Regulation of DENND3, the exchange factor for the small GTPase Rab12 through an intramolecular interaction [Membrane Biology]

The Rab family of small GTPases function in multiple aspects of cellular membrane trafficking. Proteins bearing a DENN (differentially expressed in normal and neoplastic cells) domain have emerged as the largest family of Rab-activating guanine nucleotide exchange factors (GEFs). Rab12 functions in the initiation of starvation-induced autophagy and our previous work revealed that its activator, DENND3 is phosphorylated and activated upon starvation. However, how the GEF activity of DENND3 towards Rab12 is regulated at the molecular level is still not understood. Here we combine size-exclusion chromatography, Forster resonance energy transfer, pull-down and in vitro GEF assays to demonstrate that regulation of GEF activity is achieved through an intramolecular interaction that is controlled by a key residue in DENND3, tyrosine 940. Our study sheds light on the regulation of Rab12 activation, and lays the groundwork for characterizing the regulation of other DENN domain-containing proteins.

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