Approximately 5% of the Arabidopsis (Arabidopsis thaliana) proteome is predicted to be involved in the ubiquitination/26S proteasome pathway. The majority of these predicted proteins have identity to conserved domains found in E3 ligases, of which there are multiple types. The RING-type E3 is characterized by the presence of a cysteine-rich domain that coordinates two zinc atoms. Database searches followed by extensive manual curation identified 469 predicted Arabidopsis RING domaincontaining proteins. In addition to the two canonical RING types (C3H2C3 or C3HC4), additional types of modified RING domains, named RING-v, RING-D, RING-S/T, RING-G, and RING-C2, were identified. The modified RINGs differ in either the spacing between metal ligands or have substitutions at one or more of the metal ligand positions. The majority of the canonical and modified RING domain-containing proteins analyzed were active in in vitro ubiquitination assays, catalyzing polyubiquitination with the E2 AtUBC8. To help identity regions of the proteins that may interact with substrates, domain analyses of the amino acids outside the RING domain classified RING proteins into 30 different groups. Several characterized protein-protein interaction domains were identified, as well as additional conserved domains not described previously. The two largest classes of RING proteins contain either no identifiable domain or a transmembrane domain. The presence of such a large and diverse number of RING domain-containing proteins that function as ubiquitin E3 ligases suggests that targetspecific proteolysis by these E3 ligases is a complex and important part of cellular regulation in Arabidopsis.The covalent attachment of the 76-amino acid protein ubiquitin to target proteins is a central and essential part of numerous cellular processes in eukaryotes. Ubiquitin attachment is a multistep reaction involving three enzymes referred to as E1, E2, and E3 (Glickman and Ciechanover, 2002). The conjugation cascade begins with the formation of thioester-linked ubiquitin by the ubiquitin-activating enzyme or E1 in an ATPdependent reaction. Thioester-linked ubiquitin is then transferred to a cysteinyl residue of an ubiquitinconjugating (UBC) enzyme or E2. Ubiquitin is then transferred from the E2-ubiquitin intermediate to a lysyl group on the substrate. This final step is mediated by the ubiquitin ligase or E3 enzyme. Uniquely, E3 ligases of the HECT domain class form a ubiquitin thioester prior to transfer to the substrate, whereas all the other ligases interact noncovalently with the E2 carrying a thioester-linked ubiquitin. The E3 is also responsible for recruiting the target protein for ubiquitination, and hence is considered the major substrate recognition component of the pathway.Ubiquitin-dependent protein degradation selectively targets a diverse range of substrates, including receptors, nuclear transcription activators and repressors, abnormal proteins, and other short-lived regulatory proteins for degradation by the 26S proteasome (Glickman and Ciec...

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This is the first time that DC have been studied in prostate cancer using the relatively new DC specific monoclonal antibodies CD83 and CMRF-44. These findings suggest that there is no active recruitment of DC into prostate cancer and those DC present are only minimally activated.

The RelB gene product is a member of the nuclear factor (NF)-kappaB family of transcription factors. It has been identified recently within mouse antigen-presenting cells and human monocyte-derived dendritic cells (DC). Disruption of the mouse RelB gene is accompanied, amongst other phenotypes, by abnormalities in the antigen-presenting cell lineages. In order to define RelB expression during human DC differentiation, we have analysed RelB mRNA by reverse transcriptase-polymerase chain reaction and RelB protein by intracellular staining in CD34+ precursors and different types of DC preparations. RelB mRNA was not detected in CD34+ precursor populations. Fresh blood DC (lineage-human leucocyte antigen-DR+ (lin-HLA-DR+)) lacked RelB mRNA and cytoplasmic RelB protein but a period of in vitro culture induced RelB expression in blood DC. Purified Langerhans' cells (LC) (CD1a+ HLA-DR+) failed to express RelB mRNA. Immunocytochemical staining identified RelB protein in human skin epithelium. RelB protein was expressed in a very few CD1a+, CD83+ or CMRF-44+ dermal DC but was not present in CD1a+ LC. Tonsil DC (lin-HLA-DR+ CMRF-44+) were positive for RelB mRNA and RelB protein. Intestinal DC (HLA-DR+) also lacked immunoreactive RelB protein. The majority of interdigitating CD83+, CMRF-44+, CMRF-56+ or p55+ DC located in paracortical T-lymphocyte areas of lymph node and tonsil contained RelB protein. The expression of RelB mRNA and RelB protein correlates with the activated phase of blood DC and the postmigration cell (activated) stage of tissue DC development.

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