Influence of mutation in pea (Pisum sativum L.) cdt (cadmium tolerance) gene on histological and ultrastructural nodule organization

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Abstract

Background. A comparative analysis out of the structural organization of the symbiotic nodules of the pea initial line SGE and the mutant line SGECdt, characterized by increased tolerance to cadmium and increased its accumulation, was carried out.

Materials and methods.Nodules of initial line SGE and mutant SGECdt were analyzed using light and transmission electron microscopy.

Results. The non-treated nodules of SGE and SGECdt were characterized by a similar histological and ultrastructural organization. In the nodules of SGE exposed to 100 µM CdCl2 in infected cells, the following abnormalities were observed: expansion of the peribacteroid space, destruction of the symbiosome membrane, fusion of symbiosomes and, as a result, the formation of symbiosomes containing several bacteroids. In the nodules of SGECdt, infected cells did not undergo pronounced changes. In the nodules of SGE exposed to 1 mM CdCl2, at the base of the nodule, senescent infected cells with completely destroyed cytoplasm and degrading bacteroids appeared. Also there were present cells in which the contents of symbiosomes were lysing, and only the “ghosts” of the bacteroids remained in them. In SGECdt, in some infected cells, abnormalities were manifested in an increase in the peribacteroid space, partial destruction of symbiosome membranes, fusion of symbiosomes, and release of bacteroids into the vacuole.

Conclusions. The tolerance of pea nodules to cadmium can be significantly increased due to a single recessive cdt mutation.

About the authors

Anna V. Tsyganova

All-Russia Research Institute for Agricultural Microbiology

Email: isaakij@mail.ru
ORCID iD: 0000-0003-3505-4298

Leading Scientist, Laboratory of Molecular and Cellular Biology

Russian Federation, 3, Podbelsky highway, Pushkin, Saint-Petersburg, 196608

Elena V. Seliverstova

All-Russia Research Institute for Agricultural Microbiology; Sechenov Institute of Evolutionary Physiology and Biochemistry of the RAS

Email: elena306@yandex.ru
ORCID iD: 0000-0003-2096-693X

Senior Scientist, Laboratory of Molecular and Cellular Biology; Senior Scientist, Laboratory of Kidney Physiology and Water-Salt Metabolism

Russian Federation, 3, Podbelsky highway, Pushkin, Saint-Petersburg, 196608; 44, Thorez prospekt, St.-Petersburg, 194223

Viktor E. Tsyganov

All-Russia Research Institute for Agricultural Microbiology

Author for correspondence.
Email: tsyganov@arriam.spb.ru
ORCID iD: 0000-0003-3105-8689
SPIN-code: 6532-1332
Scopus Author ID: 7006136325
ResearcherId: Q-5634-2016
http://arriam.ru/departments/laboratoriya-molekulyarnoj-i-kletochnoj-biologii/

Head of the Laboratory, Laboratory of Molecular and Cellular Biology

Russian Federation, 3, Podbelsky highway, Pushkin, Saint-Petersburg, 196608

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2. Fig. 1. Histological organization of pea nodules of the initial line SGE and mutant line SGECdt, untreated and treated with 100 μM CdCl2, and ultrastructural organization of untreated SGECdt nodules: a – histological organization of untreated SGE nodules; b – histological organization of untreated SGECdt nodules; c, e – histological organization of SGE nodules treated with 100 μM CdCl2; d, f – histological organization of SGECdt nodules treated with 100 μM CdCl2; g – infected cell of SGECdt from the infection zone with infection thread and juvenile bacteroids; h – infected cell of SGECdt from the nitrogen fixation zone with pleomorphic bacteroids. I – meristem, II – infection zone, II-III – interzone between infection and nitrogen fixation zones, III – nitrogen fixation zone, ic – infected cell, uic – uninfected cell, dic – degrading infected cell, IT – infection thread, CW – cell wall, B – bacterium, Ba – bacteroid, JBa – juvenile bacteroid; arrows indicate symbiosome membrane. Scale bar: a-d – 100 µm, e, f – 20 µm, g, h – 1 µm

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3. Fig. 2. Ultrastructural organization of pea nodules of the initial line SGE, treated with 100 μM CdCl2: a, b – infected cells from the nitrogen fixation zone with pleiomorphic bacteroids; c, d – infected cells from the nitrogen fixation zone with the primary signs of senescence (degradation of the cytoplasm and developmental abnormalities of the infection thread); e, f – degrading infected cells from the senescence zone. IC – infected cell, DIC – degrading infected cell, N – nucleus, A – amyloplast, IT – infection thread, Ba – bacteroid, MS – “multiple” symbiosome, formed as a result of symbiosome fusion and containing several bacteroids; arrows indicate a symbiosome membrane, arrowheads indicate destruction of symbiosome membranes, asterisks indicate outgrowths of infection thread containing a matrix without bacteria. Scale bar: a, c, e – 5 µm; b, d, f – 1 µm

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4. Fig. 3. Ultrastructural organization of pea nodules of the mutant line SGECdt, treated with 100 μM CdCl2: a – infected cell from the nitrogen fixation zone with infection thread and infection droplet; b – infected cell from the infection zone with infection thread and juvenile bacteroids; c – infected cells from the nitrogen fixation zone, filled with symbiosomes with expanded peribacteroid spaces; d – pleomorphic bacteroids from the nitrogen fixation zone. IC – infected cell, N – the nucleus, CW – cell wall, IT – infection thread, ID – infection droplet, Ba – bacteroid, MS – “multiple” symbiosome, formed as a result of symbiosome fusion and containing several bacteroids; arrows indicate symbiosome membrane, arrowheads indicate the destruction of symbiosome membranes. Scale bar: a – 5 μm; b, d – 1 µm; c – 2 µm

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5. Fig. 4. Ultrastructural organization of pea nodules of the initial line SGE, treated with 1 mM CdCl2: a – infected cells from the nitrogen fixation zone with premature signs of degradation; b – pleomorphic bacteroids from the nitrogen fixation zone; c – degrading infected cell from the senescence zone with infection thread and infection droplet; d – bacteroids of irregular shape and with rugose surface in degrading infected cell from the senescence zone; e – degrading infected cell from the senescence zone, filled with the “ghosts” of bacteroids; f – degrading bacteroids with a partly cleared matrix and destroyed symbiosome membrane. IC – infected cell, DIC – degrading infected cell, N – nucleus, V – vacuole, CW – cell wall, IT – infection thread, ID – infection droplet, Ba – bacteroid, DBa – degrading bacteroid, MS – “multiple” symbiosome, formed as a result of symbiosome fusion and containing several bacteroids; arrows indicate symbiosome membrane, arrowheads indicate the destruction of symbiosome membranes, and triangles indicate the “ghosts” of bacteroids. Scale bar: a, c, e – 5 µm; b, d, f – 1 µm

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6. Fig. 5. Ultrastructural organization of pea nodules of the mutant line SGECdt, treated with 1 mM CdCl2: а – infected cell from the infection zone with juvenile bacteroids; b – pleomorphic bacteroids in the infected cell from the nitrogen fixation zone; c – infected cell from the nitrogen fixation zone with primary signs of symbiosome degradation: expansion of the peribacteroid space, destruction of the symbiosome membrane and fusion of symbiosomes; d – degrading infected cell from the senescence zone. IC – infected cell, V – vacuole, CW – cell wall, A – amyloplast, Ba – bacteroid, JBa – juvenile bacteroid, MS – “multiple” symbiosome, formed as a result of symbiosome fusion and containing several bacteroids, arrows indicate symbiosome membranes, arrowheads indicate the destruction of symbiosome membranes. Scale bar: a, b – 1 µm; c – 2 µm; d – 5 µm

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Copyright (c) 2019 Tsyganova A.V., Seliverstova E.V., Tsyganov V.E.

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