Macrophages are responsible for defending against diverse pathogens and play a crucial role in the innate immune system. contribute to understanding the regulation of autophagy and apoptosis in macrophages, and shed lights on death receptor\targeted therapy for cancer, inflammation and autoimmune diseases. (Wei et?al., 2010). In this study, we find that TRAIL treatment influences death receptor expression in U937 cells, indicating that death receptor mediates TRAIL\induced apoptosis and autophagy in macrophages. These data further demonstrate that TRAIL plays an important role in innate immunity. Autophagy is a cell survival process involving macromolecule and organelle degradation. It has been reported that autophagy is connected to various physiological processes and an astonishing number of human diseases (Jostins et?al., 2012; Levine and Kroemer, 2008; Liu et?al., 2011; Mizushima et?al., 2008). A unique report on TRAIL\induced autophagy by Mills et?al. showed that TRAIL is required for the induction of autophagy during lumen formation (Mills et?al., 2004). Here we demonstrate that TRAIL induces both autophagy and apoptosis; inhibition of autophagy facilitates apoptosis in macrophages. These results suggest that TRAIL\induced autophagy is a cyto\protective mechanism, favoring stress adaptation and inhibiting cell death. TNF\R\mediated regulation of cell fate is closely related to the assembly of the DISC complex, which involves the aggregation of the intracellular domain of the death receptor, caspase\8, FADD, TRADD and others. (Cao et?al., 2011; Vanlangenakker et?al., 2011; Zhang et?al., 2011). A recent study shows that RIP1\dependent signal transduction pathways are involved in regulating cell survival, apoptosis and necrosis (Festjens et?al., 2007). In these pathways, as in TNF\R\mediated signaling, RIP1 is positioned at the center of cell\fate decisions; survival, apoptosis or necroptosis pathways are followed by the formation of complex I, complex II or the necrosome, respectively (Micheau and Tschopp, 2003; Rothe et?al., 1995). In complex I (TRADD, RIP1, TRAF2, etc.), RIP1 quickly recruits IKK complex and activates NF\B. RIP1 and NEMO can also form a stable complex with a linear 196868-63-0 IC50 ubiquitin chain, thereby inhibiting cell death (Haas et?al., 2009; Tokunaga et?al., 2009). If RIP1 is not ubiquitinated, the complex I (TRADD\RIP1\TRAF2) is dissociated from the death receptor to allow FADD and caspases to bind and cause cell death by apoptosis (Bertrand et?al., 2008; Petersen et?al., 2007). When caspase activation is inhibited by viral infection, RIP1 and RIP3 induce necroptosis (Vandenabeele et?al., 2010). We show here that the dynamic disintegration of RIP1 expression and deubiquitination suppress? autophagy and increase apoptosis in TRAIL\treated macrophages. This result suggests that the ubiquitination status of RIP1 may tune its activity in different signaling pathways. Our observations provide new CDX2 evidence that RIP1 plays a critical role in the regulation of death receptor mediated conversion of autophagy to apoptosis in macrophages. Beclin 1, the mammalian orthologue of yeast Atg6, plays a central role in autophagy (Liang et?al., 1998; Wang, 2008). We observed that knockdown of RIP1 suppresses the expression of Beclin 1 during TRAIL\induced autophagy and apoptosis, suggesting that Beclin 1 is a downstream modulator of RIP1 signaling. It is known that both RIP1 and Beclin 1 are substrates of caspase\8 and that caspase\mediated cleavage of Beclin 1 promotes crosstalk between apoptosis and autophagy (Djavaheri\Mergny et?al., 2010; Kang et?al., 2011). Moreover, Cho et?al. report that TRAIL can trigger the caspase\mediated cleavage of Beclin 1 in HeLa cells (Cho et?al., 2009a). Another study (Hou et?al., 2010) suggests that caspase\8 activity in the TRAIL\mediated autophagic response is counter\balanced by the TRAIL\mediated apoptotic response; the proposed mechanism involves continuous sequestration of the large caspase\8 subunit in the autophagosomes of Bax?/? HCT116 colon cancer cells, which supports the existence of a feedback mechanism that cross\regulates autophagy and apoptosis. Further clarification of the mechanism and downstream targets of Beclin 1 in the autophagy\apoptosis shift would be valuable for the development of novel therapeutic strategies for the treatment of cancer and other diseases. In summary, we have found that TRAIL induces both autophagy and apoptosis in macrophages, with the inhibition of autophagy 196868-63-0 IC50 significantly enhancing apoptosis. TRAIL treatment increased DR4 expression, decreased DR5 expression, indicating that TRAIL\induced apoptosis and autophagy are mediated by death 196868-63-0 IC50 receptors in macrophages. RIP1 expression and ubiquitination are dynamically regulated, and the phosphorylation of IKK\, IKK\ (NEMO), and IB is strikingly decreased in the TRAIL\induced conversion of autophagy to apoptosis. Knockdown of RIP1 suppresses the expression of LC3\II, which is the hallmark of autophagy. In.