MODULE 4 - Plant-Microbe Interactions

Question 1

what part of a plant hosts microbial communities?

Answer 1

every single part from leaves to roots and even the soil around it

Question 2

what is the rhizosphere?

Answer 2

zone of root influence - extends less than 5mm from root

Question 3

what is the rhizoplane?

Answer 3

root surface

Question 4

outline the microbial community in the rhizosphere?

Answer 4

populations around plant roots usually 20-100x more than surrounding soil (called the rhizosphere effect)

home to a vast array of species but beta and alpha proteobacteria often predominate, their growth is dependent on root exudates

around the root tip is where many microbes accumulate sloughed off cells make this area nutrient rich

bacteria grow as microcolonies over about 5% of the root surface meaning it is mostly sterile

microbes prefer to colonise the gaps between plants cells and around the root tip

Question 5

what are the bacterial communities in the rhizosphere strongly influenced by?

Answer 5

soil type and genetic constitution of host plant

Question 6

what are the four main phylum of bacteria which dominate the rhizosphere of many plants?

Answer 6

actinobacteria, bacteroidetes, firmicutes and proteobacteria

of these, alpha and beta-proteobacteria are the two most dominant classes of bacteria

Question 7

outline the multi-step process of how bacteria colonise plant roots?

Answer 7

bacteria swim up a gradient of exudates to reach root surface; especially root tip

primary attachment occurs (reversible) which is initially weak and mediated by hydrophobic and electrostatic forces

adhesion strengthened by something like flagella, fimbriae, adhesins or pili)

secondary attachment occurs (irreversible) where cellulose is produced further strengthening attachment

micro colonies form at site of adhesion and biofilms form through secretion of exopolysaccharides

Question 8

what are the three groups which the rhizosphere microbiome can be classified in?

Answer 8

beneficial microbes

commensals (neutral) (however they do modulate ability of beneficial and pathogenic bacteria to colonise root through normal microbe-microbe interactions)

pathogens (can infect plant root or secrete compounds which harm plant root)

Question 9

how might rhizosphere research help us reduce reliance on agrochemicals such as pesticides and fertilisers?

Answer 9

by tipping the balance of rhizosphere microbes in the favour of the beneficial ones

Question 10

what are some examples of rhizosphere microbes that have harmful effects on plant growth?

Answer 10

pathogenic fungi, oomycetes, nematodes and bacteria

root rot fungi such as fusarium, pythium

phytophthora agathidicida causes kauri dieback

Question 11

what are some of the harmful effects of pathogenic rhizosphere microbes other than the diseases they cause?

Answer 11

major problem for food production and ecosystem stability worldwide

e.g. pathogen resistance to applied agents, environmental impacts of pesticides, cost of pesticides, demand for pesticide-free food

Question 12

what are beneficial rhizosphere bacteria called?

Answer 12

plant growth-promoting rhizobacteria (PGPR)

Question 13

what are some of the direct beneficial effects of rhizosphere microbes like PGPR?

Answer 13

facilitate uptake of nutrients from enviro e.g. phosphate solubilisation, associative nitrogen fixation, siderophore production (iron acquisition)

synthesise compounds that affect plant growth e.g. plant growth regulators such as IAA which produces ACC deaminase that inactivates precursor of stress-hormone ethylene

Question 14

what do auxin-producing bacteria do?

Answer 14

promote plant growth

e.g. Pseudomonas corrugata with radish plants, Azospirillum with maize

Question 15

how do ACC deaminase-producing bacteria benefit plants?

Answer 15

halotolerant bacteria which protect plants from salt stress

Question 16

what are some indirect effects of beneficial rhizosphere microbes such as PGPR?

Answer 16

lessen or prevent effects of plant pathogens

e.g. pseudomonas fluorescens can control root rots caused by fungal pathogens such as Ggt which causes take-all disease

Question 17

how does Pseudomonas fluorescens prevent take-all disease from occurring in wheat and how was this discovered?

Answer 17

produces antibiotic (2,4-DAPG) which is toxic to Ggt, the take-all causing pathogen

this was sound out as many cases where wheat grown year after year in same soil and natural suppression of fungus occurred which is referred to as take-all decline (TAD)

TAD correlated with development of specific fluorescent Pseudomonas

important example of how bacteria control pathogens (biocontrol)

Question 18

why is soil type important in shaping the rhizosphere bacterial community?

Answer 18

it is recruited from the microorganisms present in the soil, thus soil type important in shaping rhizosphere microbiome

Question 19

why is plant genotype important in shaping the plant microbiome?

Answer 19

is a driving force for selection of specific elements from the bulk soil microbial community

when under stress/threat, evidence shows that plants can modify their rhizosphere microbiome to select for specific elements

Question 20

what are the three main mechanisms of biocontrol of plant diseases by bacteria in the rhizosphere?

Answer 20

antibiosis

induced systematic resistance

competition for nutrients and niches

Question 21

what is antibiosis?

Answer 21

bacterium colonises root system and delivers antibiotic molecules around the root thereby harming pathogens that approach the root

also commonly used to kill fungal pathogens which colonise same area as bacteria (gaps between plant cells)

Question 22

what is induced systematic resistance (ISR)?

Answer 22

local root colonisation is sufficient to induce ISR. Many bacterial products induce systemic signalling via plant receptors which can result in protection of the whole plant against diseases caused by different organisms

Question 23

how does biocontrol occur through competition for nutrients and niches?

Answer 23

biocontrol bacteria acting through this mechanism excel in fast chemotactic movement along the growing root in their efficient hunt for root exudate components, thereby outcompeting the pathogen in scavenging nutrients and in occupying niches on the root

basically just colonise quick as a form of niche exclusion

Question 24

what is the phylloplane?

Answer 24

leaf surface

Question 25

what is the phyllosphere?

Answer 25

the total surface area of the plant above ground

Question 26

why is the phyllosphere much harsher than the rhizosphere?

Answer 26

cause its exposed to rapid moisture fluctuations, rapid temperature fluctuations and UV radiation

so to colonise this enviro you have to be able to withstand these conditions

Question 27

describe the microbial population in the phylloplane?

Answer 27

microbial populations fluctuate rapidly

common inhabitants include epiphytic bacteria, gram negative bacteria e.g. Pseudomonas, Erwinia, Xanthomonas

Question 28

where are bacteria mostly concentrated in the phyllosphere?

Answer 28

large populations of bacteria around the trichome, majority of the leaf is sterile

Question 29

what are three Pseudomonas syringae methods of colonising the phylosphere?

Answer 29

production of syringomycin which acts as phytotoxin and surfactant to force release of nutrients by damaging plant cells and breaking down the waxy cuticle to allow its dispersal

auxin production and secretion by bacteria which provokes release of saccharides from plant cell wall

production of exopolysaccharide by bacteria to form biofilm which provides protection from environmental stresses

Question 30

what are some common bacterial traits involved in adaptation to the phyllosphere?

Answer 30

quorum sensing signalling molecules

antibiotics (often controlled by above so all released at same time)

pigments to protect from UV

auxin production

protective proteins

efflux pumps

EPS production

biosurfactants (get through waxy cuticle)

metabolic adaptations

Question 31

how does Pseudomonas syringae worsen frost damage?

Answer 31

it is ice nucleation active meaning that the bacteria can function as a nuclei for formation of ice crystals that can spread into plant tissues

this prevents supercooling of leaves so that frost damage occurs at -3 degs rather than the usual -8 degs without bacterial presence

this process is dependent on production of INA outer-membrane protein

Question 32

what has been done to try and prevent frost damage caused by P. syringae involving competitive exclusion?

Answer 32

production of mutant INA- strains of P. syringae released into environment to see if they competitively exclude INA+ wild type which are a problem for orchards

Question 33

how are INA+ strains of P. syringae used commercially?

Answer 33

in snow making machines to allow snow production at much warmer temperatures

Question 34

what bacteria can live in the atmosphere and induce rain/hail and why is this?

Answer 34

P. syringae

due to its adaptations to surviving in the phyllosphere, it can survive in the atmosphere

being swept up by wind and then inducing and falling with precipitation could be a method of spreading the bacteria to other plants

Question 35

what bacteria causes fireblight?

Answer 35

Erwinia amylovora

spread by pollinating insects

Question 36

how can we use Pseudomonas strains to prevent fireblight?

Answer 36

you can spray blossom with fast-colonising Pseudomonas strain which can’t cause disease but can competitively exclude and produce antibiotics to prevent Erwinia amylovora colonisation which is the pathogen causing fireblight

Question 37

what are legumes?

Answer 37

plants which have seeds in pods such as soybeans, clover, beans etc.

only plant that has developed ability to form symbiotic relationship with rhizobia

Question 38

what does infection of legume roots by rhizobia lead to?

Answer 38

root nodule formation and nitrogen fixation, a process where gaseous nitrogen is converted to combined nitrogen

Question 39

how is there host specificity in the rhizobium-legume symbiosis?

Answer 39

only a specific strain of rhizobia is able to nodulate a specific type of legume

Question 40

what are rhizobia?

Answer 40

gram-negative rod shaped soil bacteria that can form nodule on legume

rhizobia very diverse and fall into two groups - alpha-proteobacteria and beta-proteobacteria

nodule forming ability was passed around through HGT, this ability was developed AFTER they split into all these different genera

Question 41

what is the rhizobia-legume symbiosis characterised by?

Answer 41

a high degree of host specificity

Question 42

what are the stages of forming the rhizobia-legume symbiosis?

Answer 42

recognition of correct plant partner by rhizobia and then attachment of rhizobia to root hairs

this causes root hair to curl and induces cortical cell division opposite to where rhizobia binds (one side growing faster causes root hair to curl)

micro colony now trapped in curled root, plant cell wall begins to grow inwards forming an infection threat which rhizobia enters (meaning rhizobia always protected from plant cell cytoplasm)

cortical cell division induced in cells of inner cortex of plant root which forms organ which will be nodule

infection thread branches to contact dividing cells and rhizobia bud off into plant cell via endocytosis surrounded by membrane of plant origin

here the rhizobia terminally differentiate into misshapen bacterial cells called bacteroids which can fix nitrogen

nutrient transmission across plant cell membrane to bacteroid provides carbon source so bacteroid can make energy for N fixation in form of ammonia which it sends back across membrane

Question 43

what is a determinate nodule?

Answer 43

nodule grows not only by plant cells dividing but also by plant cells expanding after division

Question 44

what are the developmental stages which are present (can be seen in cross section) in a nodule?

Answer 44

meristematic zone = the part of the nodule where the cells are continuing to divide making it larger and larger

infection zone = where rhizobia are budding into dividing plant cells

nitrogen-fixing zone = where all the plant cells have bacteroids in them fixing nitrogen

Question 45

what is the purpose of the non-differentiated rhizobia living in the nitrogen-fixing zone?

Answer 45

these released back into soil when the nodule dies off to go infect other plants i.e. nodule provided an ideal sheltered space for rhizobia to grow and proliferate

Question 46

how does host specificity occur in the rhizobia-legume symbiosis i.e. how does it recognise its specific host?

Answer 46

host legume releases exudates into rhizosphere incl phenolic-like compounds called flavonoids

rhizobia can recognise particular flavonoid as opposed to other kinds of flavonoid so that it knows its in presence of specific host legume

rhizobia then induces a set of genes called nod genes which are required for nodule formation. These nod genes make a chemical of their own which has complex oligosaccharide structure which has a fatty acid which differs between rhizobia

so particular nod factors unique for particular rhizobia and legume recognises this via receptors on plant cells

so basically there is two-way signalling between host legume and rhizobia to induce nodule formation/N fixation

Question 47

what are nod genes encoded on?

Answer 47

encoded on a accessory genetic elements called symbiosis islands

e.g. large Sym plasmids in Rhizobium

e.g. chromosomal symbiosis islands in Mesorhizobium loti

this means that nod genes are part of the accessory genome

Question 48

what is Mesorhizobium loti?

Answer 48

rhizobia which modulates the legumes of the genus Lotus

symbiosis genes are on the chromosome

Question 49

what is the Mesorhizobium loti R7A symbiosis island?

Answer 49

mobile, chromosomally integrated element which integrates into a phe-tRNA gene in a process mediated by phage-type integrase

can excise from chromosome and transfer via conjugation (involving oriT, trb and tra genes) thus inserting itself into chromosome of recipient

has been shown in lab and in enviro to convert non-symbiotic mesorhizobia to symbionts (due to HGT of symbiosis island)

Question 50

what are the two groups of nod genes?

Answer 50

regulatory - the nodD gene product turns on other nod genes in response to flavonoid signal from plant

structural - responsible for synthesis of a lipochito-oligosaccharide signalling molecule called nod factor

so once recognition of flavonoid occurs by nodD gene transcription is activated for structural genes which make nod factor

Question 51

how are nod genes turned on?

Answer 51

host legume secretes specific signal molecule in root exudate. These are different for each rhizobia/legume pair and most are secondary plant metabolites called flavonoids (three-ring aromatic compound)

different nodD genes recognise different flavonoids and then turn on other nod genes which are recognised by receptors on legume and allow for nodulation to occur

(two-way signalling)

Question 52

what do nod genes produce?

Answer 52

a nod factor

this is an oligomer of N-acetyl-glucosamine (called a lipochitooligosaccharide) which is decorated by species specific bells and whistles (main one is a fatty acid (acyl group) which is unique for different rhizobia)

it is these bells and whistles that allow for host-specificity

Question 53

what are the plant responses to nod factors?

Answer 53

influx of calcium at root hair tip

calcium spiking in root hair nucleus

curling of root hair

initiation of cortical cell division

(nod factor share many characteristics with plant hormones)

Question 54

what is calcium spiking?

Answer 54

big calcium conc. changes through effluxes and influxes of root hair nucleus which occur about 10 minutes after adding nod factor

Question 55

what are NFR1 and NFR5?

Answer 55

two genes in Lotus japonicus which make proteins which form a receptor for nod factor

extracellular LysM domains bind nod factors and then transduce the symbiotic signals through the intracellular kinase domain of NFR1 to downstream signalling cascades

this leads to rhizobia infection and nodulation

the cascade includes activation of calcium-calmodulin kinase through calcium spiking

Question 56

what are the two pathways which can lead to formation of an infected nodule in Lotus japonicus?

Answer 56

both start with nod-factor perception by NFR1 and NFR5 and both end with infected nodule being formed

pathway 1 involves formation of infection thread

pathway 2 involves organogenesis

Question 57

outline the infection thread pathway to nodule formation?

Answer 57

nod-factor perception and then root hair curling which is followed by rearrangements of actin and then growth down infection thread to infect nodule

Question 58

outline the organogenesis pathway to nodule formation?

Answer 58

nod-factor perception and then signal transduction through a bunch of receptors which leads to calcium spiking and this eventually leads to organogenesis which leads to infected nodule

Question 59

what does cross-signalling between the two pathways to nodule formation in L. japonicus allow?

Answer 59

keeps the pathways in tune with each other (idk just know that there is cross-signalling)

Question 60

what two rhizobia genes are essential for symbiotic N fixation?

Answer 60

nif and fix genes

Question 61

outline the reaction involved in nitrogen fixation in root nodules?

Answer 61

reaction is catalysed by nitrogenase enzyme and has a very high energy requirement

rhizobia are obligate aerobes cause they needa make lots of ATP for N fixation and so have oxygen as terminal electron acceptor

problem is oxygen denatures the nitrogenase enzyme and inhibits expression of nif genes

Question 62

how do rhizobia allow for efficient aerobic respiration while protecting the nitrogenase enzyme?

Answer 62

their ECT has a very high oxygen affinity so can function well at low oxygen concentrations

plants have leg-haemoglobin which serves as an oxygen carrier within the nodule and facilitates diffusion of oxygen to bacteroids electron acceptors at high flux but low conc. i.e. ensures that its getting enough O2 for respiration but it is in a bound form so that it cannot harm nitrogenase inside the bacteria

N fixation genes only expressed in absence of intracellular oxygen - regulated by FixLJ

Question 63

what nutrient exchanges are occurring in the nodule between the plant and bacteroid?

Answer 63

host plant supplies bacteroid w C4-dicarboxylic acids as energy source

bacteria reduce N2 to NH3 which is translocated to host and assimilated in plant cell cytoplasm

Question 64

what happens to rhizobium mutants which are defective in C4-dicarboxylic acid transport?

Answer 64

they form ineffective nodules as no carbon source to turn into energy

Question 65

what is ammonium assimilation?

Answer 65

NH3 enters host cytosol via passive diffusion

NH3 assimilated through actions of glutamine synthetase and glutamate synthetase producing glutamine and glutamate

these products are used to synthesise amides or ureides (other nitrogen containing compounds) which are transported to the rest of the plant as a nitrogen source

Question 66

what nutrients do mycorrhizal fungi get from plants and what do they give in return?

Answer 66

carbs and lipids from plant

give back phosphates, nitrogen, water and also increase surface area tapped by roots

Question 67

why do many arbuscular mycorrhizae absolutely need the fatty acids synthesised by the host plant?

Answer 67

many of them are fatty acid auxotrophs

Question 68

what is the difference between ectomycorrhizae and endomycorrhizae?

Answer 68

ecto stay on outside of plant cells

endo form an intracellular infection

Question 69

describe ectomycorrhizae?

Answer 69

form sheath around root with little intercellular penetration and no intracellular penetration

found mostly in temperate forest trees

colonises short roots and forms sheath around root which protects from pathogens

Question 70

what is the main interface between fungi and plant where nutrients are exchanged?

Answer 70

the Hartig net

Question 71

describe endomycorrhizae?

Answer 71

three types - orchid, erichoid and arbuscular

orchid - restricted to orchids and characterised by non-pathogenic penetration of root cortex by septate fungal hyphae that sometimes form intracellular coils. Orchids require fungi to survive but produce orcinol to keep fungal growth checked i.e. fine balance between parasitism and mutualism

ericoid - restricted to Ericaceae and similar morphology to orchid mycorrhizae

arbuscular - most common type and found in crops, mycelia penetrates plant cells and forms branches called arbuscules. Differ from above two endomycorrhizae as hyphae are non-septate. Very little host specificity. Can be cultured w addition of fatty acid

Question 72

describe the arbuscule?

Answer 72

fungal branches inside plant cell from arbuscular endomycorrhizae

each branch surrounded by plant-derived periarbuscular membrane (PAM) which is continuous w plant plasma membrane and so fungus excluded from plant cytoplasm (why it isn’t pathogenic)

interface between fungal plasma membrane and PAM is called the periarbuscular space (PAS) which is comprised of both fungal and plant material

Question 73

which type of mycorrhizae forms a mantle on plant cells?

Answer 73

ectomycorrhizae

Question 74

how does communication occur between arbuscular mycorrhizae (AM) and root and what does this allow to form?

Answer 74

plant root secretes strigolactones, fungal spores sense this and germinate causing rapid growth/branching of mycelia

mycorrhizae secretes signalling molecules such as lipochitooligosaccharides with fatty chains (like nod factors)

these molecules trigger reactions in plant root such as calcium spiking which leads to fungal gene expression leading to formation of pre-penetration apparatus

root secretes cutin which signals to fungi to form hypha-odium and initiate arbuscular growth (penetrate and form arbuscules)

this allows symbiosis to form and is a less specific form of communication than that of plant-rhizobia

Question 75

what is the common symbiotic signalling pathway?

Answer 75

there is a common symbiotic signalling pathway shared between mycorrhizae and rhizobia

differences are AM fungus initiates it with myc factor well rhizobium initiates with nod factor

infection thread solely rhizobia and mycorrhizae less specific

however a lot of the genes for the pathways leading to nodulation and mycorrhization are shared

Question 76

what are the benefits of mycorrhizae?

Answer 76

essential for colonisation and growth of plants in nutrient-poor environments so they are key organisms in nutrient/carbon cycles in forest ecosystems

provide phosphate, nitrogen and other minerals and improve nutrient absorption for mycorrhizal plants through greater root SA provided by mycelia

enhanced resistance to drought stress cause greater water acquisition

protection against pathogens (especially ectomycorrhize as form mycelial sheath on plant root)

link plants together into communities which are more stress resilient than single plants

enable plant-plant communication

Question 77

describe how resource sharing occurs in mycorrhizal networks?

Answer 77

one mycorrhizal individual colonises different plants of different species

molecules such as carbon and nitrogen can be acquired by fungus and form nutrient patches which can then be passed between plants via the fungus

Question 78

how does plant-plant communication occur via common AM fungal networks?

Answer 78

plant infected with aphids increases its defences against them by emitting chemicals which are repellent to aphids but attract wasps which kill aphids

it is connected to other plants via underground hyphal network which also start to emit the same chemicals despite no aphids

when there is no hyphal network the other plants to not up regulate their defences

Question 79

what are some NZ examples of plant pathogens which pose major threats to some plants?

Answer 79

Pseudomonas syringae causing death of kiwifruit vines on orchards

Phytopthora agathidicida causing kauri dieback

Myrtle rust fucking up põhutakawa

Question 80

what kind of pathogens can attack plants?

Answer 80

bacteria, fungi, viruses and nematodes

Question 81

what occurs in most plant-pathogen interactions?

Answer 81

most pathogens only attack one or a limited number of plant species

often find resistant varieties to certain pathogens within plant species

even in susceptible plants, pathogen damage usually limited

plants are often able to recognise pathogen and mount co-ordinated defence against it. Once induced the plant defence response is usually effective

Question 82

what are the two defence systems that plant cells have to detect bacterial and fungal pathogens?

Answer 82

basal defence (PAMP triggered immunity - PTI)

gene-for-gene defence (effector triggered immunity - ETI)

Question 83

outline basal defence?

Answer 83

similar to innate immunity

induced upon infection by almost all microbes and based on recognition of general elicitors known as PAMPS (ubiquitous molecules found in microbes) hence PAMP-triggered immunity (PTI)

defence activated rapidly but generally without a hypersensitive response

Question 84

outline gene-for-gene defence?

Answer 84

induced upon infection by specialised pathogens

based upon recognition of highly specific Avr-gene products (effectors) by specialised, matching R-gene products hence effector-triggered immunity (ETI)

recognition results in rapid reaction called hypersensitive response which kills infected cells

Question 85

outline the plant-pathogen arms race?

Answer 85

plant pathogens have effector proteins which they secrete into plant cytoplasm which interfere with PAMP recognition by plant PRRs

this allows pathogen to cause disease by suppressing the plant basal defence

in response to this however the plant has evolved resistance proteins which recognise the impact of effector proteins and activates a seperate defence response (effector triggered immunity)

Question 86

why do plant pathogens develop effector proteins rather than just mutate PAMPs so that they are unrecognisable?

Answer 86

PAMPs are on highly conserved proteins

Question 87

what are some common PAMPs in bacteria and fungi?

Answer 87

bacteria: flagellin, elongation factor, LPS, cold-shock protein

fungi: chitin, beta-glucan, ergosterols

Question 88

describe flagellin as a PAMP?

Answer 88

flagellin is the main protein of the flagellum and is very conserved in the N terminal of flagellin where there is a peptide called flg22

FLS2 (flagellin sensing 2) is a receptor like kinase (RLK) which recognises flg22 and triggers ion fluxes, generation of ROS, protein phosphorylation, MAP kinase activation which then activates a shit load of genes which make antibiotics and shit and toughens up the cell wall making penetration harder

ROS also has a role in linking proteins in plant cell wall to make it tougher against penetration as well

so basically all these things protect the cell from infection

Question 89

how has the importance of the FLS2 receptor been shown in the lab?

Answer 89

FLS2- plants clearly more susceptible to infection from bacteria

Question 90

how many PAMP receptors do plants usually have?

Answer 90

lots!

Question 91

what is CERK1?

Answer 91

a PRR for chitin which is a common fungal PAMP

CERK1 PRR is homologous to the nod factor receptors NFR1 and NFR5 which mediate perception of lipo-chitin nod factors

so there is a lot of homology. between defence systems and systems that recognise rhizobia in plants

Question 92

what is it that makes a plant resistant to a particular pathogen?

Answer 92

if it is a pathogen it is virulent i.e. can cause disease

however the pathogen may have an avirulence gene (Avr) and if the plant has a resistance gene which matches the Avr gene, then no disease occurs

so for each resistance gene in the plant there is a corresponding Avr (avirulence) gene in the pathogen. The reason the pathogen cannot cause disease is not cause it lacks a virulence gene but because it has an avirulence gene which the plant will have a corresponding resistance gene for (R gene recognises the Avr gene to induce resistance i.e. no disease (incompatible reaction)

so a plant may have a resistance gene but if the pathogen doesn’t have corresponding Avr gene then disease occurs (compatible reaction)

a plant may have several R genes and to escape recognition and cause disease, a pathogen will have to lack all the Avr genes which correspond to the plant’s R genes

this is referred to as gene-for-gene complementarity

Question 93

what are the main plant defence responses elicited by pathogen recognition?

Answer 93

rapid oxidative burst w production of H2O2

rapid oxidative cross-linking of pre-existing cell wall proteins; toughens cell wall

expression of antimicrobial compounds called phytoalexins

expression of pathogenesis-related (PR) proteins e.g. things that lyse bacteria

rapid death of infected plant cells (hypersensitive response)

systemic acquired resistance (chemical signals diffuse through plant so other parts of the plant express defence mechanisms despite not having contacted pathogen yet)

Question 94

outline the entire defence response in plants in response to PAMP recognition?

Answer 94

PAMPs such as chito-oligosaccharide (COS) recognised by PRRs. PRR activation triggers MAP kinase pathway leading to up regulation of hundreds of defence related genes. Several WRKY transcription factors activated that are directly responsible for expression of defence genes. Activated basal defences include cell wall fortifications, production of ROS and in the case of ETI (effector-triggered immunity), the hypersensitive response. Other cellular responses include release of phytohormones which activate signalling pathways leading to increased expression of antimicrobial genes, cell wall reinforcement and signal transduction to the whole plant

Question 95

after avr genes were cloned, what were most of them found to encode?

Answer 95

hydrophilic proteins with no functional homologues in data bases

now we know that these proteins are secreted into the host cell cytoplasm by Type III secretion systems and are called effector proteins

Question 96

outline the proteins encoded by avr genes?

Answer 96

effector proteins which are secreted into host cell cytoplasm via type III secretion systems (T3SS)

most can be mutated to restore pathogenicity

can function as virulence factors on hosts lacking the corresponding R gene

in absence of R gene, Avr effectors function to suppress basal defence

Question 97

which is going to be more virulent against a plant without a resistance gene, a pathogen with no avr gene or a pathogen with an avr gene?

Answer 97

pathogen with avr gene as it will encode an effector protein which suppresses the plants basal defence

however if plant had compatible R gene there would be no disease caused as R gene and compatible avr gene leads to incompatible reaction (no disease caused)

note this is only with some avr genes

Question 98

after R genes were cloned, what were most of them found to encode?

Answer 98

most encode hydrophilic proteins with nucleotide-binding (NB) site and leucine-rich-repeat (LRR) region involved in protein-protein interactions. Some others encode protein kinases

the NB-LRR class can be subdivided based on N-terminal structural features:

• many have homology to intracellular signalling domains of the Drosophila Toll and mammalian IL-1 receptors (TIR-NB-LRR) (TIR structure)
• others contain coiled-coil domains (CC-NB-LRR) (coil-coil structure)

Question 99

what is the guard hypothesis for how Avr and R gene products interact?

Answer 99

R protein essentially guarding the target of the effector (Avr) protein and when the pathogen effector protein interacts with its target protein the resistance gene can sense this interaction, know its target is under attack and then is activated so it can signal on for other defence responses

other idea is that the R protein is seperate from the target of the effector but when the effector interacts with the target the R protein is is recruited and this sets off the cascade

bottom line is that the actual interaction is being sensed and it is the effect of that interaction that leads to activation of resistance

Question 100

what is the main point of the guard model/hypothesis and why is this beneficial?

Answer 100

the pathogen virulence proteins are recognised as a consequence of their virulence function, rather than by direct interaction with a plant R protein

this is beneficial because by recognising pathogen virulence proteins based on enzymatic activity rather than their shape, plants could likely detect a large number of pathogen effectors with a limited number of R proteins

Question 101

what are some genomic strategies we could use to breed durable resistance to plant pathogens?

Answer 101

use sequencing technologies to sequence genomes of pathogens causing disease

identify R genes that are activated by effectors

use bioinformatics to identify the most successful effectors in particular strains

Question 102

what bacteria causes crown gall formation?

Answer 102

Agrobacterium tumefaciens

Question 103

what is Agrobacterium tumefaciens?

Answer 103

alpha-proteobacterium closely related to Rhizobium

causes crown-gall disease which is important in stone fruits, grapes, rose, apples

only wounded cells can be transformed from stuff like freeze damage, grafting, mechanical injury

majority of genes for crown gall formation on large plasmid called Ti (tumour inducing) plasmid

Question 104

why can Agrobacterium lose the ability to induce crown galls and other bacteria gain it?

Answer 104

the Ti plasmid encodes most of the genes for crown gall

Question 105

outline the steps in crown gall formation?

Answer 105

first step is attachment - involves loose attachment via acidic scapular polysaccharide followed by production of cellulose fibrils that enmesh large numbers of bacteria at the wound surface

vir genes then expressed and T-DNA excised as single strand and is transferred to plant nucleus and integrated into plant genome so now these genes expressed in plant and not agrobacterium

tumorigenesis occurs in plant

Question 106

what are the two main regions on the Agrobacterium plasmid?

Answer 106

vir region

T-DNA region

Question 107

how does the Agrobacterium tumefaciens recognise the wounded plant cell?

Answer 107

wounded plant cell secretes phenolic compounds (acetosyringone)

these single molecules are recognised by VirA/VirG two-component regulatory system and this activates other vir genes

VirA senses them and VirG is a transcriptional activator which turns on the other virulence genes

Question 108

what parts of the T-DNA on the Ti plasmid are required for transfer?

Answer 108

only the borders required

all the DNA between these borders are transferred

Question 109

what does the T-DNA encode?

Answer 109

enzymes for auxin and cytokinin synthesis which cause hormone imbalance leading to tumour formation

also encodes opine synthesis genes and the bacteria uses the opines (v uncommon compounds) as carbon and nitrogen sources for growth

Question 110

how does the T-DNA get excised from the Ti plasmid of Agrobacterium tumefaciens?

Answer 110

process starts with nicking of T-DNA at its right border by a mob protein called VirD2

VirD2 covalently attaches to nicked strand

single strand of T-DNA unwinds and is released from the Ti plasmid by nicking at the left border

gap in Ti plasmid is repaired

T-strand transferred through a pilus encoded by verb operon; this VirD4 also required

VirD2 mob protein acts as pilot protein leading the strand to mating pore for transfer to plant cell

coating of T-strand with VirE2 occurs within plant cell

Question 111

describe the Type IV Secretion System (T4SS) in Agrobacterium?

Answer 111

bacterial cells contact unidentified host cell receptor using VirB2 pilin and VirB5 adhesin

a DNA-dependent ATPase called VirD4 delivers DNA bound by relaxase VirD2 to inner membrane secretion channel

ATPases VirB4 and VirB11 provide energy for secretion of DNA and effectors through a VirB6 inner membrane channel and a secretion complex in host cell

Question 112

once the T-DNA is inside the plant cell, how does it target the nucleus?

Answer 112

once inside plant cell T-DNA has to get to nucleus

VirD2 and VirE2 both contain two plant-active nuclear localisation (NLS) sequences and are both necessary for nuclear targeting

VirE2 interacts with plant protein VIP1 which is a transcription factor that is phosphorylated by MAP kinase which is activated by Agrobacterium infection as a defence response

Phosphorylation targets VIP1 to nucleus and T-DNA hitches ride through interaction with VirE2

VirD2 required for integration of T-DNA and integration occurs by illegitimate recombination

T-DNA encoded genes then expressed

cell division and opine synthesis occurs

Question 113

what are the responses to the opines synthesised by the tumours?

Answer 113

generally each Agrobacterium strain catabolises only the opines synthesised by the tumours it induces (catabolic genes encoded on Ti plasmid (tumour is their ecological niche)

some opines induce conjugal transfer of the Ti plasmid to other strains of Agrobacterium that may be present (opines are the Ti plasmids way of ensuring its propagated)