The synaptojanin-like protein Inp53/Sjl3 functions with clathrin in a yeast TGN-to-endosome pathway distinct from the GGA protein-dependent pathway.
Yeast TGN resident proteins that frequently cycle between the TGN and endosomes are much more slowly transported to the prevacuolar/late endosomal compartment (PVC) than other proteins. However, TGN protein transport to the PVC is accelerated in mutants lacking function of Inp53p. Inp53p contains a SacI polyphosphoinositide phosphatase domain, a 5-phosphatase domain, and a proline-rich domain. Here we show that all three domains are required to mediate "slow delivery" of TGN proteins into the PVC. Although deletion of the proline-rich domain did not affect general membrane association, it caused localization to become less specific. The proline-rich domain was shown to bind to two proteins, including clathrin heavy chain, Chc1p. Unlike chc1 mutants, inp53 mutants do not mislocalize TGN proteins to the cell surface, consistent with the idea that Chc1p and Inp53p act at a common vesicular trafficking step but that Chc1p is used at other steps also. Like mutations in the AP-1 adaptor complex, mutations in INP53 exhibit synthetic growth and transport defects when combined with mutations in the GGA proteins. Taken together with other recent studies, our results suggest that Inp53p and AP-1/clathrin act together in a TGN-to-early endosome pathway distinct from the direct TGN-to-PVC pathway mediated by GGA/clathrin.
Ha
Seon-Ah
SA
Division of Biological Sciences, University of Missouri, Columbia, Missouri 65211, USA.
Torabinejad
Javad
J
DeWald
Daryll B
DB
Wenk
Markus R
MR
Lucast
Louise
L
De Camilli
Pietro
P
Newitt
Richard A
RA
Aebersold
Ruedi
R
Nothwehr
Steven F
SF
D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D033922:Clathrin Heavy Chains / Q000737:chemistry / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism; D005800:Genes, Fungal; D006056:Golgi Apparatus / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism
D033942:Adaptor Proteins, Vesicular Transport; D002352:Carrier Proteins; C407306:GGA adaptor proteins; D009419:Nerve Tissue Proteins; D029701:Saccharomyces cerevisiae Proteins; D018808:Transcription Factor AP-1; D033922:Clathrin Heavy Chains; C110553:INP53 protein, S cerevisiae; C097702:synaptojanin; D010744:Phosphoric Monoester Hydrolases; D020727:ADP-Ribosylation Factors