TY - JOUR
T1 - Pal depletion results in hypervesiculation and affects cell morphology and outer-membrane lipid asymmetry in bordetellae
AU - de Jonge, Eline F
AU - van Boxtel, Ria
AU - Balhuizen, Melanie D
AU - Haagsman, Henk P
AU - Tommassen, Jan
N1 - Funding Information:
We would like to thank Nathalie Devos (GlaxoSmithKline Biologicals SA) for providing BrkA antiserum. We also thank Gosse Metz and Puck Roos for their contributions in initial experiments. This work received funding from the domain Applied and Engineering Sciences (TTW) of The Netherlands Organization for Scientific Research (NWO) (TTW Perspectief grant numbers 14921 and 14924), which received financial contributions for this grant from GlaxoSmithKline Biologicals SA and PULIKE Biological Engineering Inc.
Funding Information:
We would like to thank Nathalie Devos (GlaxoSmithKline Biologicals SA) for providing BrkA antiserum. We also thank Gosse Metz and Puck Roos for their contributions in initial experiments. This work received funding from the domain Applied and Engineering Sciences (TTW) of The Netherlands Organization for Scientific Research (NWO) (TTW Perspectief grant numbers 14921 and 14924 ), which received financial contributions for this grant from GlaxoSmithKline Biologicals SA and PULIKE Biological Engineering Inc .
Publisher Copyright:
© 2022 The Author(s)
PY - 2022/5/1
Y1 - 2022/5/1
N2 - Current vaccines against Bordetella pertussis do not prevent colonization and transmission of the bacteria, and vaccine-induced immunity wanes rapidly. Besides, efficacy of vaccines for Bordetella bronchiseptica remains unclear. Novel vaccines could be based on outer-membrane vesicles (OMVs), but vesiculation of bordetellae needs to be increased for cost-effective vaccine production. Here, we focused on increasing OMV production by reducing the anchoring of the outer membrane to the peptidoglycan layer. Inactivation of rmpM, tolR, and pal failed, presumably because their products are essential in bordetellae. Conditional pal mutants were constructed, which were hypervesiculating under Pal-depletion conditions. SDS-PAGE and Western blot analyses showed that the protein composition of OMVs produced under Pal-depletion conditions resembled that of the outer membrane but differed from that of OMVs released by the wild type. Pal depletion affected the cell morphology and appeared to increase the amounts of cell-surface-exposed phospholipids, possibly reflecting a role for the Tol-Pal system in retrograde phospholipid transport. We also identified additional lipoproteins in bordetellae with a putative peptidoglycan-anchoring domain. However, their inactivation did not influence OMV production. We conclude that the conditional pal mutants could be valuable for the development of OMV-based vaccines.
AB - Current vaccines against Bordetella pertussis do not prevent colonization and transmission of the bacteria, and vaccine-induced immunity wanes rapidly. Besides, efficacy of vaccines for Bordetella bronchiseptica remains unclear. Novel vaccines could be based on outer-membrane vesicles (OMVs), but vesiculation of bordetellae needs to be increased for cost-effective vaccine production. Here, we focused on increasing OMV production by reducing the anchoring of the outer membrane to the peptidoglycan layer. Inactivation of rmpM, tolR, and pal failed, presumably because their products are essential in bordetellae. Conditional pal mutants were constructed, which were hypervesiculating under Pal-depletion conditions. SDS-PAGE and Western blot analyses showed that the protein composition of OMVs produced under Pal-depletion conditions resembled that of the outer membrane but differed from that of OMVs released by the wild type. Pal depletion affected the cell morphology and appeared to increase the amounts of cell-surface-exposed phospholipids, possibly reflecting a role for the Tol-Pal system in retrograde phospholipid transport. We also identified additional lipoproteins in bordetellae with a putative peptidoglycan-anchoring domain. However, their inactivation did not influence OMV production. We conclude that the conditional pal mutants could be valuable for the development of OMV-based vaccines.
KW - Bordetella
KW - Outer-membrane vesicles
KW - Peptidoglycan-associated lipoprotein
KW - RmpM
KW - Tol-Pal system
UR - http://www.scopus.com/inward/record.url?scp=85129635976&partnerID=8YFLogxK
U2 - 10.1016/j.resmic.2022.103937
DO - 10.1016/j.resmic.2022.103937
M3 - Article
C2 - 35248703
SN - 0923-2508
VL - 173
SP - 1
EP - 13
JO - Research in Microbiology
JF - Research in Microbiology
IS - 4-5
M1 - 103937
ER -