Sudarshan Seshadri

My Story:

Thesis title:
Role of IkappaBzeta and Pyrin as Modulators of Macrophage Innate Immune Function

Abstract:
Innate immunity is the first line of defense mounted by the host against the pathogens. Innate immune responses are initiated at receptors that get activated upon pathogen recognition to turn on several proinflammatory pathways. The present study concentrated on understanding the role of two proteins, MAIL (IBzeta) in the signalosome pathway and pyrin in the inflammasome pathway. The signalosome pathway leads to the production of pro-inflammatory cytokines that are involved in host defense and host cell survival. On the other hand, the inflammasome pathway leads to the activation of caspase-1, an enzyme which has been implicated in processing and release of proIL-1, proIL-18 and proIL-33.
MAIL is a recently described homologue of IkBalpha that is rapidly induced by lipopolysaccharide (LPS) in monocytes. MAIL regulates the transcription of a number of inflammatory genes including IL-6 in the mouse. Although the role of IL-6 is well established in cancer and sepsis, the regulation of IL-6 in human monocytes and macrophages is poorly understood. Here we show that MAIL expression is suppressed with differentiation of monocytes to macrophages which correlates with IL-6 production. Furthermore, suppression of MAIL by small interfering RNA decreases LPS and Nod2 ligand induced IL-6 production. Our data suggests that MAIL is a key regulator of IL-6 production in monocytes and plays an important role in inflammation.
Dysregulation in IL-1 synthesis and production leads to many autoinflammatory diseases like familial Mediterranean fever (FMF). FMF is characterized by recurrent fever and inflammation and is caused by mutations in the pyrin gene (MEFV). Pyrin interacts with the proteins of the inflammasome and thus regulates its activity. However, the exact role of pyrin in the regulation of inflammasome assembly is poorly understood and a subject of controversy. Both activating and suppressing roles of pyrin have been proposed. In the studies outlined here, Pyrin levels in monocytes correlate positively with IL-1 processing and release. Macrophages are deficient in activating caspase-1, which correlates well with the decreased expression levels of pyrin. Suppression of pyrin by siRNA decreases IL-1 processing and release in THP-1 cells and in monocytes. In the case of an in vitro endotoxin model, pyrin tends to augment IL-1 processing and release.
The mechanism of how mutations of the pyrin gene causes increased inflammation has not been elucidated. In the present study, the role of the two common mutations, E148Q and M694V, in caspase-1 activation were analyzed. Both these mutant forms of pyrin were similar to wild type pyrin in co-localization and interaction with ASC. Importantly, the mutations in pyrin did not affect IL-1 processing and release in a HEK293 cells overexpression model. Our data suggests that FMF is not due to enhanced caspase-1 activation.
This study has elucidated the role of MAIL and pyrin in the regulation of two distinct pathways which are important for the innate host defense. Understanding the role of MAIL and pyrin in monocytes will be helpful in understanding mechanisms of inflammation and may open new avenues for therapeutic interventions in inflammation and inflammation induced diseases, such as cancer and sepsis.