Linked Life Data

21730954

Source:http://linkedlifedata.com/resource/pubmed/id/21730954

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
52
pubmed:dateCreated
2011-7-6
pubmed:abstractText
Apoptosis, or programmed cell death, is a conserved and highly regulated pathway by which cells die¹. Apoptosis can be triggered when cells encounter a wide range of cytotoxic stresses. These insults initiate signaling cascades that ultimately cause the release of cytochrome c from the mitochondrial intermembrane space to the cytoplasm². The release of cytochrome c from mitochondria is a key event that triggers the rapid activation of caspases, the key cellular proteases which ultimately execute cell death³??. The pathway of apoptosis is regulated at points upstream and downstream of cytochrome c release from mitochondria?. In order to study the post-mitochondrial regulation of caspase activation, many investigators have turned to direct cytoplasmic microinjection of holocytochrome c (heme-attached) protein into cells???. Cytochrome c is normally localized to the mitochondria where attachment of a heme group is necessary to enable it to activate apoptosis¹??¹¹. Therefore, to directly activate caspases, it is necessary to inject the holocytochrome c protein instead of its cDNA, because while the expression of cytochrome c from cDNA constructs will result in mitochondrial targeting and heme attachment, it will be sequestered from cytosolic caspases. Thus, the direct cytosolic microinjection of purified heme-attached cytochrome c protein is a useful tool to mimic mitochondrial cytochrome c release and apoptosis without the use of toxic insults which cause cellular and mitochondrial damage. In this article, we describe a method for the microinjection of cytochrome c protein into cells, using mouse embryonic fibroblasts (MEFs) and primary sympathetic neurons as examples. While this protocol focuses on the injection of cytochrome c for investigations of apoptosis, the techniques shown here can also be easily adapted for microinjection of other proteins of interest.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
1940-087X
pubmed:author
pubmed:issnType
Electronic
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:dateRevised
2011-11-17
pubmed:meshHeading
pubmed:year
2011
pubmed:articleTitle
Activation of apoptosis by cytoplasmic microinjection of cytochrome c.
pubmed:affiliation
Department of Cell and Developmental Biology, Neuroscience Center, University of North Carolina.
pubmed:publicationType
Journal Article, Research Support, N.I.H., Extramural, Video-Audio Media