Programmed cell death can be divided into apoptosis and autophagic cell death. We describe the biological activities of TMEM166 (transmembrane protein 166, also known as FLJ13391), which is a novel lysosome and endoplasmic reticulum-associated membrane protein containing a putative TM domain. Overexpression of TMEM166 markedly inhibited colony formation in HeLa cells. Simultaneously, typical morphological characteristics consistent with autophagy were observed by transmission electron microscopy, including extensive autophagic vacuolization and enclosure of cell organelles by double-membrane structures. Further experiments confirmed that the overexpression of TMEM166 increased the punctate distribution of MDC staining and GFP-LC3 in HeLa cells, as well as the LC3-II/LC3-I proportion. On the other hand, TMEM166-transfected HeLa and 293T cells succumbed to cell death with hallmarks of apoptosis including phosphatidylserine externalization, loss of mitochondrial transmembrane potential, caspase activation and chromatin condensation. Kinetic analysis revealed that the appearance of autophagy-related biochemical parameters preceded the nuclear changes typical of apoptosis in TMEM166-transfected HeLa cells. Suppression of TMEM166 expression by small interference RNA inhibited starvation-induced autophagy in HeLa cells. These findings show for the first time that TMEM166 is a novel regulator involved in both autophagy and apoptosis.

Apoptosis is a genetically determined cell suicide program. Mitochondria play a central role in this process and various molecules have been shown to regulate apoptosis in this organelle. In the present study, we firstly identified that protein tyrosine phosphatase interacting protein 51 (PTPIP51) is a novel mitochondrial protein, which may induce apoptosis in HEK293T and HeLa cell lines. PTPIP51 transfection resulted in the externalization of phosphatidylserine (PS), activation of caspase-3, cleavage of PARP, and condensation of nuclear DNA. Further investigation revealed that PTPIP51 over-expression caused a decrease in mitochondrial membrane potential and release of cytochrome c, suggesting that it may be involved in a mitochondria/cytochrome c mediated apoptosis pathway. We also found that a putative TM domain near the N terminus of PTPIP51 is required for its targeting to mitochondria, as evidenced by the finding that deletion of the PTPIP51 TM domain prevented the protein's mitochondiral localization. Furthermore, this deletion significantly influenced the ability of PTPIP51 to induce apoptosis. Taken together, the results of the present study suggest that PTPIP51 is a mitochondrial protein with apoptosis-inducing function and that the N-terminal TM domain is required for both the correct targeting of the protein to mitochondria and its apoptotic functions.

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CD133 is a pentaspan transmembrane protein that can serve as a biomarker for cancer stem cells, although its biochemical mechanism remains unclear. Here we report that CD133 expression enhances glioma cell tolerance of a nutrient-deprived microenvironment. Under starvation conditions, CD133-positive cells exhibited higher survival and decreased levels of apoptosis. These changes were dependent on activation of autophagy-associated gene signaling and were impaired by the autophagic inhibitor chloroquine. Furthermore, rapamycin up-regulated the level of autophagy and inversely reduced CD133 expression. Immunofluorescence confirmed that starvation promoted release of CD133 from the plasma membrane to the cytoplasm, with CD133 also partially co-localizing with LC3 upon starvation. Additionally, CD133 partially co-localized with Beclin1, Atg5, and lysosomes, indicating that CD133 directly participates in the autophagosome membrane fusion process and ultimately undergoes lysosomal degradation. Collectively, our results demonstrate that CD133 contributes to cell survival by regulating autophagy, and that targeting CD133-linked signaling and autophagy may be useful in improving anti-cancer treatments.