BCL-2, a novel regulator of apoptosis

The bcl-2 gene has a unique function among mammalian oncogenes as a negative regulator of apoptosis. Its expression pattern in embryonic and adult tissues is consistent with a role in maintaining in vivo survival of specific cell types. The biochemical function of bcl-2 is unknown, but its localization to mitochondrial and microsomal membranes suggests several possibilities, bcl-2 is protective against oxidative stress in mammalian cells and can be replaced by antioxidants in a factor-deprivation model of apoptosis. These results are consistent with a model of apoptotic death involving oxidative stress in a central pathway. The recent discovery of several bcl-2-related genes, some of which also inhibit apoptosis and others that unexpectedly promote apoptosis, has shed new light on several aspects of bcl-2 action. © 1996 Wiley-Liss, Inc.     

Schlafen-3: A novel regulator of intestinal differentiation

Schlafen-3 (Slfn-3), a novel gene, has been shown to be a negative regulator of proliferation. The current investigation was undertaken to determine whether Slfn-3 might play a role in regulating cellular differentiation. Butyric acid, a short chain fatty acid, which induced differentiation of intestinal cells as evidenced by increased alkaline phosphatase (ALP) activity in the rat small intestinal IEC-6 cells, also produced a marked increase in Slfn-3 expression. Furthermore, overexpression of Slfn-3 caused stimulation of ALP activity in IEC-6 cells, which was exacerbated by butyrate. On the other hand, downregulation of Slfn-3 by slfn-3-si-RNA greatly attenuated the butyrate-mediated induction of differentiation of IEC-6 cells. Additionally, we observed that increased expression of Slfn-3 in colon cancer HCT-116 cells stimulated TGF-β expression and modulated expression of its downstream effectors as evidenced by increased expression of p27kip1 and downregulation of CDK-2. In addition, Slfn-3 increases E-cadherin expression but downregulates β-catenin. In conclusion, our data show that Slfn-3 plays a critical role in regulating intestinal mucosal differentiation. Furthermore our data also show that TGF-β signaling pathway plays an important role in mediating slfn-3 induced differentiation.     

Connexin 43 a potential regulator of cell proliferation and apoptosis within the seminiferous epithelium

The gap junction proteins, connexins (Cx), are present in the testis and among them Cx43 play an essential role in spermatogenesis. By using an in vitro proliferation model of germ cells and Sertoli cells, we tempted here to clarify the role of Cx43 in the control of Sertoli and germ cell proliferation and apoptosis. Cx43 was detected in purified preparations of Sertoli cells and spermatogonia and immunolocalized in both cell types identified by vimentin and c-kit, respectively. Inhibition of gap junction coupling by the gap junction inhibitor α-GA significantly enhanced BrdU incorporation in Sertoli cells and reduced the number of activated caspase-3 positive germ cells. Similarly, inhibitory Cx43 and pan-Cx mimetic inhibitory peptides increased proliferation of Sertoli cells and stimulated survival of germ cells. Cx32 mimetic inhibitory peptide also stimulated Sertoli cell proliferation without altering germ cell proliferation and apoptosis. The present results reveal that Cx43 gap junctions between Sertoli cells participate in the control of Sertoli cell proliferation and that Cx43 gap junctions between Sertoli cells and spermatogonia are indirectly involved in germ cell number increase by controlling germ cell survival rather than germ cell proliferation.     

Cell cycle and apoptosis regulatory protein-1: a novel regulator of apoptosis in the colonic mucosa during aging

Although the regulatory mechanisms for the age-related rise in proliferation and reduction in apoptosis in the colonic mucosa are yet to be fully delineated, we have demonstrated that these events are associated with increased expression and activation of epithelial growth factor receptor (EGFR)/ErbB-1 and some of its receptor family members (EGFRs), indicating their involvement in these processes. However, the downstream signaling events of EGFR and/or its family members regulating age-related changes in mucosal proliferation and apoptosis remain to be delineated. Cell cycle and apoptosis regulatory protein-1 (CARP-1), a novel growth signaling regulator that we isolated, participates in EGFR-dependent signaling. In the current investigation, we examined the involvement of CARP-1 in colonic mucosal growth-related processes during aging. We report that the age-related reduction in apoptosis in the colonic mucosa is associated with increased expression and tyrosine phosphorylation of not only EGFR but also ErbB-2 and ErbB-3. In contrast, protein and mRNA levels of CARP-1 as well as tyrosine phosphorylation of CARP-1 are decreased. Additionally, we have observed that administration of wortmannin, an inhibitor of phosphatidylinositol 3-kinase activity that accelerates apoptosis in the colonic mucosa of aged rats, causes a marked increase in expression and tyrosine phosphorylation of CARP-1. The age-related decline in CARP-1 expression could partly be attributed to increased methylation of the CARP-1 promoter. Taken together, our data suggest that not only EGFR but also its other members are involved in regulating colonic mucosal growth during aging and that CARP-1 may play a crucial role in transducing EGFRs signals.     

Pyridoxamine protects intestinal epithelium from ionizing radiation-induced apoptosis

Reactive oxygen species (ROS) and reactive carbonyl species (RCS) are the major causes of biological tissue damage during exposure to ionizing radiation (IR). The existing strategies to protect normal tissues from the detrimental effects of IR suffer from several shortcomings including highly toxic side effects, unfavorable administration routes, and low efficacy. These shortcomings emphasize a need for radioprotective treatments that combine effectiveness with safety and ease of use. In this paper, we demonstrate that pyridoxamine, a ROS and RCS scavenger with a very favorable safety profile, can inhibit IR-induced gastrointestinal epithelial apoptosis in cell culture and in an animal model. Pyridoxamine was more effective at protecting from radiation-induced apoptosis than amifostine, a synthetic thiol compound and the only FDA-approved radioprotector. We suggest that pyridoxamine has potential as an effective and safe radioprotective agent.     

Characterization of a novel negative regulator (DOC-2/DAB2) of c-Src in normal prostatic epithelium and cancer

DOC-2/DAB2 is a potent tumor suppressor in many cancer types including prostate cancer. In prostate cancer, expression of DOC-2/DAB2 can inhibit its growth. Our recent studies demonstrate that DOC-2/DAB2 can suppress both protein kinase C and peptide growth factor-elicited signal pathways via the Ras-mitogen-activated protein kinase pathway. In this study, we further showed that the proline-rich domain of DOC-2/DAB2 could also interact with proteins containing the Src homology 3 domain, such as Src and Fgr. The binding of c-Src to DOC-2/DAB2 was enhanced in cells treated with growth factor, and this interaction resulted in c-Src inactivation. The c-Src inactivation was evidenced by the decreased tyrosine 416 phosphorylation of c-Src and reduced downstream effector activation. It appears that DOC-2/DAB2 can bind to Src homology 3 domain of c-Src and maintain it in an inactive conformation. Thus, this study provides a new mechanism for modulating c-Src in prostatic epithelium and cancer.     

O-GlcNAc: a novel regulator of immunometabolism

The rapidly expanding field of immunometabolism focuses on how metabolism controls the function of immune cells. CD4⁺ T cells are essential for the adaptive immune response leading to the eradication of specific pathogens. However, when T cells are inappropriately over-active, they can drive autoimmunity, allergic disease, and chronic inflammation. The mechanisms by which metabolic changes influence function in CD4⁺ T cells are not fully understood. The post-translational protein modification, O-GlcNAc (O-linked β-N-acetylglucosamine), dynamically cycles on and off of intracellular proteins as cells respond to their environment and flux through metabolic pathways changes. As the rate of O-GlcNAc cycling fluctuates, protein function, stability, and/or localization can be affected. Thus, O-GlcNAc is critically poised at the nexus of cellular metabolism and function. This review highlights the intra- and extracellular metabolic factors that influence CD4⁺ T cell activation and differentiation and how O-GlcNAc regulates these processes. We also propose areas of future research that may illuminate O-GlcNAc’s role in the plasticity and pathogenicity of CD4⁺ T cells and uncover new potential therapeutic targets.     

ARMER,apoptotic regulator in the membrane of the endoplasmic reticulum,a novel inhibitor of apoptosis

We have identified a novel protein, apoptotic regulator in the membrane of the endoplasmic reticulum (ARMER), which protects HT1080 fibrosarcoma cells from apoptosis induced by various stimuli. We demonstrate that ARMER is an endoplasmic reticulum (ER) integral membrane protein with four predicted transmembrane domains and a COOH-terminal KKXX ER retrieval motif. We used an inducible expression system (pIND) to study the effects of regulated ARMER overexpression. Cells in which ARMER was overexpressed exhibited protection from multiple apoptotic inducers including serum starvation, doxorubicin, UV irradiation, tumor necrosis factor alpha, and the ER stressors brefeldin A, tunicamycin, and thapsigargin. Analysis of the caspase proteolytic cascade reveals that ARMER inhibits proteolysis of the caspase-9-specific fluorogenic substrate LEHD-AFC as well as endogenous substrates downstream of caspase-9; however, it does not inhibit cytochrome c release or cleavage of caspase-9 itself. Apoptotic stimuli cause endogenous levels of ARMER protein and RNA to decrease, leading to cell death; however, sustaining ARMER protein levels through exogenous expression inhibits apoptosis. These data suggest that ARMER is a novel ER integral membrane protein which protects cells by inhibiting caspase-9 activity and reveal a possible role for ARMER in cell survival.     

微RNA:天然免疫的新型调节子

微RNA（microRNA,miRNA）是多种生物学过程的有效调节子,并表现为基因的定量调节。新出现的证据表明miRNA与天然免疫反应的调节有关。这种调节作用有助于维持宿主免疫反应和保护感染组织间的平衡。深入理解miRNA对天然免疫反应的调节有助于鉴定免疫调节的新靶标和建立基于miRNA的有效疗法。本综述重点总结miRNA在调节免疫细胞发育、Toll样受体和炎症细胞因子信号中的作用。     

Nir2, a novel regulator of cell morphogenesis

Cell morphogenesis requires dynamic reorganization of the actin cytoskeleton, a process that is tightly regulated by the Rho family of small GTPases. These GTPases act as molecular switches by shuttling between their inactive GDP-bound and active GTP-bound forms. Here we show that Nir2, a novel protein related to Drosophila retinal degeneration B (RdgB), markedly affects cell morphology through a novel Rho-inhibitory domain (Rid) which resides in its N-terminal region. Rid exhibits sequence homology with the Rho-binding site of formin-homology (FH) proteins and leads to an apparent loss of F-actin staining when ectopically expressed in mammalian cells. We also show that Rid inhibits Rho-mediated stress fiber formation and lysophosphatidic acid-induced RhoA activation. Biochemical studies demonstrated that Nir2, via Rid, preferentially binds to the inactive GDP-bound form of the small GTPase Rho. Microinjection of antibodies against Nir2 into neuronal cells markedly attenuates neurite extension, whereas overexpression of Nir2 in these cells attenuates Rho-mediated neurite retraction. These results implicate Nir2 as a novel regulator of the small GTPase Rho in actin cytoskeleton reorganization and cell morphogenesis.     

Speedy: a novel cell cycle regulator of the G2/M transition

Stage VI Xenopus oocytes are suspended at the G2/M transition of meiosis I, and represent an excellent system for the identification and examination of cell cycle regulatory proteins. Essential cell cycle regulators such as MAPK, cyclins and mos have the ability to induce oocyte maturation, causing the resumption of the cell cycle from its arrested state. We have identified the product of a novel Xenopus gene, Speedy or Spy1, which is able to induce rapid maturation of Xenopus oocytes, resulting in the induction of germinal vesicle breakdown (GVBD) and activation of M-phasepromoting factor (MPF). Spy1 activates the MAPK pathway in oocytes, and its ability to induce maturation is dependent upon this pathway. Spy1-induced maturation occurs much more rapidly than maturation induced by other cell cycle regulators including progesterone, mos or Ras, and does not require any of these proteins or hormones, indicating that Spy1-induced maturation proceeds through a novel regulatory pathway. In addition, we have shown that Spy1 physically interacts with cdk2, and prematurely activates cdk2 kinase activity. Spy1 therefore represents a novel cell cycle regulatory protein, inducing maturation through the activation of MAPK and MPF, and also leading to the premature activation of cdk2.     

Oxygen stress: a regulator of apoptosis in yeast.

Oxygen radicals are important components of metazoan apoptosis. We have found that apoptosis can be induced in the yeast Saccharomyces cerevisiae by depletion of glutathione or by low external doses of H2O2. Cycloheximide prevents apoptotic death revealing active participation of the cell. Yeast can also be triggered into apoptosis by a mutation in CDC48 or by expression of mammalian bax. In both cases, we show oxygen radicals to accumulate in the cell, whereas radical depletion or hypoxia prevents apoptosis. These results suggest that the generation of oxygen radicals is a key event in the ancestral apoptotic pathway and offer an explanation for the mechanism of bax-induced apoptosis in the absence of any established apoptotic gene in yeast.