We describe the lethal, recessive accelerated-cell-death11 Arabidopsis mutant (acd11). Cell death in acd11 exhibits characteristics of animal apoptosis monitored by flow cytometry, and acd11 constitutively expresses defense-related genes that accompany the hypersensitive response normally triggered by avirulent pathogens. Global transcriptional changes during programmed cell death (PCD) and defense activation in acd11 were monitored by cDNA microarray hybridization. The PCD and defense pathways activated in acd11 are salicylic acid (SA) dependent, but do not require intact jasmonic acid or ethylene signaling pathways. Light is required for PCD execution in acd11, as application of an SA-analog to SA-deficient acd11 induced death in the light, but not in the dark. Epistatic analysis showed that the SA-dependent pathways require two regulators of SA-mediated resistance responses, PAD4 and EDS1. Furthermore, acd11 PR1 gene expression, but not cell death, depends on the SA signal tranducer NPR1, suggesting that the npr1-1 mutation uncouples resistance responses and cell death in acd11. The acd11 phenotype is caused by deletion of the ACD11 gene encoding a protein homologous to a mammalian glycolipid transfer protein (GLTP). In contrast to GLTP, ACD11 accelerates the transfer of sphingosine, but not of glycosphingolipids, between membranes in vitro.
The ocular lens is the only organ that does not develop spontaneous tumor. The molecular mechanism for this phenomenon remains unknown. Through examination of the signaling pathways mediating stress-induced apoptosis, here we presented evidence to show that different from most other tissues in which the extracellular signal-regulated kinases (ERKs) pathway is generally implicated in mediation of survival signals activated by different factors, the RAF/MEK/ERK signaling pathway alone plays a key role in stress-activated apoptosis of lens epithelial cells. Treatment of N/N1003A cells with calcimycin, a calcium mobilizer, activates the RAF/MEK/ERK pathway through RAS, which is indispensable for the induced apoptosis because inhibition of this pathway by either pharmacological drug or dominant negative mutants greatly attenuates the induced apoptosis. Calcimycin also activates p38 kinase and JNK2, which are not involved in calcium-induced apoptosis. Downstream of ERK activation, p53 is essential. Activation of RAF/MEK/ERK pathway by calcimycin leads to distinct up-regulation of p53. Moreover, overexpression of p53 enhances calcimycin-induced apoptosis, whereas inhibition of p53 expression attenuates calcimycin-induced apoptosis. Up-regulation of p53 directly promotes Bax expression, which changes the integrity of mitochondria, leading to release of cytochrome c, activation of caspase-3 and eventually execution of apoptosis. Overexpression of alphaB-crystallin, a member of the small heat-shock protein family, blocks activation of RAS to inhibit ERK1/2 activation, and greatly attenuates calcimycin-induced apoptosis. Together, our results provide 1) a partial explanation for the lack of spontaneous tumor in the lens, 2) a novel signaling pathway for calcium-induced apoptosis, and 3) a novel antiapoptotic mechanism for alphaB-crystallin.
HET-C2 is a fungal protein that transfers glycosphingolipids between membranes and has limited sequence homology with human glycolipid transfer protein (GLTP). The human GLTP fold is unique among lipid binding/transfer proteins, defining the GLTP superfamily. Herein, GLTP fold formation by HET-C2, its glycolipid transfer specificity, and the functional role(s) of its two Trp residues have been investigated. X-ray diffraction (1.9 Å ) revealed a GLTP fold with all key sugar headgroup recognition residues (Asp 66 , Asn 70 , Lys 73 , Trp 109 , and His 147 ) conserved and properly oriented for glycolipid binding. Far-UV CD showed secondary structure dominated by ␣-helices and a cooperative thermal unfolding transition of 49°C, features consistent with a GLTP fold. Environmentally induced optical activity of Trp/Tyr/Phe (2:4:12) detected by near-UV CD was unaffected by membranes containing glycolipid but was slightly altered by membranes lacking glycolipid. Trp fluorescence was maximal at ϳ355 nm and accessible to aqueous quenchers, indicating free exposure to the aqueous milieu and consistent with surface localization of the two Trps. Interaction with membranes lacking glycolipid triggered significant decreases in Trp emission intensity but lesser than decreases induced by membranes containing glycolipid. Binding of glycolipid (confirmed by electrospray injection mass spectrometry) resulted in a blue-shifted emission wavelength maximum (ϳ6 nm) permitting determination of binding affinities. The unique positioning of Trp 208 at the HET-C2 C terminus revealed membrane-induced conformational changes that precede glycolipid uptake, whereas key differences in residues of the sugar headgroup recognition center accounted for altered glycolipid specificity and suggested evolutionary adaptation for the simpler glycosphingolipid compositions of filamentous fungi.Self/nonself recognition is a universally important process, encompassing intercellular interactions ranging from vertebrate immune responses to somatic chimera formation in protists, filamentous fungi, sponges, ascidians, and tunicates. Self/ nonself discrimination becomes critical for filamentous fungi during hyphal fusion, which enables the exchange of cytoplasm and nuclei during the assimilative growth phase (1-4). Nonself recognition triggers a postfusion, programmed cell death process known as vegetative incompatibility, which leads to heterokaryon death. The benefits of vegetative incompatibility include prevention of transmission of deleterious cytoplasmic elements (i.e. viruses) as well as restriction of plundering by parasitic genotypes.het genes are known to play a major role in vegetative incompatibility processes that occur in Neurospora crassa and Podospora anserina. het genes exhibit extensive polymorphism and generally encode proteins carrying a HET domain (1-4). Originally, this domain was linked to the het-c2 gene of P. anserina, where it was shown to encode a protein similar in size and with limited sequence homology to mammalian glycolipid transfer...
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