TY - JOUR T1 - Elucidation of the 14-3-3ζ interactome reveals critical roles of RNA splicing factors during adipogenesis JF - bioRxiv DO - 10.1101/184499 SP - 184499 AU - Yves Mugabo AU - Mina Sadeghi AU - Nancy N. Fang AU - Thibault Mayor AU - Gareth E. Lim Y1 - 2017/01/01 UR - http://biorxiv.org/content/early/2017/09/05/184499.abstract N2 - Adipogenesis is facilitated by a complex signaling network requiring strict temporal and spatial organization of effector molecules. Molecular scaffolds, such as 14-3-3 proteins, coordinate such events, and we have previously identified 14-3-3ζ as an essential scaffold in adipocyte differentiation. The interactome of 14-3-3ζ is large and diverse, and it is possible that novel adipogenic factors may be present within it. Mouse embryonic fibroblasts from mice over-expressing a TAP-epitope-tagged 14-3-3ζ molecule were generated, and following the induction of adipogenesis, TAP-14-3-3ζ complexes were purified, followed by mass spectrometry analysis to determine the 14-3-3ζ interactome. Over 100 proteins were identified as being unique to adipocyte differentiation, of which 56 were novel interacting partners. Previously established regulators of adipogenesis (ie, Ptrf/Cavin1 and Phb2) were found within the 14-3-3ζ interactome, confirming the ability of this approach to identify regulators of adipocyte differentiation. An enrichment of proteins in the interactome related to RNA metabolism, processing, and splicing was identified, and analysis of transcriptomic data revealed that 14-3-3ζ depletion in 3T3-L1 cells affected the alternative splicing of mRNA during adipocyte differentiation. Of the RNA splicing factors within the 14-3-3ζ interactome, depletion of Hnrnpf, Hnrnpk, Ddx6, and Sfpq by siRNA revealed essential roles of these proteins in adipogenesis and their roles in the alternative splicing of Lpin1. In summary, novel adipogenic factors can be detected within the 14-3-3ζ interactome, and further characterization of additional proteins within the 14-3-3ζ interactome has the potential of identifying novel targets to block the expansion of adipose tissue mass that occurs in obesity. ER -