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Interleukin 1: The patterns of translation and intracellular distribution support alternative secretory mechanisms
Author(s) -
Stevenson Frazier T.,
Torrano Frank,
Locksley Richard M.,
Lovett David H.
Publication year - 1992
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041520202
Subject(s) - polysome , intracellular , cytoskeleton , biology , microbiology and biotechnology , immunoelectron microscopy , biochemistry , ribosome , cell , rna , antibody , gene , immunology
Interleukin‐1 (IL‐1) is synthesized as a 31 kDa precursor protein, whose multiple extracellular activities are attributed to receptor binding of a processed, carboxyterminal 17 kDa peptide. Unlike other secreted proteins, the IL‐1 precursor lacks a hydrophobic leader sequence and is not found in organelles composing the classical secretory pathway. In order to further clarify the intracellular processing of IL‐1, we studied its site of synthesis in human monocytes. Secreted and integral membrane proteins are translated on membrane‐bound polyribosomes, while intracellular proteins are translated on free polyribosomes. Free and membrane‐bound polysomes were isolated from Lipid A‐stimulated monocyte lysates and immunoblotted using antibodies specific to the N‐terminal regions of the IL‐1α and β precursors. Free polysome fractions showed multiple small bands consistent with nascent peptide chains; membrane‐bound polysomes yielded no detectable IL‐1. Polysome fractions were then analyzed by immunoelectron microscopy; nascent IL‐1α and β peptide chains were readily seen emerging from cytoskeletal‐associated free polyribosomes, but not membrane‐bound polyribosomes. Electron microscopic in situ hybridization revealed IL‐1 mRNA chains attached to cytoskeletal‐associated free, but not membrane‐bound polyribosomes. The intracellular distribution of the fully synthesized IL‐1β precursor was studied in human mesangial cells (HMC), whose cytoskeletal organization is more readily evaluated than that of monocytes. Dual immunofluorescence microscopy of these cells revealed a complex intracellular distribution of the fully synthesized 31 kDa IL‐1 precursors. IL‐1 was asymmetrically distributed between cytosolic, microtubule, and nuclear compartments, without association with actin or intermediate filaments. This demonstration of the sites of IL‐1 synthesis and patterns of intracellular distribution provide further evidence for an extracellular release mechanism which is clearly distinct from the classical secretory pathway. © 1992 Wiley‐Liss, Inc.