MIC 4124 7a - Bacterial Effector Proteins

Simplistic overview of a bacterial infection

- Exposure to pathogens

- Adherence to skin or mucosa

- Invasion through epithelium

- Colonization and Growth (production of virulence factors)

- Tissue damage and disease

Reality of bacterial infections

Virulence Factors mediate infection and are expressed throughout entire course of infection. Are complex

Complexity of virulence factors

- Function by interacting with host; often modify host function for the benefit of pathogen

- Expression is dynamic; different virulence factors are made at different timepoints of infection for different purposes

Strategies to establish infection (3)

secretion of virulence factors...

- Extracellular: within interstitial space

- Intracellular (vacuoles): remain in compartments

- Intracellular (cytosolic): break free from compartments; live within our cytosol

Where does E. Coli secrete virulence factors?

Extracellularly (within interstitial space)

Where does Salmonella enterica and Legionella pneumophilia secrete virulence factors?

Intracellular (vacuoles): remain in compartments

Where does Listeria monocytogenes secrete virulence factors?

Intracellular (cytosolic): break free from compartments; live within our cytosol

The most successful virulence factors...

are often not the most toxic so that it can manipulate the host without killing the host

How Do Bacterial Pathogens Cause Infection?

- Toxins (endotoxins and exotoxins)

- Other virulence proteins (effectors)

Endotoxins

Normal constituents of bacterial cell wall (ex, LPS)

- Active only after released: during cell division or bacterial cell lysis

Exotoxins

Protein synthesized & secreted by the pathogen

- Cause damage to host cells

- Susceptible to antibodies, therefore are highly immunogenic

- Diverse structures

- A pathogen may secrete more than one different toxin

Endotoxins vs. Exotoxins

- Endotoxins exhibit relatively low toxicity compared to exotoxins (but can be fatal at high doses)

- Endotoxins exhibit more non-specific effects (ex, fever, aches, shock) than exotoxins

What are bacterial effectors?

Specialized virulence proteins that are injected into host cells

Bacterial effectors mode of action

- Secreted

- Manipulate host pathways to mediate infection

- Usually not overtly toxic

Effector protein function

manipulate host cell pathways to facilitate infection

- Evolved to function in the host cell, not the bacterial cell

How do effector proteins enter the host cell?

specialized bacterial secretion systems that must pass bacterial cell wall and host cell membrane

- Ex: T3SS (needle-like apparatus)

Why are bacterial effectors considered dynamic?

effectors are expressed at different timepoints of infection, and evolved to function only during a limited time

- ex: effectors made immediately after infection are often different than those made during late infection

What do bacterial effectors target?

host protein or protein complex; host-pathogen protein-protein interaction to alter function of host protein

- diverse cellular processes

Examples of complex mechanisms of effectors

- Some can mimic mammalian proteins to manipulate host cell processes

- Some can have enzymatic activity on host proteins

- Some can have multiple domains with different functions

Co-operative nature of effectors

Effectors are often co-operative

- several effectors co-evolve to target different host proteins in same pathway

Purpose of bacterial effectors

Usually, to aid bacterial survival and/or replication in the host, without causing death

Salmonella effector proteins example

Effectors make it possible for Salmonella to get inside many different cell types for infection

- salmonella is an intracellular microbe that can enter any cell in the body. it uses effector proteins to enter any non-phagocytic cell

How does Salmonella enter a non-phagocytic cell?

Salmonella will bind the host cell and inject effector proteins into the host cell, these proteins will change the regulation of actin and cause a movement

Listeria effector proteins example

Listeria is an intracellular microbe that uses a single effector to polymerize host actin to give it motility (for replication & spread)

Are bacterial effectors only relevant in human pathogens?

No, plant and animal pathogens use them as well

Plant pathogen bacterial effector example

Pseudomonas syringae infects agriculturally important crops such as tomato, tobacco & beans

- effectors help evade host immunity

Animal pathogen bacterial effector example

Pathogenic E. coli strains causes respiratory and reproductive diseases in chickens, pigs and cows

- effectors help mediate infection

Other roles of bacterial effectors (2)

Mediate symbiotic relationships between organisms

- Nitrogen fixation

- Microbial Competition

Nitrogen fixation and effector proteins

Some nitrogen-fixing bacteria in root nodules use effectors to promote nodulation

Microbial competition and effector proteins

T6SS effectors often play roles in killing competing microbes

- Bacterial Pathogen Serratia marcescens

- Candida albicans & other fungal strains

Components of the host cell that bacterial effectors target (6)

Diverse Cell Processes

- Tight Junctions

- Biochemical Activities

- Cytoskeleton (Actin and microtubules)

- Protein Degradation

- Cellular Defense Pathways (Endocytic Pathway, Autophagy, Immune Cell Function)

- Organelle Function (Mitochondria, Golgi, Nucleus)

Bacterial effector targets studied in lecture (6)

- Host small GTPases

- Membrane lipids

- Innate and adaptive immunity

- Host cytoskeleton

- Nuclear function

- Autophagy

Host Small GTPases

Small GTPases that belong to the Ras superfamily control important host cell processes

- Target for bacterial effectors

- Subclassified into 5 families (Ras, Rab, Rho, Arf and Ran); >150 members

What host cell processes to small GTPases control?

response pathways to infection, intracellular trafficking, and cytoskeletal reorganization, etc.

Intracellular trafficking

the process of how intracellular organelles move around

How do GTPases help define membrane identity in host cells?

Within host cells, compartment membranes have unique GTPase compositions

- this allows researchers to identify and study them

Host Small GTPases and bacterial effectors

effectors can target all 5 families of GTPases

→ important targets for infection

Ras Superfamily GTPases

small G-proteins that function as molecular switches for signaling pathways

Two forms of GTPases

Cycle between active (GTP-bound) and inactive (GDP-bound) forms

Only when GTPases are active...

do they interact with host binding partners to mediate cellular effects

Regulatory proteins that associate with GTPases

GEF and GAP

GEF

guanine nucleotide exchange factor → activates GTPase

GAP

GTPase activating protein (enzyme) → deactivates GTPase

Membrane lipids and bacterial effectors

Bacterial effectors often bind/target membrane lipids

- Membrane lipids such as phosphoinosides (PIs) are important to organelle function

How are membrane lipids defined membrane identity in host cell?

Intracellular compartments have unique membrane lipid compositions of PIs

PI

Phosphoinosides, a membrane lipid

Phosphoinosides (PI) role in host cell (3)

Recruit host proteins to mediate function

- Organelle trafficking

- Secretion

- Compartment fusion

When bacterial effectors bind PIs what happens?

- effector recruitment to compartments

- manipulate PI content at compartments

- manipulate host proteins that are endogenously recruited to membranes by PIs

What type of immunity do bacterial effectors target?

Innate and adaptive

- Bacterial pathogens use effectors to either reverse or finely tune immune responses for the benefit of the pathogen

NF-𝜅B pathway role

plays a role in the innate immune response to infection

Bacterial effectors that target NF-𝜅B pathway

- E. coli: NleE1, NleB1, NleC, and NleH

- S. flexneri: IpaH family

- Yersinia: YopJ

E. coli effector NleC

a metalloprotease that cleaves p65 to inactivate it → inhibits NF-𝜅B pathway

Cytoskeleton role

- maintains structure of the cell

- endocytosis

- trafficking

- cell division

Components of the host cell cytoskeleton (3)

Actin, microtubules, intermediate filaments

Bacterial effectors can manipulate the cytoskeleton to... (4)

- Induce pathogen uptake into host cells

- Manipulate organelle movement (and often function)

- Create a means of bacterial motility within host cells

- Mediate cell-to-cell spread

Example of bacterial effector action on host cytoskeleton

Listeria effector ActA mimics a host protein that regulates actin polymerization

→ allows motility within host cell

→ uses motility for cell-to-cell spread

Nucleus role in host

important in cell function (gene expression)

How does a bacterial infection alter nuclear function?

it can trigger gene transcription in the host to deal with the stress of infection

Ways that bacterial effectors can manipulate nuclear function (3)

- Suppress transcription of genes for host response to infection

- Induce expression of host proteins that might benefit the pathogen

- manipulate the cell cycle to benefit the pathogen

Example of bacterial effector manipulating nuclear function

Shigella Infection (gastrointestinal function)

- to penetrate the frequently shedding intestinal epithelium the bacterial effector protein IpaB sequesters host protein Mad2L2 to stall the cell cycle and block epithelial cell shedding

- now shigella can penetrate the intestinal epithelium and cause disease

Mad2L2

Host protein that functions with the anaphase-promoting complex (APC) to allow progression to mitosis

Autophagy

cellular pathway to remove unneeded/dysfunctional cell components or invading pathogens

- targets organelles or pathogens for degradation

- includes intracellular bacteria (xenophagy)

How do pathogens manipulate autophagy?

- Suppress antibacterial autophagy to help the pathogen survive

- Activate autophagic degradation of cell components to gain nutrients to survive

Examples of effectors that suppress antibacterial autophagy

S. enterica - SopF

S. enterica - AvrA

L. pneumophila - RavZ

Examples of effectors that activate autophagic degradation of cell components

Coxiella burnetii - CvpB

Burkholderia pseudomallei - BPSS0180

How many effector proteins does Salmonella enterica have?

>30

How many effector proteins does Legionella pneumophila have?

>300