Analysis of the dual function of the ESCRT-III protein Snf7 in endocytic trafficking and in gene expression

Peter Weiss , Stefanie Huppert , Ralf Kölling (1)

1 Lebensmittelwissenschaft und Biotechnologie

ESCRT-III mediates budding and scission of intralumenal vesicles into multivesicular endosomes. For the main ESCRT-III subunit Snf7, an additional role in activation of the transcription factor Rim101 ("Rim pathway") is now firmly established. Here, we investigate how the two Snf7 functions are related to each other. By generating SNF7 mutations that severely affect endocytic trafficking, but leave the Rim pathway function intact, we show that the two functions of SNF7 can be separated genetically. We analyzed in detail, how the SNF7 mutations affect the interaction of Snf7 with its various binding partners. While interactions with Rim-related functions (Rim13, Rim20) were not altered by the mutations, there was a strong effect on interactions with components of the ESCRT pathway. The interaction with the ESCRT-III subunits Vps20 and Vps24 was strongly increased by the mutations, while the interaction with functions acting downstream of ESCRT-III (Vps4 and Bro1) was reduced. Since Vps4 is required for disassembly of ESCRT-III, these data suggest that ESCRT-III is more stable in our SNF7 mutants. In line with this notion, a higher fraction of mutant Snf7 was detected at the membrane. Upon shift to alkaline pH, a stronger binding signal for virtually all interaction partners (except for Vps4) was observed. This indicates that the ESCRT network at the endosomal membrane is more extensive under these conditions.

Mots clés: Life Sciences

Peter Weiss, Stefanie Huppert, Ralf Kölling. Analysis of the dual function of the ESCRT-III protein Snf7 in endocytic trafficking and in gene expression. Biochemical Journal, 2009, 424 (1), pp.89-97. ⟨10.1042/BJ20090957⟩. ⟨hal-00479213⟩