Expression of IL-1 genes in human and bovine chondrocytes: a mechanism for autocrine control of cartilage matrix degradation

In this report we describe the presence of interleukin-1 activity in medium conditioned by bovine articular cartilage. Preparations partially purified by Sephacryl S200 chromatography (Mr 18000-25000) stimulate murine thymocyte proliferation in the lymphocyte activation factor assay. Furthermore, the factor(s) activate cartilage tissue to secrete a protease which is essential for the activity of purified synovial collagenase. We also demonstrate the presence of mRNA coding for IL-1 alpha and beta in human articular chondrocytes and conclude that the human monocytic and chondrocytic mRNAs are identical. Our results demonstrating cartilage expression of IL-1 genes suggest the possibility of an autocrine mechanism whereby chondrocyte production of matrix degrading proteases is initiated by chondrocyte derived IL-1.     

Hedgehog secretion and signal transduction in vertebrates

Signaling by the Hedgehog (Hh) family of secreted proteins is essential for proper embryonic patterning and development. Dysregulation of Hh signaling is associated with a variety of human diseases ranging from developmental disorders such as holoprosencephaly to certain forms of cancer, including medulloblastoma and basal cell carcinoma. Genetic studies in flies and mice have shaped our understanding of Hh signaling and revealed that nearly all core components of the pathway are highly conserved. Although many aspects of the Drosophila Hh pathway are conserved in vertebrates, mechanistic differences between the two species have begun to emerge. Perhaps the most striking divergence in vertebrate Hh signaling is its dependence on the primary cilium, a vestigial organelle that is largely absent in flies. This minireview will provide an overview of Hh signaling and present recent insights into vertebrate Hh secretion, receptor binding, and signal transduction.     

The Mechanism of Pre-transfer Editing in Yeast Mitochondrial Threonyl-tRNA Synthetase

Accurate translation of mRNA into protein is a fundamental biological process critical for maintaining normal cellular functions. To ensure translational fidelity, aminoacyl-tRNA synthetases (aaRSs) employ pre-transfer and post-transfer editing activities to hydrolyze misactivated and mischarged amino acids, respectively. Whereas post-transfer editing, which requires either a specialized domain in aaRS or a trans-protein factor, is well described, the mechanism of pre-transfer editing is less understood. Here, we show that yeast mitochondrial threonyl-tRNA synthetase (MST1), which lacks an editing domain, utilizes pre-transfer editing to discriminate against serine. MST1 misactivates serine and edits seryl adenylate (Ser-AMP) in a tRNA-independent manner. MST1 hydrolyzes 80% of misactivated Ser-AMP at a rate 4-fold higher than that for the cognate threonyl adenylate (Thr-AMP) while releasing 20% of Ser-AMP into the solution. To understand the mechanism of pre-transfer editing, we solved the crystal structure of MST1 complexed with an analog of Ser-AMP. The binding of the Ser-AMP analog to MST1 induces conformational changes in the aminoacylation active site, and it positions a potential hydrolytic water molecule more favorably for nucleophilic attack. In addition, inhibition results reveal that the Ser-AMP analog binds the active site 100-fold less tightly than the Thr-AMP analog. In conclusion, we propose that the plasticity of the aminoacylation site in MST1 allows binding of Ser-AMP and the appropriate positioning of the hydrolytic water molecule.     

Differential effects of hyperosmotic challenge on interleukin-1-activated pathways in bovine articular cartilage

Chondrocytes in situ experience fluctuations in extracellular osmolarity resulting from mechanical loading. The objective of this study was to determine whether hyperosmotic stress causes or exacerbates interleukin-1 (IL-1)-mediated effects in bovine articular cartilage. Disks of cartilage cut from the articular surface of calf radiocarpal joints were incubated for 24h in the presence or absence of IL-1 in Dulbecco's modified Eagle's medium adjusted to various osmolalities with sucrose or NaCl. Cyclooxygenase (COX)-2 levels in the cartilage were examined by Western blot. Culture media were assayed for prostaglandin E(2) (PGE(2)), nitrite as an indicator of nitric oxide (NO) production, and sulfated glycosaminoglycan as an indicator of proteoglycan degradation. We report the osmolality-dependent potentiation of COX-2 and PGE(2) production, and the osmolality-dependent inhibition of NO production and proteoglycan degradation in IL-1-activated cartilage. The data demonstrate that osmotic and cytokine signaling interact to differentially modulate IL-1-stimulated effects in calf articular cartilage.