Formation of stacked ER cisternae by low affinity protein interactions.
The endoplasmic reticulum (ER) can transform from a network of branching tubules into stacked membrane arrays (termed organized smooth ER [OSER]) in response to elevated levels of specific resident proteins, such as cytochrome b(5). Here, we have tagged OSER-inducing proteins with green fluorescent protein (GFP) to study OSER biogenesis and dynamics in living cells. Overexpression of these proteins induced formation of karmellae, whorls, and crystalloid OSER structures. Photobleaching experiments revealed that OSER-inducing proteins were highly mobile within OSER structures and could exchange between OSER structures and surrounding reticular ER. This indicated that binding interactions between proteins on apposing stacked membranes of OSER structures were not of high affinity. Addition of GFP, which undergoes low affinity, antiparallel dimerization, to the cytoplasmic domains of non-OSER-inducing resident ER proteins was sufficient to induce OSER structures when overexpressed, but addition of a nondimerizing GFP variant was not. These results point to a molecular mechanism for OSER biogenesis that involves weak homotypic interactions between cytoplasmic domains of proteins. This mechanism may underlie the formation of other stacked membrane structures within cells.
Cell Biology and Metabolism Branch, National Institutes of Child Health and Human Development, National Institutes of Health, 18 Library Dr., Bldg. 18T, Rm. 101, Bethesda, MD 20892, USA. jlippin@helix.nih.gov
Erik L
EL
Snapp
Ramanujan S
RS
Hegde
Maura
M
Francolini
Francesca
F
Lombardo
Sara
S
Colombo
Emanuela
E
Pedrazzini
Nica
N
Borgese
Jennifer
J
Lippincott-Schwartz
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure; D018871:Endoplasmic Reticulum, Smooth / Q000378:metabolism / Q000648:ultrastructure; D049452:Green Fluorescent Proteins; D007425:Intracellular Membranes / Q000378:metabolism* / Q000648:ultrastructure; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D008164:Luminescent Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins