Electron microscopy and subunit-subunit interaction studies reveal a firstarchitecture of COP9 signalosome

The COP9 signalosome is involved in signal transduction, whereas the 26 S p roteasome Lid is a regulatory subcomplex of the 26 S proteasome responsible for degradation of ubiquitinated proteins. COP9 signalosome and Lid posses s significant sequence homologies among their eight core subunits and are l ikely derived from a common ancestor. Surprisingly, from our two-dimensiona l electron microscopy data, a common architectural plan for the two complex es could not be deduced. None-the-less, the two particles have structural f eatures in common. Both COP9 signalosome and lid lack any symmetry in subun it arrangement and exhibit a central groove, possibly qualified for scaffol ding functions. Filter-binding assays with recombinant COP9 signalosome components revealed a multitude of subunit-subunit interactions, supporting the asymmetrical a ppearance of the complex in electron microscopy. On the basis of two-dimens ional images and subunit interaction studies, a first architectural model o f COP9 signalosome was created. The fact that four distinct classes of particle views were identified and t hat only 50% of the selected particles could be classified indicates a high degree of heterogeneity in electron microscopic images. Different orientat ions with respect to the viewing axis and conformational variety, presumabl y due to different grades of phosphorylation, are possible reasons for the heterogeneous appearance of the complex. Our biochemical data show that rec ombinant COP9 signalosome subunits 2 and 7 are phosphorylated by the associ ated kinase activity. The modification of COP9 signalosome subunit 2 might be essential for c-Jun phosphorylation. Dephosphorylation does not inactiva te the associated kinase activity. Although substrate phosphorylation by CO P9 signalosome is significantly decreased by lambda protein phosphatase tre atment, "autophosphorylation" is increased. (C) 2000 Academic Press.