RopB protein of Rhizobium leguminosarum bv. viciae adopts amyloid state during symbiotic interactions with pea (Pisum sativum L.)

Abstract

Amyloids represent protein aggregates with highly ordered fibrillar structure associated with the development of various disorders in humans and animals and involved in implementation of different vital functions in all three domains of life. In prokaryotes, amyloids perform a wide repertoire of functions mostly attributed to their interactions with other organisms including interspecies interactions within bacterial communities and host-pathogen interactions. Recently, we demonstrated that free-living cells of Rhizobium leguminosarum, a nitrogen-fixing symbiont of legumes, produce RopA and RopB which form amyloid fibrils at cell surface during the stationary growth phase thus connecting amyloid formation and host-symbiont interactions. Here we focused on a more detailed analysis of the RopB amyloid state in vitro and in vivo, during the symbiotic interaction between R. leguminosarum bv. viciae with its macrosymbiont, garden pea (Pisum sativum L.). We confirmed that RopB is the bona fide amyloid protein since its fibrils exhibit circular x-ray reflections indicating its cross-β structure specific for amyloids. We found that fibrils containing RopB and exhibiting amyloid properties are formed in vivo at the surface of bacteroids of R. leguminosarum extracted from pea nodules. Moreover, using pea sym31 mutant we demonstrated that formation of extracellular RopB amyloid state occurs at different stages of bacteroid development but is enhanced in juvenile symbiosomes. Proteomic screening of potentially amyloidogenic proteins in the nodules revealed the presence of detergent-resistant aggregates of different plant and bacterial proteins including pea amyloid vicilin. We demonstrated that preformed vicilin amyloids can cross-seed RopB amyloid formation suggesting for probable interaction between bacterial and plant amyloidogenic proteins in the nodules. Taken together, we demonstrate that R. leguminosarum bacteroids produce extracellular RopB amyloids in pea nodules in vivo and these nodules also contain aggregates of pea vicilin amyloid protein, which is able to cross-seed RopB fibrillogenesis in vitro. Thus, we hypothesize that plant nodules contain a complex amyloid network consisting of plant and bacterial amyloids and probably modulating host-symbiont interactions.

OmpC and OmpF Outer Membrane Proteins of Escherichia coli and Salmonella enterica Form Bona Fide Amyloids

“…For example, a number of outer membrane porins from bacterial species belonging to the order Rhizobiales were bioinformatically predicted to be potentially amyloidogenic [53]. Among Rhizobiales proteins, the RopA and RopB Omps of Rhizobium leguminosarum root nodule bacterium were experimentally proven to form bona fide amyloids in vitro and extracellular amyloid fibrils in vivo that are likely to modulate the interaction of Rhizobia with its multicellular host, Pisum sativum L. [33,54]. The Mannheimia haemolytica Omp2-like outer membrane protein was shown to participate in the formation of extracellular fibrils that bind CR and in the adhesion to the adenocarcinomic human alveolar basal epithelial cells [55].…”

Amyloid Fibrils of Pisum sativum L. Vicilin Inhibit Pathological Aggregation of Mammalian Proteins

“…At the same time, recent studies indicate that proteins in the amyloid state can perform important biological functions [ 6 , 7 ] in different organisms: archaea, bacteria, fungi, plants, insects and mammals [ 8 , 9 , 10 , 11 ]. These functions include but are not limited to the mechanical protection and modification of the cell surface properties, biotic or abiotic surface adhesion, pigment biosynthesis, storage and release of hormones and toxins and participation in host–pathogen and host–symbiont interactions [ 12 , 13 , 14 , 15 , 16 , 17 , 18 ].…”

On the anniversary of the President of the Vavilov Society of Geneticists and Breeders, Academician Igor Anatolyevich Tikhonovich

“…Игорь Анатольевич создал ведущую отечественную школу по генетике растительно-микробных взаимодействий: под его руководством защитили диссертации десятки кандидатов и докторов наук, а исследования получили широкую известность и признание на мировом уровне. Масштаб этой научной школы подтверждается не только сотнями работ по данной тематике, опубликованных в международных периодических изданиях (см., например, Kosolapova et al, 2022;Safronova et al, 2020;Dolgikh et al, 2019;Kitaeva et al, 2016;Serova et al, 2017;Sulima et al, 2017;Kulaeva et al, 2017;Samorodova et al, 2018;Kirienko et al, 2018;Serova et al, 2019;Dolgikh et al, 2020;Leppyanen et al, 2021) и монографиях (см., например, Tikhonovich, Provorov, 2009Provorov et al, 2016), но и целым рядом международных грантов и крупнейших конференций, проведенных под руководством И.А. Тихоновича, таких как Международный конгресс по биологической азотфиксации (Санкт-Петербург, 1995 год, около 700 участников из 50 стран мира) и Международный конгресс по молекулярным микробно-растительным взаимодействиям (Санкт-Петербург, 2003 год, около 800 участников из 70 стран).…”