PMID- 22125069
OWN - NLM
STAT- MEDLINE
DCOM- 20120316
LR  - 20211203
IS  - 1940-6029 (Electronic)
IS  - 1064-3745 (Linking)
VI  - 821
DP  - 2012
TI  - Quantitative visualization of autophagy induction by mTOR inhibitors.
PG  - 239-50
LID - 10.1007/978-1-61779-430-8_14 [doi]
AB  - Autophagy is a catabolic pathway that degrades bulk cytosol in lysosomal 
      compartments enabling amino acids and fatty acids to be recycled. One of the key 
      regulators of autophagy is the mammalian target of rapamycin (mTOR), a conserved 
      serine/threonine kinase which suppresses the initiation of the autophagic process 
      when nutrients, growth factors, and energy are available. Inhibition of mTOR, 
      e.g., by small molecules such as rapamycin, results in activation of autophagy. 
      To quantify autophagy induction by mTOR inhibitors, we use an mCherry-GFP-LC3 
      reporter which is amenable to retroviral delivery into mammalian cells, stable 
      expression, and analysis by fluorescence microscopy. Here, we describe our 
      imaging protocol and image recognition algorithm to visualize and measure changes 
      in the autophagic pathway.
FAU - Nyfeler, Beat
AU  - Nyfeler B
AD  - Developmental and Molecular Pathways, Novartis Institutes for BioMedical 
      Research, Cambridge, MA, USA.
FAU - Bergman, Philip
AU  - Bergman P
FAU - Wilson, Christopher J
AU  - Wilson CJ
FAU - Murphy, Leon O
AU  - Murphy LO
LA  - eng
PT  - Journal Article
PL  - United States
TA  - Methods Mol Biol
JT  - Methods in molecular biology (Clifton, N.J.)
JID - 9214969
RN  - 0 (MAP1LC3A protein, human)
RN  - 0 (Microtubule-Associated Proteins)
RN  - 0 (Morpholines)
RN  - 0 (Pyrimidines)
RN  - 147336-22-9 (Green Fluorescent Proteins)
RN  - 81HJG228AB (Ku 0063794)
RN  - EC 2.7.11.1 (TOR Serine-Threonine Kinases)
RN  - W36ZG6FT64 (Sirolimus)
SB  - IM
MH  - Amino Acid Sequence
MH  - Autophagy/*drug effects
MH  - Genetic Vectors
MH  - Green Fluorescent Proteins/analysis
MH  - Humans
MH  - Microscopy, Fluorescence/*methods
MH  - Microtubule-Associated Proteins/analysis
MH  - Molecular Sequence Data
MH  - Morpholines/pharmacology
MH  - Pyrimidines/pharmacology
MH  - Sirolimus/pharmacology
MH  - TOR Serine-Threonine Kinases/*antagonists & inhibitors
EDAT- 2011/11/30 06:00
MHDA- 2012/03/17 06:00
CRDT- 2011/11/30 06:00
PHST- 2011/11/30 06:00 [entrez]
PHST- 2011/11/30 06:00 [pubmed]
PHST- 2012/03/17 06:00 [medline]
AID - 10.1007/978-1-61779-430-8_14 [doi]
PST - ppublish
SO  - Methods Mol Biol. 2012;821:239-50. doi: 10.1007/978-1-61779-430-8_14.