Functional compartmentalization and metabolic separation in a prokaryotic cell

Abstract

The prokaryotic cell is traditionally seen as a "bag of enzymes," yet its organization is much more complex than in this simplified view. By now, various microcompartments encapsulating metabolic enzymes or pathways are known for Bacteria These microcompartments are usually small, encapsulating and concentrating only a few enzymes, thus protecting the cell from toxic intermediates or preventing unwanted side reactions. The hyperthermophilic, strictly anaerobic Crenarchaeon Ignicoccus hospitalis is an extraordinary organism possessing two membranes, an inner and an energized outer membrane. The outer membrane (termed here outer cytoplasmic membrane) harbors enzymes involved in proton gradient generation and ATP synthesis. These two membranes are separated by an intermembrane compartment, whose function is unknown. Major information processes like DNA replication, RNA synthesis, and protein biosynthesis are located inside the "cytoplasm" or central cytoplasmic compartment. Here, we show by immunogold labeling of ultrathin sections that enzymes involved in autotrophic CO₂ assimilation are located in the intermembrane compartment that we name (now) a peripheric cytoplasmic compartment. This separation may protect DNA and RNA from reactive aldehydes arising in the I. hospitalis carbon metabolism. This compartmentalization of metabolic pathways and information processes is unprecedented in the prokaryotic world, representing a unique example of spatiofunctional compartmentalization in the second domain of life.

Keywords: Archaea; CO2 fixation; compartmentalization; immunogold labeling.

Fig. 1.

The DC/HB cycle (adapted from ref. 3) (A), ultrastructure (B), and 3D model (C) of I. hospitalis. Enzymes: 1) pyruvate synthase, 2) pyruvate:water dikinase, 3) PEP carboxylase, 4) malate dehydrogenase, 5) fumarate hydratase, 6) fumarate reductase (natural electron donor is not known), 7) succinyl-CoA synthetase, 8) succinyl-CoA reductase (natural electron acceptor is not known), 9) succinic semialdehyde reductase, 10) 4-hydroxybutyrate-CoA ligase, 11) 4-hydroxybutyryl-CoA dehydratase, 12) crotonyl-CoA hydratase, 13) (S)-3-hydroxybutyryl-CoA dehydrogenase, and 14) acetoacetyl-CoA β-ketothiolase. Fd, ferredoxin; MV, methyl viologen. The enzymes studied in this work are highlighted in red. Ultrathin section (B) and 3D model of a semithin section (C) of a cryo-fixed, freeze-substituted, Epon-embedded cell. CCC (central cytoplasmic compartment), PCC (peripheric cytoplasmic compartment), ICM (inner cytoplasmic membrane), TN (tubular network), OCM (outer cytoplasmic membrane), and Neq (Nanoarchaeum equitans). 3D model highlights the TN originating from the CCC. (Scale bars, 500 nm.)

Fig. 2.

Localization of enzymes involved in the DC/HB cycle (resin sections). (A–D) Image sections of SI Appendix, Fig. S3. Localization of PEP carboxylase (A), malate dehydrogenase (B), succinic semialdehyde reductase (C), and crotonyl-CoA hydratase/(S)-3-hydroxybutyryl-CoA dehydrogenase (D). The black arrow in D indicates gold particles in protrusions derived from the central CC. (Scale bars, 100 nm.)

Fig. 3.

Localization of DNA (cryosection). I. hospitalis cell is labeled with an anti-DNA antibody according to Tokuyasu (16). (Scale bar, 500 nm.)