The lysosome-associated apoptosis-inducing protein containing the pleckstrin homology (PH) and FYVE domains (LAPF), representative of a novel family of PH and FYVE domain-containing proteins, induces caspase-independent apoptosis via the lysosomal-mitochondrial pathway.
Lysosomes have recently been identified as important apoptotic signal integrators in response to various stimuli. Here we report the functional characterization of LAPF, a novel lysosome-associated apoptosis-inducing protein containing PH and FYVE domains. LAPF is a representative of a new protein family, the Phafins (protein containing both PH and FYVE domains), which consists of 14 unidentified proteins from various species. Overexpression of LAPF in L929 cells induces apoptosis and also increases cell sensitivity to TNFalpha-induced apoptosis, concomitant with its translocation to lysosomes. Two mutants of LAPF, either lacking the PH or FYVE domain, failed to induce cell death and translocate to lysosomes, suggesting that both domains are required for its apoptosis-inducing activity and relocation. We demonstrate that LAPF may induce apoptosis via the following steps: LAPF translocation to lysosomes, lysosomal membrane permeabilization (LMP), release of cathepsin (cath) D and L, mitochondrial membrane permeabilization (MMP), release of apoptosis-inducing factor (AIF), and caspase-independent apoptosis. The cath D-specific inhibitor attenuates LAPF-induced apoptosis, indicating a pivotal role of lysosomes in LAPF-initiated apoptosis. We also demonstrate that the lysosomal pathway was employed in the typical apoptotic model in which high dose TNFalpha was used to stimulate L929 cells. Silencing of LAPF expression by small RNA interference protected L929 cells from hTNFalpha-induced apoptosis by impairing hTNFalpha-triggered LMP and MMP. Therefore, LAPF may launch caspase-independent apoptosis through the lysosomal-mitochondrial pathway.
Chen
Wei
W
Institute of Immunology, Zhejiang University, 353 Yanan Road, Hangzhou 310031, Zhejiang, China.
Li
Nan
N
Chen
Taoyong
T
Han
Yanmei
Y
Li
Changfei
C
Wang
Yuzhen
Y
He
Weigang
W
Zhang
Lihuang
L
Wan
Tao
T
Cao
Xuetao
X
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D015152:Blotting, Northern; D020169:Caspases / Q000378:metabolism*; D002402:Cathepsin D / Q000378:metabolism; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D005354:Fibrosarcoma; D015870:Gene Expression; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D051379:Mice; D008854:Microscopy, Electron; D008928:Mitochondria / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D051017:Apoptosis Regulatory Proteins; D018076:DNA, Complementary; C508358:PLEKHF1 protein, human; D011506:Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D002403:Cathepsins; D020169:Caspases; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L; C534278:Ctsl protein, mouse; D002402:Cathepsin D