Protein phosphatase 2Cepsilon is an endoplasmic reticulum integral membrane protein that dephosphorylates the ceramide transport protein CERT to enhance its association with organelle membranes.
Protein phosphatase 2Cepsilon (PP2Cepsilon), a mammalian PP2C family member, is expressed in various tissues and is implicated in the negative regulation of stress-activated protein kinase pathways. We show that PP2Cepsilon is an endoplasmic reticulum (ER) transmembrane protein with a transmembrane domain at the amino terminus and the catalytic domain facing the cytoplasm. Yeast two-hybrid screening of a human brain library using PP2Cepsilon as bait resulted in the isolation of a cDNA that encoded vesicle-associated membrane protein-associated protein A (VAPA). VAPA is an ER resident integral membrane protein involved in recruiting lipid-binding proteins such as the ceramide transport protein CERT to the ER membrane. Expression of PP2Cepsilon resulted in dephosphorylation of CERT in a VAPA expression-dependent manner, which was accompanied by redistribution of CERT from the cytoplasm to the Golgi apparatus. The expression of PP2Cepsilon also enhanced the association between CERT and VAPA. In addition, knockdown of PP2Cepsilon expression by short interference RNA attenuated the interaction between CERT and VAPA and the sphingomyelin synthesis. These results suggest that CERT is a physiological substrate of PP2Cepsilon and that dephosphorylation of CERT by PP2Cepsilon may play an important role in the regulation of ceramide trafficking from the ER to the Golgi apparatus.
Saito
Satoko
S
Department of Biochemistry, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryo-machi, Aoba-ku, Sendai 980-8575, Japan.
Matsui
Hiroyuki
H
Kawano
Miyuki
M
Kumagai
Keigo
K
Tomishige
Nario
N
Hanada
Kentaro
K
Echigo
Seishi
S
Tamura
Shinri
S
Kobayashi
Takayasu
T
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D001921:Brain / Q000201:enzymology*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002518:Ceramides / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000201:enzymology* / Q000235:genetics; D006056:Golgi Apparatus / Q000201:enzymology* / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000071636:Protein Phosphatase 2C; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D013109:Sphingomyelins / Q000096:biosynthesis / Q000235:genetics; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism
D002352:Carrier Proteins; D002518:Ceramides; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D013109:Sphingomyelins; C114051:VAPA protein, human; C491920:Vapa protein, mouse; D033921:Vesicular Transport Proteins; C119090:CERT1 protein, human; D017346:Protein Serine-Threonine Kinases; D010749:Phosphoprotein Phosphatases; D000071636:Protein Phosphatase 2C