Methods br Results br Discussion In this study
Discussion In this study we make the novel observation that villin-1 and gelsolin are down-regulated in response to diverse cellular stressors, including bacteria associated with the pathogenesis of CD. The high plasticity and dynamics of the Tylosin tartrate australia cytoskeleton are exploited by bacterial and viral pathogens to enter host cells. Not surprisingly then, changes in the expression and/or loss of actin severing functions of villin-1 and gelsolin have been shown to bolster the immune response by limiting pathogen entry into mammalian cells.8, 26, 35 We provide a molecular basis for this function of villin-1 and gelsolin by identifying them as targets of EIF2A signaling, which we suggest serves to achieve broad anti-viral and anti-bacterial protection. Our study establishes that changes in actin dynamics are crucial determinants of cell fate during cellular adaptation to stress. Villin-1 and gelsolin serve as biosensors of cellular stress to integrate environmental sensing pathways with determinants of cell fate. The dynamic conversion of cellular G- and F-actin levels allows rapid restoration of cellular homeostasis through the regulation of PP1.21, 36 G-actin binding to PP1 and its regulatory subunits stabilizes the phosphatase activity of PP1, which dephosphorylates pEIF2A and restores homeostasis.4, 37 We confirm the role of villin-1 and gelsolin in regulating the association of G-actin with PP1. A good measure of cell fate during cell stress may be this ability of PP1 to bind G-actin or not. We also demonstrate that stabilization of F-actin is the cue for initiating cell death, and we suggest that a cell measures the amount of damage during stress by whether cellular F-actin levels exceed their tolerance limits. The limits of F-actin levels required to initiate cell death may be unique to different cell types and may be achieved by tissue-specific actin regulatory proteins. We suggest that changes in cellular F-actin levels could provide a quantitative marker of cellular damage. This also confirms that actin-modifying proteins have important signaling properties that influence cell fate. It may be noted that while advillin has been identified as a specific target of DDIT3, its function in EIF2A signaling remains unknown. We propose that, like its homologs villin-1 and gelsolin, changes in advillin activity in response to cellular stress could regulate constitutive phosphorylation of EIF2A to prevent recovery from cellular stress. Advillin is an actin-bundling protein and pEIF2A signaling increases advillin expression, which is expected to increase cellular F-actin levels to induce cell death downstream of DDIT3 activation. Similarly, down-regulation of cofilin-1 increases cellular F-actin levels and induces cell death downstream of DDIT3 activation. Thus, changes in cellular actin dynamics may be more widely used as an adaptive response to cellular stress. More importantly, this regulatory EIF2A-actin pathway is highly conserved and intersects with signaling pathways pivotal to inflammation and disease pathogenesis. IRGM plays a role in innate immunity by regulating autophagy in response to several intracellular pathogens. Exposure to microbial products or bacterial invasion increases IRGM expression and IRGM guides autophagic clearance of these bacteria. Based on that, the thinking has been that by direct elimination of intracellular bacteria and preventing the activation of pattern recognition receptor signaling, IRGM regulates gut homeostasis. In CD a single nucleotide polymorphism (SNP) lying upstream of the IRGM gene in the non-coding region results in sustained increase not decrease in IRGM protein expression and, more importantly, despite the high levels of IRGM in CD patients, it is accompanied by an increase in intracellular bacteria. So the role of IRGM in the etiology of CD is unknown. Our studies provide for the first time a molecular basis for the paradoxical function of IRGM in CD. Our study highlights the autophagy-independent function of IRGM in inducing necroptosis, releasing DAMPs, and initiating intestinal inflammation, thus providing a molecular basis for IRGM in the pathogenesis of CD.