JIP4 is recruited by the phosphoinositide-binding protein Phafin2 to promote recycling tubules on macropinosomes

Abstract

Macropinocytosis allows cells to take up extracellular material in a non-selective manner into large vesicles called macropinosomes. After internalization, macropinosomes acquire phosphatidylinositol 3-phosphate (PtdIns3P) on their limiting membrane as they mature into endosomal-like vesicles. The molecular mechanisms that mediate recycling of membranes and transmembrane proteins from these macropinosomes still need to be defined. Here we report that JIP4, a protein previously described to bind to microtubule motors, is recruited to tubulating subdomains on macropinosomes by the PtdIns3P-binding protein Phafin2. These JIP4-positive tubulating subdomains on macropinosomes contain F-actin, the retromer recycling complex, and a retromer cargo, VAMP3. Disruption of the JIP4-Phafin2 interaction, deletion of Phafin2, or inhibition of PtdIns3P production by VPS34 impairs JIP4 recruitment to macropinosomes. While knockout of JIP4 suppresses tubulation, overexpression enhances tubulation from macropinosomes. JIP4 knockout cells display increased retention of macropinocytic cargo in both early and late macropinosomes. Collectively, these data identify JIP4 and Phafin2 as components of a tubular recycling pathway that operates from macropinosomes.

CYRI proteins: controllers of actin dynamics in the cellular ‘eat vs walk’ decision

“…The protein Phafin2 regulates the maturation of macropinosomes by detecting the presence of PtdIns3P and PtdIns4P simultaneously and promoting the reorganisation of the actin cytoskeleton to free the macropinosome from its actin network [ 21 ]. During early engulfment, macropinosomes undergo explosive tubulation, presumably to retrieve membrane and receptors back to the plasma membrane [ 22 ].…”

JNK‐interacting protein 4 is a central molecule for lysosomal retrograde trafficking

“…JIP4 interacts with phafin2 (also known as PLEKHF2), whose FYVE domain binds to a PtdIns(3)P localized in the macropinosome membrane. JIP4 binds to the PH domain of phafin2 via a poorly conserved region at 566‐767 aa, resulting in tubulating subdomains on macropinosomes (Figure 3B) [ 72 ] .…”

Phafins Are More Than Phosphoinositide-Binding Proteins

“…Functionally, both Phafin1 and Phafin2 proteins have been reported to play significant roles in apoptosis and autophagy. Phafin2 promotes macropinocytosis through the rearrangement of the subcortical actin cytoskeleton and recruits JNK (c-Jun NH2-terminal kinase)-interacting protein 4 (JIP4) to tubulating macropinosomes [21,22]; whereas the role of Phafin1 in macropinocytosis remains to be determined.…”

“…Previous studies showed that the Btk PH domain interacts with protein kinase C (PKC) and the Akt PH domain forms a tight complex with calmodulin [78,79]. The Phafin2 PH domain directly binds F-actin and JIP4 (Table 1) [21,22]. The presence of both the PH and FYVE domains in Phafin proteins enables their interactions with membrane-embedded lipids and other peripheral proteins, which characterize them as adaptor proteins; however, it remains unknown whether any of these associations and PIP-binding are mutually exclusive.…”