B Y A L E X P O L I D O R E

TYPE III SECRETION SYSTEM

BACKGROUND OF SECRETION SYSTEMS

• There are six different types of secretion systems in bacteria

• Type I, Type II, Type III, Type IV, Type V, and Type VI

• Type I and IV are found in both gram-negative and gram-positive bacteria

• All other types of secretion systems are found only in gram-negative

bacteria

• Type III secretion system (TTSS) is responsible for interactions

with host cell membranes in which virulence factors are

injected directly in to the host cell

• Knowledge about TTSS is important to understanding bacterial pathogenesis and developing possible antibiotics which target TTSS components

• Also note that Flagella and TTSS share very similar cellular components and structure, but do not function in a similar way

GRAM-NEGATIVE VS. GRAM-POSITIVE

BACKGROUND OF TYPE III SECRETION SYSTEM (TTSS)

• Contains more than 20 proteins that make up the

apparatus

• Most complex protein secretion system known in

bacteria

• Components of the TTSS’s are highly conserved,

however different TTSS’s release unique effectors

despite this conservation

• Evolutionarily related to bacterial flagellum

• TTSS operons found in plasmids or in the genome or

both

Ref. 2

TYPE OF PROTEINS INVOLVED IN TTSS

• Structural proteins

• Compose the basal body, inner rod, needle, and the bulb

• Effector proteins

• The proteins that are secreted by the apparatus into host

• Chaperone proteins

• Bind effectors to prepare them for secretion and target them to

the TTS-apparatus

STRUCTURAL COMPONENTS OF TTSS

• Basal body

• Outer rings (OR1 and OR2)

• Inner rings (IR1 and IR2)

• Needle complex

• Inner rod

• Needle

• Translocator

• Bulb

• ATPase

• Chaperones

• Other cytoplasmic

associated proteinsRef. 4

Ref. 11

ASSEMBLY OF TTSS

1) Base structure is assembled first via a sec pathway (socket,

septum, outer rings, inner rings, and export apparatus)

2) Then needle complex is assembled (addition of inner rod and

needle proteins)

Ref. 7 Ref. 9

ASSEMBLY OF TTSS

Ref. 7Ref. 13

Ref. 13

MATURATION OF THE TTS-APPARATUS

• The needle complex switches substrate affinity from

structure proteins to effector molecules

• Termination of inner rod and firm anchoring or needle leads

to downward shift in cup-like protrusion in bulb cytoplasmic face and other conformational changes in the inner and

outer rings

• Now the secretion system is in its mature form and is

ready to secrete effector molecules such as virulence

factors into host cells

TARGETING PROTEINS FOR TTSS

• Contain a 20-30 amino acid signal sequence in the N-terminal region of the polypeptide

• These signal sequences vary depending on the effector

• Unlike sec-dependent signal sequences, TTSS signals are not cleaved upon secretion

• Some evidence suggests the secretion signal located in the coding messenger RNA

• Much debate over where the signal sequence lies, but in at least some TTSS proteins the secretion signal does in fact reside within the amino acid sequence and not the mRNA

• A second method of targeting effectors to TTSS involves the accessory proteins called chaperones

ROLE OF CHAPERONES IN TARGETING PROTEINS TO TTSS

• TTSS chaperones are small, acidic, dimeric proteins

that lack ATP-binding or ATP-hydrolyzing activities

• Share a common crystal structure

• Bind downstream of the N-terminal secretion signal

Ref. 7

ROLE OF CHAPERONES IN TARGETING PROTEINS TO TTSS

• Functions of chaperone proteins:

• Prime secreted proteins for unfolding before secretion

• Prevent interactions with other components of the TTSS machinery

• Target secreted proteins to the desired TTS-apparatus

• Chaperone proteins contain a conserved motif

• This motif is a good target for pharmacological disruption

Ref. 1

ROLE OF ATPASE IN MODIFYING SECRETED PROTEIN

• The chaperone-effector complex cannot be transported through the needle

• Modification by ATPase activity of this complex occurs before secretion

• Functions of ATPase associated with the TTSS

• Recognizes chaperone-effector complex by a C-terminal domain on the chaperone

• Releases effector protein from chaperone using ATP hydrolysis

• Unfolds the effector domains in

Ref. 10

MODEL OF ATPASE FUNCTION AND SECRETION OF PROTEIN

Ref. 10

ROLE OF TRANSLOCATORS IN SECRETION OF PROTEINS

• Much unknown to how exactly the needle delivers effectors into the target cells

• A current model involves translocators

• Function of translocatorproteins:

• Insert into host cell membrane forming a channel

• Recruits the needle (needle docks onto the channel)

Ref. 7

REGULATION OF TTSS

• Regulatory mechanisms specific for each TTSS

• One hypothesis involves the needle as a sensor for cell

contact

• Needle senses host cell

• Transduces a signal to the cytoplasmic side of the TTSS

• Bulb structure “opens” allowing effector proteins to pass through the

needle

• Regulatory proteins ensure the system is reset and the

system is re-loaded with the same effector proteins or new

ones

HOW EFFECTORS INTERACT WITH HOST

• Effector proteins have been optimized through

evolution to suit the bacteria’s specific needs

• Effectors delivered by the TTSS can effect many

cellular functions such as actin and tubulin dynamics,

gene expression, vesicular trafficking, apoptosis, and

cell cycle progression

• General theme involving effector proteins is mimicry

• TTSS effector proteins mimic the function of host cell

proteins

THE FUTURE OF TTSS

• Type III secretion systems are widespread and diverse and

function to interact pathogenic (or symbiotic) bacteria to

their eukaryotic hosts

• This central role of TTSS allows for the possibility of

developing effective anti-infective strategies

• These systems may also be used for therapeutic purposes

in drug or vaccine delivery

• Much is still unknown about these systems and their

specific functions in different bacteria, but the

importance and fascination of these machines will likely

propel research in the study of these secretion systems

REFERENCES

1. Lilic M., Vujanac M., Stebbins C.E. A common structural motif in the binding of virulence factors to bacterial secretion chaperones (2006) Molecular Cell, 21 (5), pp. 653-664.

2. Dieye, Y., Ameiss, K., Mellata, M., & Curtiss, R. (2009). The salmonella pathogenicity island (spi) 1 contributes more than spi2 to the colonization of the chicken by salmonella enterica serovar typhimurium. BMC Microbiology, 9(3), 1-14.

3. Sory, M. P., Boland, A., Lambermont, I., & Cornelis, G. R. (1995). Identification of the yope and yoph domains required for secretion and internalization into the cytosol of macrophages, using the cyaa gene fusion approach. Proceedings of the National Academy of Sciences of the United States of America, 92(26), 11998-12002.

4. Tampakaki, A. P., Fadouloglou,, V. E., Gazi, A. D., Panopoulos, N. J., & Kokkinidis, M. (2004). Conserved features of type iii secretion. Cellular Microbiology, 6(9), 805-816.

5. Andrade, A., Pardo, J. P., Espinosa, N., Pe´rez-Herna´ndez, G., & Gonza´lez-Pedrajo, B. (2007) Enzymatic characterization of the enteropathogenic Escherichia coli type III secretion ATPase EscN. Archives of biochemistry and biophysics, 468,121-127.

6. Blocker, A., Komoriya, K., & Aizawa, S. I. (2003). Type iii secretion systems and bacterial flagella: Insights into their function from structural similarities. PNAS, 100(6), 3027-3030.

7. Galan, J. E., & Watz, H. W. (2006). Protein delivery into eukaryotic cells by type iii secretion machines. Nature, 444(30), 567-573.

8. Galan, J. E., & Lee, S. H. (2004). Salmonella type iii secretion-associated chaperones confer secretion-pathway specificity. Molecular Microbiology, 51(2), 483-495.

9. Marlovits, T. C., Kubori, T., Tejero, M. L., Thomas, D., Unger, V. M., & Galan, J. E. (2006). Assembly of the inner rod determines needle length in the type iii secretion injectisome. Nature, 441(1), 637-640.

10. Akeda, Y., & Galan, J. E. (2005). Chaperone release and unfolding of substrates in type iii secretion. Nature, 437(6), 911-915.

11. Aizawa, S. I. (2001). Bacterial £agella and type iii secretion systems. FEMS Microbiology Letters, 202, 157-164.

12. Journet, L., Hughes, K. T., & Cornelis, G. R. (2004). Type iii secretion: a secretory pathway serving both motility and virulence (review). Molecular Membrane Biology, 22(1-2), 41-50.

13. Marlovits, T. C., Kubori, T., Sukhan, A., Thomas, D. R., Galan, J. E., & Unger, V. M. (2004). Structural insights into the assembly of the type iii secretion needle complex. Science, 306(5698), 1040-1042.