Erythroid cells undergo enucleation and the removal of organelles during terminal differentiation 1-3 . Although autophagy has been suggested to mediate the elimination of organelles for erythroid maturation 2-6 , the molecular mechanisms underlying this process remain undefined. Here we report a role for a Bcl-2 family member, Nix (also called Bnip3L) 7-9 , in the regulation of erythroid maturation through mitochondrial autophagy. Nix −/− mice developed anaemia with reduced mature erythrocytes and compensatory expansion of erythroid precursors. Erythrocytes in the peripheral blood of Nix −/− mice exhibited mitochondrial retention and reduced lifespan in vivo. Although the clearance of ribosomes proceeded normally in the absence of Nix, the entry of mitochondria into autophagosomes for clearance was defective. Deficiency in Nix inhibited the loss of mitochondrial membrane potential (ΔΨ m ), and treatment with uncoupling chemicals or a BH3 mimetic induced the loss of ΔΨ m and restored the sequestration of mitochondria into autophagosomes in Nix −/− erythroid cells. These results suggest that Nix-dependent loss of ΔΨ m is important for targeting the mitochondria into autophagosomes for clearance during erythroid maturation, and interference with this function impairs erythroid maturation and results in anaemia. Our study may also provide insights into molecular mechanisms underlying mitochondrial quality control involving mitochondrial autophagy.Nix, a BH3-only member of the Bcl-2 family, is upregulated in erythroid cells undergoing terminal differentiation 10 . To determine the potential function for Nix in erythroid maturation, we generated Nix −/− mice using embryonic stem (ES) cells with a gene trap insertion between exons 3 and 4 of Nix ( Supplementary Fig. 2). We first examined red blood cells in the peripheral blood (RBCs), including reticulocytes and erythrocytes, in Nix −/− mice. Although RBC counts were decreased (Supplementary Table 1), polychromasia and increased reticulocytes were observed in Nix −/− mice ( Fig. 1a and Supplementary Fig. 3a). We also examined RBCs for the expression of an erythroid cell marker, glycophorin-A-associated Ter119, and for transferrin receptor CD71, which is downregulated during terminal erythroid differentiation 11,12 . Although Ter119 low CD71 high and Ter119 + CD71 high early erythroblasts 13 were absent in the peripheral blood, a significant increase in Ter119 + CD71 + reticulocytes was observed in Nix −/− mice (Fig. 1b). Electron microscopy also showed more irregularly shaped cellsCorrespondence and requests for materials should be addressed to M.C. (minc@bcm.tmc.edu) or J.W. (jinwang@bcm.tmc.edu). Author Contributions H.S. conducted the majority of the experiments, supervised by J.W. and M.C.; P.T. stained spleen sections and blood smears; S.K.D. measured osmotic fragility and assisted with biotin and CMFDA labelling; A.S. performed RT-PCR for Epo; J.T.P. and P.T. provided experimental advice; M.C. and J.W. generated the Nix −/− mice, designed experiments and...

Apoptosis of mature T lymphocytes preserves peripheral homeostasis and tolerance by countering the profound changes in the number and types of T cells stimulated by diverse antigens. T cell apoptosis occurs in at least two major forms: antigen-driven and lymphokine withdrawal. These forms of death are controlled in response to local levels of IL-2 and antigen in a feedback mechanism termed propriocidal regulation. Active antigen-driven death is mediated by the expression of death cytokines such as FasL and TNF. These death cytokines engage specific receptors that assemble caspase-activating protein complexes. These signaling complexes tightly regulate cell death but are vulnerable to inherited defects. Passive lymphokine withdrawal death may result from the cytoplasmic activation of caspases that is regulated by mitochondria and the Bcl-2 protein. The human disease, Autoimmune Lymphoproliferative Syndrome (ALPS) is due to dominant-interfering mutations in the Fas/APO-1/CD95 receptor and other components of the death pathway. The study of ALPS patients reveals the necessity of apoptosis for preventing autoimmunity and allows the genetic investigation of apoptosis in humans. Immunological, cellular, and molecular evidence indicates that throughout the life of a T cell, apoptosis may be evoked in excessive, harmful, or useless clonotypes to preserve a healthy and balanced immune system.

Apoptosis is a form of programmed cell death that is controlled by aspartate-specific cysteine proteases called caspases. In the immune system, apoptosis counters the proliferation of lymphocytes to achieve a homeostatic balance, which allows potent responses to pathogens but avoids autoimmunity. The CD95 (Fas, Apo-1) receptor triggers lymphocyte apoptosis by recruiting Fas-associated death domain (FADD), caspase-8 and caspase-10 proteins into a death-inducing signalling complex. Heterozygous mutations in CD95, CD95 ligand or caspase-10 underlie most cases of autoimmune lymphoproliferative syndrome (ALPS), a human disorder that is characterized by defective lymphocyte apoptosis, lymphadenopathy, splenomegaly and autoimmunity. Mutations in caspase-8 have not been described in ALPS, and homozygous caspase-8 deficiency causes embryonic lethality in mice. Here we describe a human kindred with an inherited genetic deficiency of caspase-8. Homozygous individuals manifest defective lymphocyte apoptosis and homeostasis but, unlike individuals affected with ALPS, also have defects in their activation of T lymphocytes, B lymphocytes and natural killer cells, which leads to immunodeficiency. Thus, caspase-8 deficiency in humans is compatible with normal development and shows that caspase-8 has a postnatal role in immune activation of naive lymphocytes.