Lecture 5, HOST DEFENSE RESPONSES-I

INTRODUCTION

Plants like human and other animals are often suffering various diseases. After a long evaluation history, surviving varieties must develop certain mechanisms to protect themselves against pathogen attack. Therefore, the mechanisms for plant to defense themselves against pathogen attack is one of the most important aspect for plant pathologists to deal with.

Plant disease resistance has been most intensively studied in Plant Pathology for over hundred years. Plant disease resistance is also the most fast developing area in plant biology in last ten to fifteen years. Thus, it is impossible to cover all aspects in two lectures, and it is also difficult to go deeply in any particular aspects. Therefore, the lecture will cover historical development and basic terminology in these areas, emphasize on some important aspects dealing with resistance mechanisms, update the knowledge on some fast developing areas and explore questions for future studies.

PASSIVE DEFENSES (PREEXISTING DEFENSES)

I). Preexisting Structural and Chemical Defense

Preexisting structural defenses

[The structures, such as wax, cutin, suberin, lignin, calcium and silicon, may present a structural barrier to physical penetration by the invading pathogen; may contain components which directly inhibit pathogen growth; may contain inhibitors of enzymes, or toxins, of the pathogen; may reduce molecular interchange between host and pathogen; may contain a signal compound(s)]

Preexisting chemical defenses

(Protocatechuic acid, for example, is exuded from red skin onion, but not from white skin onion)

(tannins, tomatine, avenacin, etc)

Saponin (avenacin) (Anne Osbourn, John Innes Center) - A secondary metabolite founded in many major crops.

Saponin deficient (sad) mutants of the diploid oat species Avena strigosa were generated (which are defective in their ability to make triterpenoid oat root saponins known as avenacins.

sad mutants are more susceptible to fungal diseases, providing good evidence that saponins may protect oats against pathogen attack.

Saponins complex with sterols in fungal membranes to cause loss of membrane integrity, although the way in which this occcurs is not fully understood. While some fungi may be saponin-resistant becaue they lack membrane sterols, others are able to detoxify saponins using saponin glycosyl hydrolases. At least one of these enzymes (avenacinase, produced by Gaeumannomyces graminis var. avenae) is essential for successful infection of saponin-containing plants. We are using the following complementary approaches to investigate the nature and phytopathological significance of saponin resistance in fungi:

1) Isolation and mutational analysis of saponin glycosyl hydrolase genes from fungal pathogens.

2) Use of yeast as a model system for the study of saponin resistance in fungi.

These topics represent several hot research areas for current studies. Recent references will be cited to PNAS 97:1897, 2000; Trend in Plant Science 5:343, 2000.

ACTIVE DEFENSE (INDUCED DEFENSES)

Preexisting mechanisms do not always protect plants against the pathogen attack. In a long evaluation process, plants develop active defense mechanisms which provide more efficient ways to protect plants. Therefore, active defenses are usually quiescent, activated by the pathogen attack or other stimuli and costly.

II). Induced Structural and Biochemical defenses

Plant-pathogen recognition and signal transduction

(hydrogen peroxide, nitride oxide, calcium, protein kinase, protein phosphotase, systemin, salicylic acid, jasmonic acid, ethelene, etc.)

Pathogen-induced structural defenses

(histological defense)

(An external, secondary tissue impermeable to water and gases. It is often formed in response to wounding or infection)

(An overgrowth of the protoplast of a parenchyma cell into an adjacent xylem vessel or tracheid)

(cellular defense)

-- A nipple-like structure deposited beneath the cell wall on the inside of a cell being attacked by a fungus, apparently serving as a defense mechanism against infection.

-- A common defense phenotype independent of the compatibility or incompatibility.

-- Cross-linking of cell wall components including polyphenolics (lignin-like), callose, proteins etc., thus resistant to common hydrolytic enzymes.

-- Generating the first active oxygen burst.

-- Roles in disease resistance

-- Many questions remain to be answered:

Pathogen-induced biochemical defenses

--Müller and Börger (1940) first termed Phytoalexin (from the greek, Phyton=plant and alexin=protecting substance).

--Pisatin, the first phyloalexin was isolated from pea(Pisum sativum) infected with Monilia fructicola by two Australia scientists Cruickshank and Perrin (1960). However, Japanese believed that it was ipomeamarone isolated from sweet potato infected by Ceratocystis fimbriata.

--It is currently defined as low molecular weight antimicrobial compounds that are both synthesized by and accumulated in plants which have been exposed to microorganisms

(A substance which inhibits the development of a micro-organism, produced in higher plants in response to certain stimuli (biological, chemical and physical).

--Classified mainly into two major groups:

Isoflavonids are common phytoalexins in legume family

Terpenoids are characteristic phytoalexins of Solanaceae such as potato, tomato tobacco and pepper.

--Many pathogenic fungi and bacteria can degrade phytoalexins, apparently mediating the toxicity of those compounds.

--Roles of phytoalexins in defense responses:

-- However, the defensive significance of these compounds is not clear for the majority of host-parasite systems.

-- Stakman (1915) is generally credited with the use of the term, hypersensitive reaction. Subsequently, Gaumann (1950s), Wood (1960s) and Tomiyama (1970s-1980s) made tremendous contributions.

-- Excessive sensitivity of plant tissues to certain pathogens. Affected cells are killed quickly, blocking the advance of obligate parasites

(A rapid local reaction of plant tissue to attack by a pathogen resulting in the death of tissue around infection sites preventing further spread of infection).

-- HR shows some similarity to animal Programmed Cell Death (PCD)

Programmed Cell Death (PCD) - activation of genetically regulated cell suicide machinery.

Apoptosis [derived from two Greek roots: apo(away) and ptosis(to fall), that was used to describe petals falling from flowers or leaves from trees].

Necrosis - in contrast to PCD, necrosis is most commonly defined as cell death that does not require the active participation of the cell in its own demise.

(the cause of HR; the role of HR)

-- These proteins were initially detected from tobacco after infection with TMV by van loon's group (1970).

-- Groups of proteins with typical chemical properties induced in plant tissues following pathogen infection. (extracellular location, extreme IEP and high stability)

-- Classically divided into 5 groups (PR-1, PR-2, PR-3, PR-4 and PR-5)

-- The putative function of these proteins

PR-1 -- unknown [Rauscher M, Adam AL, Wirtz S, Guggenheim R, Mendgen K, Deising HB., 1999. PR-1 protein inhibits the differentiation of rust infection hyphae in leaves of acquired resistant broad bean. Plant J. 19(6):625-33].

PR-2 -- beta-1,3-glucanase

PR-3 -- chitinase

PR-4 -- unknown

PR-5 -- thaumatin-like protein [Ibeas JI, Lee H, Damsz B, Prasad DT, Pardo JM, Hasegawa PM, Bressan RA, Narasimhan ML, 2000. Fungal cell wall phosphomannans facilitate the toxic activity of a plant PR-5 protein. Plant J 23:375-383]

-- The particular role as molecular markers in plant defenses.

-- Been Extensively used as resistance targets in transgenic studies.

-- Active oxygen species (AOS), a collective term for radicals and other nonradical but reactive species derived from oxygen, including O^-, HO^- and H₂O₂

Other term: Reactive Oxygen Intermediates (ROIs).

-- Doke's pioneer contribution in 1983

-- The oxidative burst required for

-- Mechanisms for generation of AOS

Using of pharmaceutical agents confirms the function of NADPH oxidase system in plant defense responses.

Mammalian NADPH oxidase homologues were identified in plant species.

However, this biochemical function of this system in plant defense responses hasn't been fully elucidated yet.

-- Nitric oxide (NO) functions as a signal in resistance response

It was just recently hypothesized that NO functions together with AOS participating in signaling of plant defense responses.

It is likely that this signal pathway exists in plant system.

However, no nitric oxide synthase (NOS) was predicted from reported plant DNA sequences. Plant may use an uncial system to generate NO.

-- More questions need to be answered

(Induced resistance develops after a primary infection (or treatment with certain chemicals) and its effective against a second infection. This phenomenon resembles acquired immunity in animals).

Induced resistance often generates a very broad spectrum against many types of pathogens.

Local Acquired Resistance (LAR)

ILR is apparently confined to the site of infection)

Systemic Acquired Resistance (SAR)

(-- SAR, which can be induced by a local infection (or by a local chemical treatment), provides the plants with long lasting, systemic resistance against a broad spectrum of pathogens).

-- SAR is a form of systemically induced disease resistance that is triggered upon infection by a necrotizing pathogen. The state of SAR is

characterized by an early increase in endogenously synthesized salicylic acid and the concomitant activation of genes encoding PR-proteins.

-- SAR has been demonstrated in many plant species and confers resistance against a broad spectrum of plant pathogens in distant, uninfected plant parts.

-- SA-dependent SAR pathway, which is induced upon pathogen infection.

-- SAR/SA marker proteins: PR-Proteins.

-- SA-nonaccumulating NahG plants (Ryan, 1994) expressing the bacterial SA hydroxylase gene NahG, are impaired in SAR.

-- Overproduction of SA in plant: Defense genes, particularly those encoding acidic pathogenesis-related (PR) proteins, were constitutively expressed in CSA plants. This expression did not affect the plant phenotype, but the CSA plants showed a resistance to viral and fungal infection resembling SAR in nontransgenic plants (Verberne et al., Nat Biotechnol 18:779, 2000).

Induced Systemic Resistance (ISR)

(Pieterse et al., Plant Cell 10:1571,1998)

-- JA-dependent ISR pathway, which is triggered by nonpathogenic Pseudomonas fluorescens (rhizobacteria).

-- In Arabidopsis, ISR is active against the fungal root pathogen Fusarium oxysporum f.sp. raphani, the oomycetous leaf pathogen Peronospora parasitica, the bacterial leaf pathogens Xanthomonas campestris pv. campestris and Pseudomonas syringae pv. tomato.

-- ISR/JA marker proteins: JA and ethylene have been shown to act in concert in activating genes

encoding defensive proteins, such as proteinase inhibitors and plant defensins.

-- ISR functions independently of SA and PR gene activation.

-- mutants: JA deficient and ethylene insensitive coil1, jar1 and ein2 mutants in Arabidopsis are impaired in the JA/ethylene signal transduction pathways.

Both SA and JA signaling are dependent on NPR1 pathway. Divergence of the SAR and the ISR pathway are downstream of NPR1. There are some conflicts on cross-talking between these two pathways.

Significance of SA and JA as endogenous signals:

-- Many other questions remain to be answered:

Engineering of Defense Responses

Many approaches have been developed in last five years. Several types of genes were transferred into plants and tested against pathogen attack including

Defensin (Wang et al., 1999)

PR-proteins (Borkowska et al., 1999)

Superparasite enzymes

Pto (Tobias et al., 1999)

Prf1

Xa21

Cf-9

Enhanced free-radical production;

Expression-hydrogen peroxide (Wu et al., 1995; 1999)

Antisense strategies: catalase (Molina et al., 1999)

Ion channels

Calmodulin (Harding et al., 1997; Harding & Roberts, 1998)

Npr1/cdr1

Elicitor/Avr-genes (Keller et al., 1999; McNellis et al., 1998; Wegener et al., 1996)

Protein kinase (Seo et al., 1999)

R-gene/Avr-gene

Advances:

Summary

The plant defense response represents one of the most quickly developing areas in plant biology.

Current genomic studies provide the most useful information in a genome-wide base of understanding plant resistance mechanisms.

These studies are important for theoretical biology of plants and for agricultural applications.

REFERENCES

PNAS, 1995, 92(10).

Plant Cell, 1996, 8(10).

Curr Opin Plant Biol. 1998, 1(4).

R.N., Goodman, Z., Kiraly and K.R., Wood, The Biochemistry and Physiology of Plant Disease, 1986, University of Missouri Press.

J.A. Callow, Biochemical Plant Pathology, John Wiley & Sons, 1983.

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