Characterization of three novel mechanosensitive channel activities in Escherichia coli

Mechanosensitive channels sense elevated membrane tension that arises from rapid water influx occurring when cells move from high to low osmolarity environments (hypoosmotic shock). These non-specific channels in the cytoplasmic membrane release osmotically-active solutes and ions. The two major mechanosensitive channels in Escherichia coli are MscL and MscS. Deletion of both proteins severely compromises survival of hypoosmotic shock. However, like many bacteria, E. coli cells possess other MscS-type genes (kefA, ybdG, ybiO, yjeP and ynaI). Two homologs, MscK (kefA) and YbdG, have been characterized as mechanosensitive channels that play minor roles in maintaining cell integrity. Additional channel openings are occasionally observed in patches derived from mutants lacking MscS, MscK and MscL. Due to their rare occurrence, little is known about these extra pressure-induced currents or their genetic origins. Here we complete the identification of the remaining E. coli mechanosensitive channels YnaI, YbiO and YjeP. The latter is the major component of the previously described MscM activity (~300 pS), while YnaI (~100 pS) and YbiO (~1000 pS) were previously unknown. Expression of native YbiO is NaCl-specific and RpoS-dependent. A Δ7 strain was created with all seven E. coli mechanosensitive channel genes deleted. High level expression of YnaI, YbiO or YjeP proteins from a multicopy plasmid in the Δ7 strain (MJFGH) leads to substantial protection against hypoosmotic shock. Purified homologs exhibit high molecular masses that are consistent with heptameric assemblies. This work reveals novel mechanosensitive channels and discusses the regulation of their expression in the context of possible additional functions.