Gene Expression and Synthesis of IL1 p

In human monocytes, IL-1P mRNA levels rise rapidly within 15 min but, depending on the stimulus, start to fall after 4 h. The decrease is thought to be due to the synthesis of a transcriptional repressor and/or a decrease in mRNA half-life (38,39). Using IL-1 itself as a stimulant of its own gene expression, IL-ip mRNA levels are sustained for over 24 h (7,40). Raising cAMP levels in monocytes with histamine enhances IL-ip gene expression and protein synthesis (41). Retinoic acid induces IL-ip gene expression, but the primary precursor transcript fails to yield mature mRNA (39). Inhibition of translation by cycloheximide results in enhanced splicing of exons, excision of introns, and increased levels of mature mRNA (superinduction). Thus, synthesis of mature IL-ip mRNA requires an activation step to overcome an apparently intrinsic inhibition to process precursor mRNA.

A dissociation between transcription and translation is characteristic of IL-ip and also of TNF-a (42). Despite a vigorous signal for transcription by a variety of agents, including C5a, adherence, or even hyperosmolar NaCl (42-44), most of the IL-ip mRNA is degraded and no significant translation into proIL-ip takes place. Although the IL-ip mRNA assembles into large polyribosomes, there is little significant elongation of the peptide (45). However, adding bacterial endotoxin or IL-i itself to cells with high levels of steady-state IL-ip mRNA results in augmented translation (42,44) in somewhat the same manner as the removal of cycloheximide following superinduction. One explanation is that stabilization of the AU-rich 3' untranslated region takes place in cells stimulated with lipopolysaccharide (LPS). These AU-rich sequences are known to suppress normal hemoglobin synthesis. The stabilization of mRNA by microbial products may explain why low concentrations of LPS or a few bacteria or Borrelia organisms per cell induce the translation of large amounts of IL-ip (46).

Another explanation is that IL-i stabilizes its own mRNA by preventing deadenylation as it does for the chemokine gro-a (47). Removal of IL-i from cells after 2 h increases the shortening of polyadenylic acid (poly-A), and IL-i apparently is an important regulator of gro synthesis, because it prevents deadenylation. In fact, of the several cytokines induced by IL-i, large amounts of the chemokine family are produced in response to low concentrations of IL-i. For example, i pM of IL-i stimulates fibroblasts to synthesize i0 nM of IL-8 (48).

Following synthesis, proIL-ip remains primarily cytosolic until it is cleaved and transported out of the cell (Fig. 2). The IL-ip propiece (amino acids i-ii6) is also myristoylated on lysine residues (34), but unlike IL-ia, proIL-ip has no known membrane form and proIL-ip is only marginally active (49). Some IL-ip is found in lysosomes (50) or associated with microtubules (5i,52) and either localization may play a role in the secretion of IL-ip. In mononuclear phagocytes, a small amount of proIL-ip is secreted from

Figure 2 Human blood monocyte producing IL-ip. mRNA coding for proIL-ip is translated on polysomes in the cytosol and associated with microtubules (52). proIL-ip remains cytosolic until cleaved by ICE. ICE is translated in the endoplasmic reticulum as an inactive precursor (proICE) and requires two internal cleavage steps to form the enzymatically active heterodimer. Two heterodimers form a tetramer in association with two molecules of proIL-ip and cleavage occurs. Active ICE is found predominantly on the inner surface of the cell membrane (68). Following cleavage, 17-kD IL-1P is secreted into the extracellular compartment through a putative membrane channel. The 16 kD IL-ip ''propiece'' can be found both inside and outside the cell. A small amount of proIL-i p can be transported into the extracellular space from intact cells, presumably using the same channel; however, when ICE activity is inhibited, more proIL-ip is found in the extracellular compartment (65).

Figure 2 Human blood monocyte producing IL-ip. mRNA coding for proIL-ip is translated on polysomes in the cytosol and associated with microtubules (52). proIL-ip remains cytosolic until cleaved by ICE. ICE is translated in the endoplasmic reticulum as an inactive precursor (proICE) and requires two internal cleavage steps to form the enzymatically active heterodimer. Two heterodimers form a tetramer in association with two molecules of proIL-ip and cleavage occurs. Active ICE is found predominantly on the inner surface of the cell membrane (68). Following cleavage, 17-kD IL-1P is secreted into the extracellular compartment through a putative membrane channel. The 16 kD IL-ip ''propiece'' can be found both inside and outside the cell. A small amount of proIL-i p can be transported into the extracellular space from intact cells, presumably using the same channel; however, when ICE activity is inhibited, more proIL-ip is found in the extracellular compartment (65).

intact cells (53,54), but the pathway for this secretion remains unknown. On the other hand, release of mature IL-ip appears to be linked to processing at the aspartic acid-alanine (ii6-ii7) peptide cleavage by the IL-ip-con-verting enzyme (ICE) (55) (see below).

There are several sites in proIL-ip which are vulnerable to cleavage by enzymes in the vicinity of alanine ii7. These are trypsin, elastase, chymotryp-sin, a mast cell chymase, and a variety of proteases which are commonly found in inflammatory fluids (reviewed in ref. 56). The extent that these proteases play in the in vivo conversion of proIL-ip to mature forms is uncertain, but in each case, a biologically active IL-ip species is produced. For example, in ICE-deficient mice given a subcutaneous injection of turpentine, biologically active IL-ip is released (56). In the discussion on the soluble IL-i receptor type II (below), the affinity of proIL-ip for this constitutively produced soluble receptor is high and may prevent haphazard cleavage of the precursor by these enzymes in inflammatory fluids.

0 0

Post a comment