Connectivity of the intracytoplasmic membrane of Rhodobacter sphaeroides: a functional approach

Abstract

The photosynthetic apparatus in the bacterium Rhodobacter sphaeroides is mostly present in intracytoplasmic membrane invaginations. It has long been debated whether these invaginations remain in topological continuity with the cytoplasmic membrane, or form isolated chromatophore vesicles. This issue is revisited here by functional approaches. The ionophore gramicidin was used as a probe of the relative size of the electro-osmotic units in isolated chromatophores, spheroplasts, or intact cells. The decay of the membrane potential was monitored from the electrochromic shift of carotenoids. The half-time of the decay induced by a single channel in intact cells was about 6 ms, thus three orders of magnitude slower than in isolated chromatophores. In spheroplasts obtained by lysis of the cell wall, the single channel decay was still slower (~23 ms) and the sensitivity toward the gramicidin concentration was enhanced 1,000-fold with respect to isolated chromatophores. These results indicate that the area of the functional membrane in cells or spheroplasts is about three orders of magnitude larger than that of isolated chromatophores. Intracytoplasmic vesicles, if present, could contribute to at most 10% of the photosynthetic apparatus in intact cells of Rba. sphaeroides. Similar conclusions were obtained from the effect of a ∆pH-induced diffusion potential in intact cells. This caused a large electrochromic response of carotenoids, of similar amplitude as the light-induced change, indicating that most of the system is sensitive to a pH change of the external medium. A single internal membrane and periplasmic space may offer significant advantages concerning renewal of the photosynthetic apparatus and reallocation of the components shared with other bioenergetic pathways.

Membrane Dynamics in Phototrophic Bacteria

“…Some ICMs exist as discrete vesicles without any physical associations with other structures, supporting the discontinuity model (25,32). By contrast, cryo-ET of R. sphaeroides Ga revealed no connections between ICMs and the plasma membrane, and most ICM vesicles are fused to neighboring vesicles rather than being discrete vesicular structures (92), in support of the continuous model (112). The connectivity of adjacent ICM vesicles results in formation of a continuous membrane reticulum that encloses a single lumenal space (92).…”

“…The connectivity of adjacent ICM vesicles results in formation of a continuous membrane reticulum that encloses a single lumenal space (92). This large-scale ICM network may provide physiological advantages in molecular trafficking and environmental adaptation (112). Despite the possible strain variation, cryo-ET imaging of R. sphaeroides DSM 158 further showed that ∼22% of ICM vesicles are unconnected, with the majority forming connections with neighboring vesicles (70).…”

Ectopic Neo-Formed Intracellular Membranes in Escherichia coli: A Response to Membrane Protein-Induced Stress Involving Membrane Curvature and Domains

“…In contrast to eukaryotic cells, most bacteria are devoid of any internal membranes since they do not have any organelles (except purple photosynthetic bacteria, such as Rhodobacter sphaeroides [ 12 ]), and do not display internal membrane trafficking such as endocytosis. However, the bacterial cell membrane (single in Gram-positive or double in Gram-negative bacteria) is not that simple [ 13 ].…”

Connectivity of centermost chromatophores in Rhodobacter sphaeroides bacteria

“…Decades of experimental effort have offered two models concerning chromatophore continuity in R. sphaeroides. The continuous model asserts that chromatophores in photosynthetic bacteria form a membrane continuum with both the cytoplasmic membrane (CM) and neighboring vesicles (Cohen-Bazire and Kunisawa, 1963;Boatman, 1964;Holt and Marr, 1965;Tauschel and Drews, 1967;Prince et al, 1975;Holmqvist, 1979;Scheuring et al, 2014;Verméglio et al, 2016). However, there is also ultrastructural evidence to support the claim that chromatophores in phototrophic bacteria manifest as naturally discrete structures in the cytoplasmic space (Vatter and Wolfe, 1958;Geyer and Helms, 2006a;Tucker et al, 2010).…”

“…Investigating chromatophore connectivity in R. sphaeroides has been hampered by the limitations of experimental techniques used to assess their structure. Many reported results are based on ex-situ studies or on the interpretation of projection images, which can alter structures or discard important 3D information about vesicle connectivity (Cohen-Bazire and Kunisawa, 1963;Boatman, 1964;Holt and Marr, 1965;Tauschel and Drews, 1967;Prince et al, 1975;Holmqvist, 1979;Scheuring et al, 2014;Verméglio et al, 2016 ). Electron tomography (ET) provides 3D visualization of peripheral architecture in whole R. sphaeroides bacteria with nanoscale detail, however, resolving centermost structures is limited by cell thickness (Tucker et al, 2010;Scheuring et al, 2014).…”