Biochemical and Biophysical Research Communications
Organelle degradation during the lens and erythroid differentiation is independent of autophagy
Section snippets
Materials and methods
Mice. Atg5+/− and Atg5−/− mice on the C57BL/6 background have been described previously [7]. Atg5+/− mice were crossed with green fluorescent protein (GFP)-fused LC3 transgenic mice [6] to produce Atg5−/− mice expressing GFP-LC3 (GFP-LC3 Atg5−/−). For caesarian delivery, pregnant mothers were injected on 17.5 and 18.5 dpc with 2 mg progesterone (Luteum Injection, Teikoku Hormone, Tokyo) to delay birth; neonates were obtained at 19.5 dpc. To prolong the survival of Atg5−/− neonates, artificial milk
Results and discussion
Since the organelle free zone (OFZ) in the lens is created between 17.5 dpc and birth, we first determined the occurrence of autophagy at this stage. LC3, a mammalian homolog of yeast Atg8, localizes to autophagosome membranes as a phosphatidylethanolamine (PE)-conjugated form [27], [28]. Taking this advantage, we have developed an autophagy-indicator mouse model, in which GFP-LC3 is systemically expressed [6]. Using these mice, autophagosomes can be detected as GFP-positive dots [29]. In lens
Acknowledgments
This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan. The authors thank the Yamada Science Foundation and the Cell Science Research Foundation for their financial support. This work was performed under the NIBB Cooperative Research Program. The authors declare that they have no competing financial interests.
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