As a soil organism, P. putida recurrently encounters filament-inducing conditions during its natural life cycle. Our data indicate that filament formation of P. putida could confer environmentally advantageous traits. Indeed, P. putida KT2440 grown at low shaking speed produced filaments and was more resistant to heat shock and saline stress. Similar observations were made for Caulobacter crescentus filaments, which showed a higher resistance to oxidative, osmotic, thermal and acid stress
The comparative proteome profile indicated that the metabolic activity of P. putida KT2440 grown at 50 rpm was significantly different from P. putida KT2440 grown at 150 rpm. The most pronounced induction occurred for the heat shock protein IbpA. This small heat shock protein belongs to the widely conserved family of α-crystallin-type heat shock proteins. The latter appears to play a very versatile role in the protection against different stress conditions via protein and membrane protection
. In addition, many small heat shock proteins form oligomers, which may vary by the degree of phosphorylation or ion concentration
 (induction of PP_2645, PP_2656 and PP_5329).
Although no observable differences in dissolved oxygen levels could be reported at the time of proteomic analysis (i.e., 15 hours, below detection limit for both conditions) (Figure
2), this does not completely rule out the role of dissolved oxygen in the observed results as the maximum oxygen transfer rate at 150 rpm is approximately 2.5 times higher than at 50 rpm
. Ohr, a protein of the OsmC family (osmotically inducible protein) was 6.25-fold down-regulated in filamented P. putida, and is involved in the resistance to oxidative stressors, such as organic peroxide, but not in osmotic stress resistance
. In addition to a decreased Ohr abundance, other proteins involved in oxidative stress resistance were present at lower levels in 50 rpm samples, including a catalase/peroxidase (PP_3668, 0.28-fold), an antioxidant AhpC (PP_1084, 0.42-fold), a glutaredoxin-related protein (PP_1081, 0.44 fold) and a putative DNA binding stress protein (PP_1210, 0.32-fold). The latter has recently been described as an oxidative stress-inducible Dps miniferritin
[22, 23], and was found up-regulated in an OxyR mutant of P. aeruginosa. The differential abundance of proteins involved in oxidative stress resistance could potentially be explained by lower oxygen levels in 50 rpm cultures (and/or decreased catabolism). The increase of OprE (PP_0234, 2.41-fold) and CyoA (PP_0812, 1.82-fold) further suggests limitations in oxygen availability in 50 rpm cultures
[24, 25]. Finally, oxygen limitation is related to bacterial filamentation and/or RecA induction
[6, 26–28]. However, Jenssen and colleagues determined that, in addition to oxygen deprivation, additional factors such as exhaustion of medium components and changes in growth rate, were important for P. putida filamentation
While RecA was more abundant in P. putida KT2440 grown at 50 rpm, the P. putida KT2440 recA mutant filamented at similar levels as the wild type. A similar observation was reported previously, showing that an E. coli recA mutant displayed similar levels of filamentation as the wild type strain in response to growth at high pressure, despite strong evidence of RecA-mediated SOS response activation
[29–31]. Gottesman et al. (1981) suggested the existence of a transient filamentation phenotype in response to UV, independent of SulA
, which could explain the RecA-independent filamentation phenotype of 50 rpm-grown P. putida KT2440 in the present study.
While the bacterial SOS response and associated filamentation is typically triggered by treatments directly affecting DNA integrity (e.g. exposure to mitomycin C or UV), a number of environmental conditions were reported to cause DNA damage in an indirect manner (e.g. starvation, aging, β-lactam antibiotics and high pressure stress)
[30, 33–36]. As such, high pressure-induced filamentation of E. coli was shown to stem from the activation of a cryptic Type IV restriction endonuclease (i.e. Mrr) endogenously present in the cell
, while β-lactam antibiotics triggered DpiA to interfere with DNA replication
[30, 36]. Even though it remains unclear which metabolic changes could indirectly lead to DNA damage and SOS response activation, the major changes in metabolism provide evidence for new triggers of the SOS response.