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.

Formation of stacked ER cisternae by low affinity protein interactions / E.L. Snapp, R.S. Hegde, M. Francolini, F. Lombardo, S. Colombo, E. Pedrazzini, N. Borgese, J. Lippincott Schwartz. - In: THE JOURNAL OF CELL BIOLOGY. - ISSN 0021-9525. - 163:2(2003 Oct 27), pp. 257-269. [10.1083/jcb.200306020]

Formation of stacked ER cisternae by low affinity protein interactions

M. Francolini;S. Colombo;
2003

Abstract

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.
Animals ; COS Cells ; Endoplasmic Reticulum ; Green Fluorescent Proteins ; Intracellular Membranes ; Recombinant Fusion Proteins ; Membrane Proteins ; Models, Biological ; Luminescent Proteins ; Cercopithecus aethiops ; Point Mutation ; Protein Structure, Tertiary ; Endoplasmic Reticulum, Smooth ; Cell Line
Settore BIO/14 - Farmacologia
27-ott-2003
Article (author)
File in questo prodotto:
File Dimensione Formato  
200306020.pdf

accesso aperto

Tipologia: Post-print, accepted manuscript ecc. (versione accettata dall'editore)
Dimensione 977.99 kB
Formato Adobe PDF
977.99 kB Adobe PDF Visualizza/Apri
Pubblicazioni consigliate

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/161950
Citazioni
  • ???jsp.display-item.citation.pmc??? 204
  • Scopus 377
  • ???jsp.display-item.citation.isi??? 355
social impact