Abstract
Activation of the GTPase Rab7/Ypt7 by its cognate guanine nucleotide exchange factor (GEF) Mon1-Ccz1 marks organelles such as endosomes and autophagosomes for fusion with lysosomes/vacuoles and degradation of their content. Here, we present a high-resolution cryogenic electron microscopy structure of the Mon1-Ccz1 complex that reveals its architecture in atomic detail. Mon1 and Ccz1 are arranged side by side in a pseudo-twofold symmetrical heterodimer. The three Longin domains of each Mon1 and Ccz1 are triangularly arranged, providing a strong scaffold for the catalytic center of the GEF. At the opposite side of the Ypt7-binding site, a positively charged and relatively flat patch stretches the Longin domains 2/3 of Mon1 and functions as a phosphatidylinositol phosphate-binding site, explaining how the GEF is targeted to membranes. Our work provides molecular insight into the mechanisms of endosomal Rab activation and serves as a blueprint for understanding the function of members of the Tri Longin domain Rab-GEF family.
Highlights
Activation of the GTPase Rab7/Ypt7 by its cognate guanine nucleotide exchange factor (GEF) Mon1-Ccz1 marks organelles such as endosomes and autophagosomes for fusion with lysosomes/ vacuoles and degradation of their content
We have previously identified the structure of Ypt7 bound to the MC1 core, which comprises a heterodimeric complex of the first Longin domains of Mon1 and Ccz1, respectively [21]
The structure of MC1 reveals that Mon1 and Ccz1 each consist of three Longin domains (LDs), which interact by β-sheet complementation via both LD1s and LD3s
Summary
Activation of the GTPase Rab7/Ypt by its cognate guanine nucleotide exchange factor (GEF) Mon1-Ccz marks organelles such as endosomes and autophagosomes for fusion with lysosomes/ vacuoles and degradation of their content. It has been demonstrated that MC1-dependent recruitment of Ypt to both late endosomes/multivesicular bodies and autophagosomes is required for the fusion of these organelles with the vacuole and degradation of the respective cargo [7, 18] This process is conserved in plants and mammalian cells [9, 14, 19]. The TLD GEF family comprises the universally conserved Rab7-GEF Mon1-Ccz (MC1) [7,8,9] and two other complexes specific to metazoans, namely BLOC-3 (biogenesis of lysosome-related organelles complex-3, which includes Hps and Hps4) and Inturned-Fuzzy. Inturned-Fuzzy, which acts as the GEF of Rab23 [5], has been implicated in the establishment of planar cell polarity and ciliogenesis and was described as part of the Significance
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