Major pathogenicity functions are encoded by hrp genes

In the beginning… it was observed that biotrophic

bacteria…

1. tended to be in contact with plant cell wall.

2. must be metabolically active to elicit disease

(or resistance).

In early 1980s- transposon tagging resulted in mutants that were unable to elicit a HR or acts a pathogen (hypersensitive response and pathogenicity).

The hrp- phenotype

Besides the above

· hrp mutants were unable to grow in planta

· but it is not required for growth on minimal media

plates – therefore they are not an auxotrophic mutant.

· Certain defence genes are still induced – phytoalexins

and PR proteins- an ancestral resistance mechanism?

Genetic organisation

It was noted that hrp functions were clustered.

P. s. pv. phaseolicola 22kb in eight transcriptional units

P.s. pv. syringae 25 eight

X.c. pv. campestris 23 six

Ralstonia solanacearum 25 seven

It was noted that avirulence genes were often linked to these

loci.

Note: hrp genes are not avirulence genes since isolated hrp genes

cannot elicit the hypersensitive response.

BUT – hrp + avirulence genes (on single clone- pHIRII) can elicit symptoms when introduced into a non-pathogen e.g. Pseudomonas fluorescens or E. coli.

Biochemical function

Database searches revealed that many hrp genes exhibited

homology to Type III secretion systems.

Type II

Sec –dependent pathway – involving a two step mechanism.

· Export to the periplasm. ATP-dependent.

Followed by cleavage of a N-terminal “segment”

by a signal peptidase.

· Passive export out of the outer membrane.

Type I

Sec-independent. A one step (ATP-dependent) mechanism

through a channel or a gated pore.

Type III

A sec-independent two-step mechanism. Commonly used

to deliver proteins from the bacterial cytoplasm to the

eukaryotic cytostol.

Key features

· Absence of signal peptide as used in sec pathway

· Requirement for host cell contact.

Hrp genes can be split into the following functional groups –

hrp A → outer membrane proteins → forms pilus-ike

structure.

One → outer membrane associated lipoprotein

(hrp C.)

Five → inner membrane associated proteins

(hrp RSTVJ)

Two → cytoplasmic proteins

(one is an ATPase, hrp N).

Seven of the nine hrp proteins exhibit homology to

other Type III particularly that found in Yersinia-

designated HR and conserved = hrc

Yersinia – Plague and gastroenteritis

Shigella - Dysentery

Salmonella – Food poisoning

E.coli - Diarrhoea

However : these pathosystems end with bacterial entry into the animal cell.

This is NOT the case with plant bacterial pathogens.

Hrp Gene Regulation

hrp gene expression influenced by Carbon/nitrogen source, pH,

osmolarity, temperature and possibly plant signal molecules.

Gene expression is dependent on hrpR and L products detecting

the metabolic status of the cell (“sated” and “hungry”) and interacting

with sigma 54 to bind to a regulatory region (the hrp box) and thereby

activating hrp gene transcription.

Hrp box – GGAACCNA – N₁₄ - CCACNNA

What is delivered?

Hrp –system is NOT required for soft-rot enzymes in

Erwinia carotovora or toxin genes in Pseudomonas

syringae or EPS genes in Ralstonia solanacearum!

What is delivered in analogous systems?

Yersinia – YopE/H : Interacts with actin cytoskeleton to

suppress phagocytosis.

Salmonella – SipA : Induces membrane ruffling by inhibiting

F-actin depolymerisation.

P. aeruginosa – ExoS : targets small GTP binding protein

exchange factor.

Surprisingly : “Avirulence” gene products are delivered.

So what are the virulence functions of these proteins?

· AvrBs2 exhibits homology to agrocinopine

synthase in Agrobacterium. Directs transformed

cells to produce a new carbon source- “cellular

metabolic reprogramming”?

· AvrBs3 localised to the nucleus where it may

activate the transcription of unknown genes.