipids. We compare lipidomes of the metabolically versatile α-proteobacterium Rhodopseudomonas palustris TIE-1 and two hopanoid mutants, detecting native hopanoids simultaneously with other types of polar lipids by electrospray ionization mass spectrometry. In all strains, the phospholipids contain high levels of unsaturated fatty acids (often >80%). The degree to which unsaturated fatty acids are modified to cyclopropyl fatty acids varies by phospholipid class. Deletion of the capacity for hopanoid production is accompanied by substantive changes to the lipidome, including a several-fold rise of cardiolipins. Deletion of the ability to make methylated hopanoids has a more subtle effect; however, under photoautotrophic growth conditions, tetrahymanols are upregulated twofold. Together, these results illustrate that the 'lipid fingerprint' produced by a micro-organism can vary depending on the growth condition or loss of single genes, reminding us that the absence of a biomarker does not necessarily imply the absence of a particular source organism.

Hopanoid lipids: from membranes to plant–bacteria interactions

“…In the purple non-sulfur bacterium Rhodopseudomonas palustris TIE-1, membranes from the Δ hpnP mutant, which lacks all 2Me-hopanoids, are equally as fluid as wild-type membranes 53 . It has been shown that hopanoid mutants overproduce other membrane-condensing lipids 68–70 , which may explain this discrepancy. Regardless, factors affecting membrane fluidity in vivo are clearly more complex than what can be captured in vitro.…”

A distinct pathway for tetrahymanol synthesis in bacteria

“…Although a direct correlation between sterol starvation and tetrahymanol production has been established in ciliates (6), the physiological role of tetrahymanol in bacteria is unknown. Recent studies have highlighted increased tetrahymanol production in R. palustris TIE-1 under certain physiological conditions (e.g., photoautotrophic growth) and also when cellular hopanoid lipid profiles are altered in gene deletion mutants (22,23), but the physiological significance of these changes is not known. Further, the biochemical mechanism of tetrahymanol synthesis in bacteria is unclear.…”

Towards measuring growth rates of pathogens during infections by D2O‐labeling lipidomics

“…The introduced heterogeneity causes broadening of chromatographic peaks, which could skew the isotope ratio observed by LC/MS as ionization efficiency varies over time . In order to evaluate how this affects isotope quantification by lipidomics, we grew an E. coli culture in 4% D 2 O (fractional D‐abundance, 2 F WATER ) and measured lipids after chromatographic separation using an electrospray ionization time‐of‐flight (ESI‐TOF) mass spectrometer …”

“…LC/MS data were collected on an Acquity I‐Class UPLC coupled to a Xevo G2‐S TOF mass spectrometer (Waters). Intact polar lipids were separated on an Acquity UPLC CSH C18 column (2.1 mm × 100 mm, 1.7 μm; Waters) at 55°C following a protocol established by Waters Corporation and adapted in our laboratory . Samples were run in three randomized instrument replicates (injection volume 5 μL).…”