Effect of cagA on ASPP2 in Helicobacter pylori associated gastritis

1. Effect of cagA on ASPP2 in Helicobacter pylori associated gastritis

2. Helicobacter pylori is a spiral shaped gram negative microaerophilic bacillus. It is one of the most common human pathogens. It is present nearly in about 50% of the world wide population (the incidence is about 20%in developed countries and more than 60% in the developing ones). The highest incidence recorded in Asia, Africa and south America.

3. H.pylori colonize the human stomach and persist for several decades causing chronic gastritis and peptic ulcer diseases. It is also considered as an important risk factor for development of gastric carcinoma. For this reason, H.pylori was classified as type I carcinogen by International Agency for Research on Cancer (IARC).

6. H.Pylori by Giemsa

7. There are many strains of H.pylori, but generally H.pylori can be classified to only two categories either cagA+ve or –ve groups according to a specific gene in the bacteria was discovered in 1989 called cag (cytotoxin associated gene) that encodes for a specific antigen called cagA. More than 90% of the isolated cagA+ve strains from east Asia including Korea, Japan and China (where the highest incidence of gastric cancer).

8. Intracellular bacterial and viral pathogens that established persistent infections have evolved multiple strategies to interfere with host signaling pathways and immune responses. they do so to create an intracellular safe heaven while avoiding killing the host cell. Several viral oncogenes inactivate the tumor suppressor activity of the host cell by targeting and degrading p53 when infection is established.

9. Although H.pylori is not an intracellular pathogen, its interactions with host cells may provide certain survival benefits for the bacterium and contribute to establishment of chronic infection. Hp uses host plasma cell membrane as a site for replication and formation of microcolonies. Colonization of the stomach by Hp is greatly enhanced when the apoptotic response of the gastric epithelial cells is impaired which is done by cagA that induce up-regulation of the antiapoptotic protein MCL-1 ,so the affected cells become more resistant to the normal cell turn over in the stomach. Hp cause down-regulation of p53 indirectly through interaction with ASPP2 proapoptotic gene.

10. Here we will discuss the role of cagA on p53 through its interaction with ASPP2 but firstly we should know how cagA is delivered from H.pylori to inside the host cell. There are many secretion systems by which the organism inject its proteins inside the host cell. Gram negative bacteria use six different secretion systems. H.pylori use type IV secretion system.

11. Type IV secretion system (T4SS or TFSS) It is homologous to conjugation machinery of bacteria. It is capable of transporting both DNA and proteins. It was discovered in Agrobacterium tumefaciens, which uses this system to introduce the T-DNA portion of the Ti plasmid into the plant host, which in turn causes the affected area to develop into a crown gall (tumor).

12. Helicobacter pylori uses a type IV secretion system to deliver cagA into gastric epithelial cells, which is associated with gastric carcinogenesis. Bortedella pertussis, the causative agent of whooping cough, secretes the pertussis toxin partly through the type IV system. Protein members of this family are components of the type IV secretion system. They mediate intracellular transfer of macromolecules via a mechanism ancestrally related to that of bacterial conjugation machineries.

15. Function In short, Type IV secretion system (T4SS), is the general mechanism by which bacterial cells secrete or take up macromolecules. Their precise mechanism remains unknown. T4SS are cell envelope-spanning complexes or in other words 11-13 core proteins that form a channel through which DNA and proteins can travel from the cytoplasm of the donor cell to the cytoplasm of the recipient cell.

16. Now cagA is inside the gastric cell, so what is cagA? cagA is directly translocated from H.pylori into the gastric epithelial cells via TFSS or type IV secretion system and localize to the host plasma membrane where the full length protein interacts with components of the apical junctional complex of the cell including E-cadherine, Jam that promote loss of cell polarity and enhance invasiveness of the cell.

17. C-terminal and N- terminal domains of cagA both are required for full activity of the protein. Expression of C-terminal domain alone of cagA initiate receptor tyrosine kinase signaling and cause scattering of the cell. However, the junctional complexes of the cell remain intact and the cell does not become migratory or invasive.

18. In absence of N- terminal domain, the C- domain of cagA is primarily localize to the cytoplasm. While co-expression of both C and N-domains induce a strong accumulation of both domains near the plasma membrane, suggesting the role of N-domain to target cagA toward plasma membrane of the affected cell. CagA modulate the activity of ASPP2 by simply binding its N domain to the ASPP2protein.

19. After this interaction; ASPP2 recruits and binds cytoplasmic p53 which is then degraded by proteisomes. CagA mediated degradation of p53 results in resistance in the apoptotic response in an ASPP2-dependent manner .

20. ASPP:-(apoptosis stimulating protein of p53)

22. ASPP are family of proteins that include ASPP1, ASPP2 and iASPP. Both ASPP1&2 are proapoptotic proteins that activate p53 while iASPP inhibite the apoptotic function of p53. ASPP2 is expressed in many human tissues such as heart, brain, placenta, lung, liver, skeletal muscle, kidney, pancreas, but at varying levels. The highest expression level of ASPP2 was detected in skeletal tissue

23. On DNA damage, cellular stress or oncogenic stimuli; the C-terminal domain of ASPP2 transiently interact with the DNA binding domain of p53 which in turn induce expression of genes involved in apoptosis like BAX& PUMA and PIG3, all of which are positive regulators of programmed cell death.. ASPP2 also bind to other factors that play a role in apoptosis as Bcl2 and Yes-associated protein. And factors that control cell growth as p65 and APC tumor suppressor.

24. The C terminus of ASPP2 interacts with the DNA-binding domain of p53, but it is unclear how this interaction leads to activation of p53. The N terminus of ASPP2 is required to enhance the transcriptional activity of p53, suggesting that this domain may have a regulatory role and determine the outcome of the ASPP2-p53 interaction . Posttranslational modification of the N terminus of ASPP2, as well as an alteration in the set of proteins that interact with this domain, may determine the fate of p53 and thereby, affect the apoptotic response. For example, cytosolic DDA3 binds ASPP2 and prevents activation of p53 without affecting the ASPP2-p53 interaction. The function of ASPP2 is not restricted to activation of p53 but includes regulation of cell–cell adhesion and polarity.

25. We did not detect the formation of a ternary complex between cagA, ASPP2 and p53 perhaps owing to a transient nature of any interactions and also unclear whether the associations between cagA-ASPP2 and ASPP2-p53 occur simultaneously or sequentially and in which cytosolic region the ASPP3-p53 complex is formed.