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Full Abstracts – TECHNIQUES AND FUNCTIONAL GENOMICS
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Functional redundancy of the
Drosophila
p38 MAP kinases probed by mass spectrometry-based interaction proteomics.
Vladimir Belozerov
1,2
, Zhen-
Yuan Lin
3
, Anne-Claude Gingras
3,4
, Michael Siu
1
, John McDermott
2
. 1) Department of Chemistry and Centre for Research in Mass Spectrometry, York
University, Toronto, Ontario, Canada; 2) Department of Biology, York University, Toronto, Ontario, Canada; 3) Centre for Systems Biology, Samuel
Lunenfeld Research Institute, Toronto, Ontario, Canada; 4) Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
The p38 MAPK pathway is a key evolutionarily conserved mediator of an organism’s response to stressful environmental stimuli. In mammals four p38
kinases form a robust signaling module believed to be supported by considerable functional redundancy. In
Drosophila
the p38 MAPK family consists of
two highly homologous kinases, Mpk2 and p38b, and the third putative kinase p38c. Recent genetic analyses of various Mpk2 and p38b alleles suggest that
the two kinases are at least partially redundant. However, the extent of this redundancy, and possible non-overlapping roles remain to be defined. To reveal
common and unique molecular functions of individual p38 kinases we generated a high-resolution protein interaction map of Mpk2, p38b, and p38c in S2
cells. The use of an optimized single-step affinity purification procedure followed by gel-free LC-MS/MS analysis allowed us to detect both stable, and more
transient, lower-affinity interactions.
In vitro
kinase reactions with a panel of recombinantly expressed interactors and activated p38 were used to identify
likely kinase substrates. The results of our study suggest limited functional overlap between Mpk2 and p38b, primarily restricted to the regulation of mRNA
processing. Another finding is a previously unacknowledged link between the p38 pathway and the regulation of carbohydrate metabolism. Validation of
these new functional connections in the context of larval muscle and gut will also be presented. More broadly, our results illustrate the use of mass
spectrometry-based interaction mapping for assigning shared and unique molecular functions to individual members of redundant protein families.