Conjugation of ubiquitin-like polypeptide to intracellular acceptor proteins.
Monoclonal nonspecific suppressor factor (MNSF), a lymphokine produced by a murine hybridoma, was originally found to inhibit the generation of LPS-induced immunoglobulin secreting cells. MNSF comprises of MNSF beta, an isoform of MNSF, and the other isoform, MNSF alpha. Ubiquitin-like segment (Ubi-L) of MNSF beta shows MNSF-like activity. Ubi-L (7.8 kDa) has 36% homology with 8.5 kDa ubiquitin. GST-Ubi-L was labeled with 125I by the chloramine T method and tested for its conjugation to acceptor in splenocyte lysates. 125I-GST-Ubi-L conjugation on SDS-PAGE showed heterogeneous bands including 95 kDa GST-Ubi-L conjugation in the splenocyte, but not reticulocyte lysates. The Ubi-L adduct appeared to be MNSF-related molecule because anti-MNSF monoclonal antibody (mAb) recognized the 95 kDa band. The pattern of the conjugations was different from that seen in ubiquitination. Unlabeled GST-Ubi-L inhibited the conjugations, while ubiquitin did not. alpha-Lactalbumin, one of the target proteins for ubiquitination, failed to conjugate to GST-Ubi-L. In addition, covalent conjugation of ubiquitin to reticulocyte lysates was also interfered by GST-Ubi-L. These results suggest that Ubi-L may conjugate to acceptor proteins in a similar, but not in the same way as ubiquitination, and might play an important role in lymphoid cells.
Nagata
T
T
Department of Otolaryngology, Shimane Medical University, Izumo, Japan. tnagata@orange.oon.ne.jp
Nakamura
M
M
Kawauchi
H
H
Tanigawa
Y
Y
D000818:Animals; D003208:Concanavalin A / Q000494:pharmacology; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D008934:Mitogens / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D012156:Reticulocytes / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000166:cytology / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D013491:Suppressor Factors, Immunologic / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*
D008934:Mitogens; D011506:Proteins; D011993:Recombinant Fusion Proteins; D013439:Sulfhydryl Reagents; D013491:Suppressor Factors, Immunologic; D014452:Ubiquitins; C072169:monoclonal nonspecific suppressor factor; D003208:Concanavalin A; D004228:Dithionitrobenzoic Acid; D004229:Dithiothreitol