Tham khảo tài liệu 'biosensors emerging materials and applications part 11', kỹ thuật - công nghệ, cơ khí - chế tạo máy phục vụ nhu cầu học tập, nghiên cứu và làm việc hiệu quả | 392 Biosensors - Emerging Materials and Applications Fig. 4. n-Rosella in yeast A Wild type cells expressing n-Rosella were imaged using fluorescence microscopy under growing and nitrogen starvation conditions. Accumulation of diffused red fluorescence in the vacuole after 24 h of commencement of nitrogen starvation indicates nucleophagy. B The absence of expression a particular gene product essential for nucleophagy influences nuclear morphology and abrogates correct delivery of n-Rosella to the vacuole. Nuclear blebs remain red and green indicating high pH environment. C The absence of the ATG15 gene abolishes degradation of n-Rosella derived vesicles in the vacuole. Some intravacuolar vesicles are both red and green indicating high pH environment whereas others are only red indicating low pH environment . The schematic right represents an interpretation of the image data. White dashed circles highlight the limits of the vacuole. of additional probes to highlight the location of the lysosome. We next demonstrate in mammalian cells that Rosella can be used to monitor delivery of the cytosol or mitochondrion to the lysosome. HeLa cells maintained in a replete growth medium and transfected with an expression vector encoding c-Rosella Rosella without any additional targeting sequence Fig. 3B when imaged using fluorescence microscopy showed both strong red and green fluorescence distributed throughout the cytosol. Rosella appears to have restricted access to the nuclear compartment less intense staining and is completely excluded from other compartments Fig. 5 . Importantly only 1-2 red and weakly green puncta cell were observed Fig. 5A white arrows suggesting that Rosella has accumulated in a relatively acidic compartment such as a lysosome. These puncta correspond to autophagolysosomes Biosensors for Monitoring Autophagy 393 and represent fusion of an autophagosome carrying the Rosella cargo and a lysosome. Low numbers of puncta observed under growth conditions are
