Membrane sphingolipids as essential molecular signals for Bacteroides survival in the intestine

Abstract

As predominant intestinal symbiotic bacteria, Bacteroides are essential in maintaining the health of the normal mammalian host; in return, the host provides a niche with plentiful nutrients for the symbionts. However, the intestinal environment is replete with chemical, physical, and biological challenges that require mechanisms for prompt and adept sensing of and responses to stress if the bacteria are to survive. Herein we propose that to persist in the intestine Bacteroides take advantage of their unusual bacterial sphingolipids to mediate signaling pathways previously known to be available only to higher organisms. Sphingolipids convey diverse signal transduction and stress response pathways and have profound physiological impacts demonstrated in a variety of eukaryotic cell types. We propose a mechanism by which the formation of specific sphingolipid membrane microdomains initiates signaling cascades that facilitate survival strategies within the bacteria. Our preliminary data suggest that sphingolipid signaling plays an important role in Bacteroides physiology, enabling these bacteria to persist in the intestine and to perform other functions related to symbiosis.

Fig. 1.

Typical structures of (A) sphingolipids and (B) phospholipids. (C) Initial steps in eukaryotic sphingolipid biosynthetic pathway.

Fig. 2.

Sphingolipids affect the survival of Bacteroides in stationary phase but not the growth of the bacteria in log phase. Growth curves are shown for the wild-type B. fragilis culture in which sphingolipids were intact (WT) and the myriocin-treated B. fragilis culture in which sphingolipid synthesis was inhibited (WT + myriocin). Optical density curves are representative of three replications. Error bars for CFUs/mL curves show the SD of triplicate samples in one representative experiment.

Fig. 3.

Sphingolipids are important in the response of Bacteroides to stress. Rich-culture cells proficient with sphingolipids (WT) were compared with those deficient in sphingolipids (WT + myriocin) in terms of survival under heat shock and oxidative stress. Error bars show the SD of triplicate samples in one representative experiment.

Fig. 4.

Sphingolipids and cholesterol are both needed for an efficient stress response. Cells grown in minimal medium with different combinations of sphingolipids and cholesterol were compared in terms of survival after DNA crosslinking and oxidative stress. Error bars show the SD of duplicate samples in one representative experiment.

Fig. 5.

Bacteroides lipids form sphingolipid- and cholesterol-dependent heterogeneous domains in supported bilayers. (A) Schematic sketch shows formation of heterogeneous membrane domains. (B–E) Representative AFM height images (1 μm × 1 μm) of reconstituted bilayers using lipid extractions from B. fragilis Sph+Chol− (B), P. aeruginosa (P.a.; C), B. fragilis Sph−Chol− (D), and B. fragilis Sph+Chol+ (E). Color bars along AFM images indicate height ranges. The extended height range in sample Sph+Chol− was due to assembly of multiple lipid bilayers instead of single bilayers. Areas of multiple lipid bilayers were excluded from calculations shown in text.