Extracellular and Intracellular Lanthanide Accumulation in the Methylotrophic Beijerinckiaceae Bacterium RH AL1

Abstract

Recent work in Methylorubrum extorquens AM1 identified intracellular, cytoplasmic lanthanide storage in an organism that harnesses these metals for its metabolism. Here, we describe the extracellular and intracellular accumulation of lanthanides in Beijerinckiaceae bacterium RH AL1, a newly isolated and recently characterized methylotroph. Using ultrathin-section transmission electron microscopy (TEM), freeze-fracture TEM (FFTEM), and energy-dispersive X-ray spectroscopy, we demonstrated that strain RH AL1 accumulates lanthanides extracellularly at outer membrane vesicles (OMVs) and stores them in the periplasm. High-resolution elemental analyses of biomass samples revealed that strain RH AL1 can accumulate ions of different lanthanide species with a preference for heavier lanthanides. Its methanol oxidation machinery is supposably adapted to light lanthanides, and their selective uptake is mediated by dedicated uptake mechanisms. Based on RNAseq analysis, these presumably include the previously characterized TonB-ABC transport system encoded by the lut -cluster, but eventually also a type VI secretion system. A high constitutive expression of genes coding for lanthanide-dependent enzymes suggested that strain RH AL1 maintains a stable transcript pool to flexibly respond to changing lanthanide availability. Genes coding for lanthanide-dependent enzymes are broadly distributed taxonomically. Our results support the hypothesis that central aspects of lanthanide-dependent metabolism partially differ between the various taxa. Importance: Although multiple pieces of evidence have been added to the puzzle of lanthanide-dependent metabolism, we are still far from understanding the physiological role of lanthanides. Given how widespread lanthanide-dependent enzymes are, only limited information is available with respect to how lanthanides are taken up and stored in an organism. Our research complements work in commonly studied model organisms and showed the localized storage of lanthanides in the periplasm. This storage occurred at comparably low concentrations. Strain RH AL1 is able to accumulate lanthanide ions extracellularly and to selectively utilize lighter lanthanides. Beijerinckiaceae bacterium RH AL1 might be an attractive target for developing biorecovery strategies to win these economically highly demanded metals in more environmentally friendly ways.

A Periplasmic Lanthanide Mediator, Lanmodulin, in Methylobacterium aquaticum Strain 22A

“…Typical ones in ΔlanM and lanM-OX are shown. Similar deposits have been also observed in M. extorquens strain AM1 lut gene mutants (Roszczenko-Jasińska et al, 2020) and Beijerinckiaceae bacterium RH AL1 (Wegner et al, 2021).…”

“…It has been suggested that the protein shuttles Ln to Ln 3+ -dependent enzymes (Cotruvo et al, 2018;Cook et al, 2019). LanM also responds to Ln 3+ in non-methylotrophy settings in a methylotrophic Beijerinckiaceae bacterium RH CH11 (Wegner et al, 2021). lanM is associated with other lut genes in strain AM1 and PA1 genomes, but it is associated with the genes of different functions in strain 22A (Supplementary Figure S2).…”

“…Regulation of MV synthesis and selection of proteins into MV cargo are now an emerging field of microbiological studies ( Toyofuku et al, 2018 ). MV formation has been suggested as one of the Ln uptake systems in the methylotrophic Beijerinckiaceae bacterium RH AL1 ( Wegner et al, 2021 ). MV is also reported to be involved in the excretion of excess copper ( Lima et al, 2022 ), suggesting that MV formation is one of the cellular strategies to excrete excess metals to maintain metal homeostasis.…”

“…It has been suggested that the protein shuttles Ln to Ln 3+ -dependent enzymes ( Cotruvo et al, 2018 ; Cook et al, 2019 ). LanM also responds to Ln 3+ in non-methylotrophy settings in a methylotrophic Beijerinckiaceae bacterium RH CH11 ( Wegner et al, 2021 ).…”

Lanpepsy is a novel lanthanide-binding protein involved in the lanthanide response of the obligate methylotroph Methylobacillus flagellatus

“…Intriguingly, crystalline lanthanum (La 3+ ) phosphate deposits were observed in the cytoplasm of M. extorquens by electron microscopy coupled with energy-dispersive X-ray spectroscopy ( 16 ). Extracellular (associated with outer membrane vesicles) as well as periplasmic deposits have also been described for the methylotroph Beijerinckiaceae bacterium RH AL1 ( 19 ). The exact composition, assembly, and function of these inclusions remain unclear.…”