Hopanoid-free Methylobacterium extorquens DM4 overproduces carotenoids and has widespread growth impairment

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.

Dichloromethane Degradation Pathway from Unsequenced Hyphomicrobium sp. MC8b Rapidly Explored by Pan-Proteomics

“…On the one hand, two of the proteins detected as differentially abundant in both DM4 and MC8b proteomics studies had already been associated with DCM metabolism. The squalene hopene cyclase shc gene was identified as essential for growth with DCM [47,48], while a transglycosylase/transpeptidase homolog (HYPMC8B_3734, annotated as penicillin-binding protein; METDI4661 in strain DM4) showed DCM-dependent synthesis [47]. These findings confirm the likely importance of these two proteins as part of a specific ensemble of proteins associated with bacterial growth on DCM and involving DNA repair and envelope processes ( Supplementary Table S5).…”

A comparative transcriptome analysis of the novel obligate methanotroph Methylomonas sp. DH-1 reveals key differences in transcriptional responses in C1 and secondary metabolite pathways during growth on methane and methanol

“…2). The function of hopanoids has been characterized in several organisms, including methylotrophic bacteria [62, 63]. A lack of hopanoid biosynthesis increases sensitivity against toxins and osmotic stress.…”

“…The role of hopanoids in maintaining membrane robustness and membrane barrier function is likely conserved across bacterial lineages. This function is possibly mediated through an interaction with lipid A in the outer membrane of Methylobacterium extorquens DM4 [63]. In addition, membrane function in the hopanoid-free Methylobacterium extorquens PA1 was lower [62].…”

Extended hopanoid loss reduces bacterial motility and surface attachment and leads to heterogeneity in root nodule growth kinetics in a Bradyrhizobium-Aeschynomene symbiosis

“…Populations within ΔhpnH also may differ in the extent to which extended hopanoid loss is compensated. In Methylobacterium extorquens and Rhodopseudomonas palustris 41,42 , hopanoid loss results in upregulation of other membrane-rigidifying lipids including carotenoids and cardiolipins, and in other plant-microbe systems, lipid exchange between hosts and microbes has been observed 43 , suggesting that ΔhpnH mutant success at a given nodulation site may relate to the local availability of exogenous plant steroids or structurally similar metabolites.…”