Plant lecture 14 - Symbiotic N fixation Flashcards
1
Q
Nitrogen fixation
Nitrogenase
A
- Dinitrogenase reductase = Fe protein, supplies reducing power, also MoFe protein that reduces N2
- Symbiotic N fixing = when N fixing bacteria associate with a plant
- E.g. rhizobia + legumes forming root nodules
2
Q
Nodules development in legumes 1
A
- Flavonoids released by roots of host plant → change in rhizobia gene expression (nod genes)
- Nod genes allow synthesis of nod factors (lipochitin oligosaccharides) that initiate change in host gene expression (ENOD genes encoding for early nodulins)
- In addition to change gene expression, Nod factors also trigger influxes, membrane depolarisation
3
Q
Nod factors
A
- Typically lipochitin oligosaccharide w/ NAG backbone
- Chemical modifications on terminal residue are important for interaction with host plant + encodes specificity of interaction
- Plant releases compounds into rhizosphere, some detected by Rhizobia which produces a particular Nod factor that if recognised by plant starts formation of nodule
4
Q
Nod factor signalling
A
- DS signalling component identified by genetic analysis of Lotus japonius
- Signal received by receptor kinase. Causes Ca2+ influx which impacts TF NSP1/2
- NSP1/2 can also be activated by DMI1/2 (ligand gated cation channel/ receptor kinase) which triggers ca2+ spiking around nucleus + activates DMI3 (calcium/calmodulin-dependent protein kinase)
- If deregulate DM13 by removing autoinhibition domain, causes nodulation w/o rhizobia (rhizobia critical in cascade)
Ubiquination
- LYK3 receptor kinase = nod factor protein that assists binding of nod factor to kinase
- W/o nod factor, PUBI = E3 ubiquitin ligase that is active. Ubiquinates a protein needed for infection + infection x
- When nod factor binds LYK3, LYK3 phosphorylates PUB1 + inactivates
Bacterial exopolysaccharide (EPS)
- Important for host invasion by symbiotic bacteria
- In Lotus japonicus, nod factor signal transduction induces transcription of the host gene Epr3
- This encodes a receptor protein similar to NRF1 family which can recognise EPS
5
Q
Nodule development in legumes 2
During infection
A
- Bacteria attach to root harms promote growth + curling
- Modification of cell wall followed by invagination of the plasma membrane → infection thread
- Invagination propagates into root cortex + nodule primordial is formed in cortex
- LOF + GOF mutations in cytokinin receptor show activation of LHK1 is sufficient to trigger nodule formation
- Signalling coordinates activity at surface vs centre of root cortex (cortex is ready for invading bacteria)
6
Q
Role of nodule inception protein in nodulation (NIN)
A
- NIN - bifunctional TF suppressing ENOD11 expression in epidermis + promoting transcription of cytokinin receptor CRE1 in root cortex
- Nod factors arrive at root hair + trigger expression of NIN
- NIN activates the cortical program leading to organogenesis
- NIN activates NPL that breaks down cell wall + allows invag. of plasma membrane to create infection thread
- Some signal triggers cytokinin production that → expression of NIN in cortex
- Sets up +ve feedback loop: production of NIN ↑ expression of CRE1, which ↑ zone within root sensitive to cytokinin. Eventually NIN inhibited
- NIN controls a diversity of functions inc. cell wall modification via NPL, GA biosynthesis by genes like CPS1, nutrient uptake + DNA synthesis
7
Q
Nodule development in legumes - 3
Nodule formation
A
- The infection thread extends into the nodule primordium, allowing rhizobia to enter the plant.
- Bacteria are released into the cytoplasm, forming symbiosomes that occupy up to 80% of the cell volume and contain up to 20 bacteria surrounded by the plant-derived peribacteroid membrane.
- The bacteria differentiate into endosymbiotic bacteroids + the nodule primordium develops into a mature nodule.
8
Q
Symbiotic bacterial gene expression
A
- Nod, nol and noe genes are required for the synthesis of the nod factors that initiate nodule development:
1. The nodD gene product activates the expression of the other nod genes after forming a complex with secondary metabolites in the root exudate
2. NodA-C gene products make lipochitin oligosaccharide backbone
3. Other nod gene products determine specificity by controlling the chemical modification of terminal residues
4. Nif genes = both symbiotic bacteria + N2 fixing. Gene products Include Fe protein + MoFe, regulatory proteins for bif gene expression, NifA/L to name a few
5. Fix genes - only occur in symbiotic N2 fixing bacteria. Gene products include components of O2 sensing system (FixJ,L,K) + structural proteins of high affinity bacterial terminal oxidase. Protect nitrogenase from denaturation by O2
9
Q
Maintaining an anerobic environment
A
3 factors help ↓ O2 conc.
- Restricted diffusion, based on a variable permeability barrier controlling O2 exchange at the nodule periphery and the reduction of intercellular air spaces.
- Binding to leghaemoglobin reduces the free O2 concentration in the host cytosol to 10-25 nM. RNAi-induced abolition of leghaemoglobin synthesis prevents symbiotic N fixation.
- Bacterial respiration, aided by the ↑ affinity terminal oxidase (Km ~ 7 nM), acts as a major oxygen sink.
- Low O2 limits nodule respiration + nitrogenase. Effect greater in bacteroid than host cytosol as host mit. localise at cell surface near intracellular air spaces
10
Q
Nitrogen sensing
A
- Free living diazotrophs use 2 component N control system to regulate Nif gene expression
- NtrB/C. Less common in symbiotic diazotrophs, reflects role of symbiotic diazotroph as source of fixed N for host + importance of O2 sensing
- NtrC-P activates NifA/L
11
Q
Symbiotic plant gene expression
A
- Early nodulins
- Tissue specific expression = essential for successful infection + nodule development
- Epidermal Nod factor-induced ENOD expression occurs in the vicinity of actively growing root hairs
- Pectate lyase induced in the nodule primordium degrades the plant cell wall around the infection thread, facilitating the formation of symbiosomes - Late nodulins
- Establishes metabolic conditions for N2 fixation in the host cytosol:
- leghaemoglobin for controlling oxygen availability
- glutamine synthetase for ammonium assimilation
- sucrose synthase for sucrose breakdown