The cellular response to unfolded and misfolded proteins in the mitochondrial matrix is poorly understood. Here, we report on a genome-wide RNAi-based screen for genes that signal the mitochondrial unfolded protein response (UPR(mt)) in C. elegans. Unfolded protein stress in the mitochondria correlates with complex formation between a homeodomain-containing transcription factor DVE-1 and the small ubiquitin-like protein UBL-5, both of which are encoded by genes required for signaling the UPR(mt). Activation of the UPR(mt) correlates temporally and spatially with nuclear redistribution of DVE-1 and with its enhanced binding to the promoters of mitochondrial chaperone genes. These events and the downstream UPR(mt) are attenuated in animals with reduced activity of clpp-1, which encodes a mitochondrial matrix protease homologous to bacterial ClpP. As ClpP is known to function in the bacterial heat-shock response, our findings suggest that eukaryotes utilize component(s) from the protomitochondrial symbiont to signal the UPR(mt).

The generation of biologically active proteins by regulated intramembrane proteolysis is a highly conserved mechanism in cell signaling. Presenilin-dependent ␥-secretase activity is responsible for the intramembrane proteolysis of selected type I membrane proteins, including ␤-amyloid precursor protein (APP) and Notch. A small fraction of intracellular domains derived from both APP and Notch translocates to and appears to function in the nucleus, suggesting a generic role for ␥-secretase cleavage in nuclear signaling. Here we show that the p75 neurotrophin receptor (p75 NTR ) undergoes presenilin-dependent intramembrane proteolysis to yield the soluble p75-intracellular domain. The p75 NTR is a multifunctional type I membrane protein that promotes neurotrophininduced neuronal survival and differentiation by forming a heteromeric co-receptor complex with the Trk receptors. Mass spectrometric analysis revealed that ␥-secretase-mediated cleavage of p75 NTR occurs at a position located in the middle of the transmembrane (TM) domain, which is reminiscent of the amyloid ␤-peptide 40 (A␤40) cleavage of APP and is topologically distinct from the major TM cleavage site of Notch 1. Size exclusion chromatography and co-immunoprecipitation analyses revealed that TrkA forms a molecular complex together with either full-length p75 or membrane-tethered C-terminal fragments. The p75-ICD was not recruited into the TrkAcontaining high molecular weight complex, indicating that ␥-secretase-mediated removal of the p75 TM domain may perturb the interaction with TrkA. Independent of the possible nuclear function, our studies suggest that ␥-secretase-mediated p75 NTR proteolysis plays a role in the formation/disassembly of the p75-TrkA receptor complex by regulating the availability of the p75 TM domain that is required for this interaction.The p75 neurotrophin receptor (p75 NTR

P58/DNAJc3 defends cells against endoplasmic reticulum (ER) stress. Most P58 molecules are translocated into the ER lumen, and here we report selective and stable binding to misfolded proteins by P58's TPR-containing N-terminal domain. In vitro, too, P58 binds selectively to a model misfolded protein and challenge of that complex with physiological concentrations of the ER lumenal Hsp70-type chaperone BiP encourages disassembly. BiP-induced dissociation of P58 from its substrate depends on the presence of ATP and on interactions with P58's J-domain, which are mediated by invariant residues BiP R197 and P58 H422 . A functional J-domain also accelerates dissociation of P58 from a model substrate, VSV-G ts045 , on the latter's re-folding in vivo. However, J-domain binding can be separated from the ability to promote substrate dissociation by the mutant BiP E201G and a wild-type J-domain fused ectopically to P58 H422Q rescues the latter's inability to dissociate from substrate in response to BiP and ATP. These findings are consistent with a model whereby localized activation of the Hsp70-type partner is sufficient to promote substrate handover from the J-domain cochaperone.

IntroductionIn the steady state, dendritic cells (DCs) capture self-antigens and maintain low expression of costimulatory molecules but nevertheless migrate to lymph nodes where they tolerize self-reactive T cells. [1][2][3][4][5][6][7] Physiologically arising apoptotic cells (ACs) are one significant source of self-antigens for DCs. 5,7 However, following their encounter with ACs, DCs can be rendered immunologically inert, immunosuppressive, or immunostimulatory. [8][9][10][11][12][13][14][15] These contradictory findings have been difficult to resolve due to the complexity of receptors on DCs and the model systems used (eg, ex vivo versus in vivo or rodent versus human sources of ACs).ACs are recognized and captured by human DCs via an array of receptors, including LOX-1, CD36, ␣v␤3, ␣v␤5, and the complement receptors (CRs) CR3 and CR4. 16 The precise contribution of individual receptors in the binding/uptake of ACs, the initiation of downstream signaling pathways, and cross-presentation of cellassociated antigens remains undefined. Blocking antibodies targeting individual receptors inhibit no more than 50% of the association with ACs, and CD36 Ϫ/Ϫ mice have no obvious defects in phagocytosis or in cross-presentation of antigens encoded within ACs, indicating a redundancy in the system and/or the incomplete characterization of AC receptors on DCs. [17][18][19][20] More recent studies have begun to evaluate the ability of specific AC receptors to modulate DC function. DCs exposed to ACs opsonized with C3bi fragments are inhibited from maturing upon stimulation with LPS or CD40L. This effect is presumably mediated through CR3 and CR4, the receptors for C3bi. 21 Studies of the scavenger receptor CD36 and the ␣v-integrin receptor CD51 have led to similar conclusions regarding their ability to modulate DC function. 12,22 However, it is not known whether such function extends to other members or components of the ␣v-integrin receptor family, particularly the well-established phagocytic receptor ␣v␤5. 17,23 Nor is it clear whether ligation of AC receptors that interfere with DC maturation consequently alters their T-cell-stimulating potential.To gain a better understanding of the phagocytic and immunomodulatory role of the various AC receptors on human DCs, we made use of an AC surrogate system 24 that permitted us to evaluate the function of individual AC receptors. We focused on ␣v␤5 and the CRs, especially CR3. We established that these different AC receptors are not equivalent in function, as might be presumed from published studies, but that there is a distinct division of labor, at least with respect to phagocytosis and tolerance induction potential. While ␣v␤5 mediates efficient phagocytosis, it does not interfere with the DC's capacity to undergo maturation or stimulate T cells. In contrast, engagement of CR3 (or CR4) inhibits the ability of DCs to undergo maturation, produce proinflammatory cytokines or M.Š . conceptualized, designed, and performed experiments; acquired, analyzed, and interpreted data; draft...