TY - JOUR
T1 - Detection of RTK pathway activation in Drosophila using anti-dpERK immunofluorescence staining.
AU - Helman, Aharon
AU - Paroush, Ze'ev
PY - 2010
Y1 - 2010
N2 - In Drosophila, like in other metazoans, receptor tyrosine kinase (RTK) signaling pathways control diverse cellular processes such as migration, growth, fate determination, and differentiation (Shilo, Development 132:4017-4027, 2005). Activation of RTKs by their extracellular ligands triggers a signal transduction cascade, mediated by the Ras/Raf/MEK cassette, which ultimately leads to dual phosphorylation and activation of the mitogen-activated protein kinase/extracellularly regulated kinase (MAPK/Erk). Once active, MAPK/Erk phosphorylates its cytoplasmic and nuclear substrates, consequently modulating (i.e., stimulating or inhibiting) their biological function (Murphy and Blenis, Trends in Biochemical Sciences 31:268-275, 2006). The currently available antibody specific for the doubly phosphorylated form of MAPK/Erk (dpERK) (Yung et al., FEBS Letters 408:292-296, 1997) provides a valuable readout for RTK signaling: it enables the spatiotemporal detection of RTK pathway activity in the developing organism, in situ (Gabay et al., Development 124:3535-3541, 1997; Gabay et al., Science 277:1103-1106, 1997). Here, we present a detailed protocol for anti-dpERK immunofluorescent staining that can be applied to the analysis of MAPK/Erk signaling in Drosophila embryogenesis.
AB - In Drosophila, like in other metazoans, receptor tyrosine kinase (RTK) signaling pathways control diverse cellular processes such as migration, growth, fate determination, and differentiation (Shilo, Development 132:4017-4027, 2005). Activation of RTKs by their extracellular ligands triggers a signal transduction cascade, mediated by the Ras/Raf/MEK cassette, which ultimately leads to dual phosphorylation and activation of the mitogen-activated protein kinase/extracellularly regulated kinase (MAPK/Erk). Once active, MAPK/Erk phosphorylates its cytoplasmic and nuclear substrates, consequently modulating (i.e., stimulating or inhibiting) their biological function (Murphy and Blenis, Trends in Biochemical Sciences 31:268-275, 2006). The currently available antibody specific for the doubly phosphorylated form of MAPK/Erk (dpERK) (Yung et al., FEBS Letters 408:292-296, 1997) provides a valuable readout for RTK signaling: it enables the spatiotemporal detection of RTK pathway activity in the developing organism, in situ (Gabay et al., Development 124:3535-3541, 1997; Gabay et al., Science 277:1103-1106, 1997). Here, we present a detailed protocol for anti-dpERK immunofluorescent staining that can be applied to the analysis of MAPK/Erk signaling in Drosophila embryogenesis.
UR - http://www.scopus.com/inward/record.url?scp=79952111781&partnerID=8YFLogxK
U2 - 10.1007/978-1-60761-795-2_24
DO - 10.1007/978-1-60761-795-2_24
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C2 - 20811997
AN - SCOPUS:79952111781
SN - 1064-3745
VL - 661
SP - 401
EP - 408
JO - Methods in Molecular Biology
JF - Methods in Molecular Biology
ER -