Specificity, promiscuity and localization of ARF protein interactions with NCS-1 and phosphatidylinositol-4 kinase-III beta.
ADP-ribosylation factor (ARF) proteins are involved in multiple intracellular vesicular transport pathways. Most studies have focused on the functions of ARF1 or ARF6 and little is known about the remaining ARF isoforms. Although the mammalian ARF proteins share a high degree of sequence identity, recent evidence has indicated that they may control distinct trafficking steps within cells. A unanswered issue is the degree of specificity of ARF family members for different interacting proteins. To investigate potential functional differences between the human ARF proteins, we have examined the localization of all human ARF isoforms and their interactions with two ARF1 binding proteins, neuronal calcium sensor-1 (NCS-1) and phosphatidylinositol-4 kinase-IIIbeta (PI4Kbeta). Use of a fluorescent protein fragment complementation method showed direct interactions between ARFs 1, 3, 5 and 6 with NCS-1 but at different intracellular locations in live HeLa cells. Photobleaching experiments indicated that complementation did not detect dynamic changes in protein interactions over short-time scales. A more specific interaction between ARFs 1/3 and PI4Kbeta was observed. Consistent with these latter findings ARF1 but not ARF5 or 6 enhanced the stimulatory effect of PI4Kbeta on regulated exocytosis, suggesting a specific role for class-I ARFs in the regulation of PI4Kbeta.
Haynes
Lee P
LP
The Physiological Laboratory, School of Biomedical Sciences, University of Liverpool, Liverpool, L69 3BX, UK.
Sherwood
Mark W
MW
Dolman
Nick J
NJ
Burgoyne
Robert D
RD
D019870:1-Phosphatidylinositol 4-Kinase / Q000378:metabolism*; D020727:ADP-Ribosylation Factors / Q000502:physiology*; D000818:Animals; D005089:Exocytosis / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D051580:Neuronal Calcium-Sensor Proteins / Q000378:metabolism*; D009479:Neuropeptides / Q000378:metabolism*; D016716:PC12 Cells; D025941:Protein Interaction Mapping; D020033:Protein Isoforms / Q000502:physiology; D051381:Rats
D051580:Neuronal Calcium-Sensor Proteins; D009479:Neuropeptides; D020033:Protein Isoforms; C101464:frequenin calcium sensor proteins; D019870:1-Phosphatidylinositol 4-Kinase; D020727:ADP-Ribosylation Factors