ER retention may play a role in sorting of the nuclear pore membrane protein POM121.
Integral membrane proteins of the nuclear envelope (NE) are synthesized on the rough endoplasmic reticulum (ER) and following free diffusion in the continuous ER/NE membrane system are targeted to their proper destinations due to interactions of specific domains with other components of the NE. By studying the intracellular distribution and dynamics of a deletion mutant of an integral membrane protein of the nuclear pores, POM121, which lacks the pore-targeting domain, we investigated if ER retention plays a role in sorting of integral membrane proteins to the nuclear envelope. A nascent membrane protein lacking sorting determinants is believed to diffuse laterally in the continuous ER/NE lipid bilayer and expected to follow vesicular traffic to the plasma membrane. The GFP-tagged deletion mutant, POM121(1-129)-GFP, specifically distributed within the ER membrane, but was completely absent from the Golgi compartment and the plasma membrane. Experiments using fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP) demonstrated that despite having very high mobility within the whole ER network (D = 0.41 +/- 0.11 micro m(2)/s) POM121(1-129)-GFP was unable to exit the ER. It was also not detected in post-ER compartments of cells incubated at 15 degrees C. Taken together, these experiments show that amino acids 1-129 of POM121 are able to retain GFP in the ER membrane and suggest that this retention occurs by a direct mechanism rather than by a retrieval mechanism. Our data suggest that ER retention might be important for sorting of POM121 to the nuclear pores.
Imreh
G
G
Södertörns Högskola (University College), S-141 89, Huddinge, Sweden.
Maksel
D
D
de Monvel
J B
JB
Brandén
L
L
Hallberg
Einar
E
D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D019556:COS Cells; D002451:Cell Compartmentation / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure; D005057:Eukaryotic Cells / Q000378:metabolism* / Q000648:ultrastructure; D036681:Fluorescence Recovery After Photobleaching; D006056:Golgi Apparatus / Q000378:metabolism; D049452:Green Fluorescent Proteins; D007425:Intracellular Membranes / Q000378:metabolism; D008164:Luminescent Proteins; D008565:Membrane Proteins / Q000172:deficiency* / Q000235:genetics; D009154:Mutation / Q000235:genetics; D022022:Nuclear Pore / Q000378:metabolism* / Q000648:ultrastructure; D009687:Nuclear Proteins; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics*; D011993:Recombinant Fusion Proteins
D008164:Luminescent Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; C082264:Pom121 protein, rat; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins